Anesthesia, brain changes, and behavior: Insights from neural systems biology.

Progress in Neurobiology 153 (2017) 121–160

Contents lists available at ScienceDirect

Progress in Neurobiology journal homepage: www.elsevier.com/locate/pneurobio

Review article

Anesthesia, brain changes, and behavior: Insights from neural systems biology Elisabeth Colona,b,* , Edward A. Bittnerc , Barry Kussmanb , Mary Ellen McCannb , Sulpicio Sorianob , David Borsooka,b a b c

Center for Pain and the Brain, 1 Autumn Street, Boston Children’s Hospital, Boston MA 02115, United States Department of Anesthesia, Perioperative, and Pain Medicine, Boston Children’s Hospital, Harvard Medical School, Boston, MA 02115, United States Department of Anesthesia, Critical Care and Pain Medicine, Massachusetts General Hospital, Harvard Medical School, Boston, MA 02114, United States

A R T I C L E I N F O

Article history: Received 15 September 2016 Received in revised form 19 January 2017 Accepted 22 January 2017 Available online 8 February 2017 Keywords: Anesthetics Toxicity Neurocognitive outcome Functional imaging Apoptosis Aging and developing brain

A B S T R A C T

Long-term consequences of anesthetic exposure in humans are not well understood. It is possible that alterations in brain function occur beyond the initial anesthetic administration. Research in children and adults has reported cognitive and/or behavioral changes after surgery and general anesthesia that may be short lived in some patients, while in others, such changes may persist. The changes observed in humans are corroborated by a large body of evidence from animal studies that support a role for alterations in neuronal survival (neuroapoptosis) or structure (altered dendritic and glial morphology) and later behavioral deficits at older age after exposure to various anesthetic agents during fetal or early life. The potential of anesthetics to induce long-term alterations in brain function, particularly in vulnerable populations, warrants investigation. In this review, we critically evaluate the available preclinical and clinical data on the developing and aging brain, and in known vulnerable populations to provide insights into potential changes that may affect the general population of patients in a more, subtle manner. In addition this review summarizes underlying processes of how general anesthetics produce changes in the brain at the cellular and systems level and the current understanding underlying mechanisms of anesthetics agents on brain systems. Finally, we present how neuroimaging techniques currently emerge as promising approaches to evaluate and define changes in brain function resulting from anesthesia, both in the short and the long-term. © 2017 Elsevier Ltd. All rights reserved.

Contents 1. 2. 3.

Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . Scope of the problem: a cause for alarm? . . . . . . . . . . . . . . . . . . . . . . Anesthetic effects on neuronal, glial, dendritic, and synaptic function Neuronal effects . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3.1. Developing brain . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3.1.1. Aging brain . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3.1.2.

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Abbreviation: AD, Alzheimer Disease; ASD, Autism Spectrum Disorder; AV, anterodorsal; BIS, Bispectral Index; BM, basomedial; CBF, Cerebral blood flow; CFA, Complete Freund’s adjuvant; CSI, Cerebral state index; CVA, cerebral vascular accident; D, day; DGC, Dentate granule cell; DL, dorsolateral; DTI, Diffusion Tensor Imaging; EEG, Electroencephalogram; fMRI, functional Magnetic Resonance Imaging; fNRIS, functional Near-infrared Spectroscopy; GA, General anesthesia; GABA, y-aminobutyrate; GMR, Rate of glucose consumption; HbO, oxygenated hemoglobin; HbR, de-oxygenated hemoglobin; HbT, total hemoglobin; ICD, International Classification of Disease; ICU, intensive care unit; IQ, Intelligence quotient; IL, Interleukin; Kg, kilogram; LD, laterodorsal; MAC, Minimum Alveolar Concentration; MD, mediodorsal; Mg, milligram; MRI, Magnetic Resonance Imaging; MRS, Magnetic Resonance Spectroscopy; NAA, N-acetylasparate; NIRS, Near-infrared Spectroscopy; NMDA, N-methyl-D-aspartate; PET, Positron Emission Tomography; phMRI, pharmacological Magnetic Resonance Imaging; PND, postnatal day; POCD, Postoperative Cognitive Dysfunction; POD, Postoperative Delirium; RA, Regional anesthesia; rCBF, regional cerebral blood flow; TNF-a, Tumor necrosis factor-a; VM, ventromedial; Y, year; 1HMRS, proton Magnetic Resonance Spectroscopy. * Corresponding author at: Center for Pain and the Brain, 1 Autumn Street, Boston Children’s Hospital, Boston MA 02115, United States. E-mail address: [email protected] (E. Colon). http://dx.doi.org/10.1016/j.pneurobio.2017.01.005 0301-0082/© 2017 Elsevier Ltd. All rights reserved.

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E. Colon et al. / Progress in Neurobiology 153 (2017) 121–160

3.1.3. Neurogenesis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . Anesthetic agents and glial effects . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . Developing brain . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3.2.1. Aging brain . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3.2.2. Dendritic effects and synaptic connectivity effects . . . . . . . . . . . . . . . . . . . . . . . . . . . 3.3. Developing brain . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3.3.1. Aging brain . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3.3.2. Secondary processes in anesthesia as contributors to altered brain function . . . . . . 3.4. 3.4.1. Surgical stress response . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . Neuroinflammation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3.4.2. Pain . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3.4.3. 3.4.4. Opioids . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . Neuroprotective factors . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3.5. Anesthetic effects across different age groups . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . Anesthesia in children: preclinical and clinical evidence of altered brain biology . . 4.1. 4.1.1. Behavioral changes in animal models . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . Behavioral changes in children . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 4.1.2. Evaluating the pediatric data: potential confounding factors . . . . . . . . . . . 4.1.3. Anesthesia in the elderly: preclinical and clinical evidence of altered brain biology 4.2. 4.2.1. Behavioral changes in animal models . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . Behavioral changes in the elderly . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 4.2.2. Evaluating the data: potential confounding factors . . . . . . . . . . . . . . . . . . . 4.2.3. Brain measures: insights into anesthetic effects on brain structure and function . . . . . . . . 5.1. Positron emission tomography (PET) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . Functional magnetic resonance imaging (fMRI) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5.2. Short-term changes . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5.2.1. Long-term changes . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5.2.2. Functional near-infrared spectroscopy (fNIRS) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5.3. Short-term changes . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5.3.1. Long-term changes . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5.3.2. Electrophysiological methods (EEG) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5.4. Short-term changes . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5.4.1. Long-term changes . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5.4.2. Conclusion . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . Acknowledgements . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3.2.

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1. Introduction Do anesthetics produce long-term alterations in brain function? From the beginning of anesthetic practice until recently, the actions of anesthetics on the brain were considered to have shortterm consequences with a complete return to the initial state upon drug elimination, or in other words, the actions were completely reversible. However, as with many drugs that act on the central nervous system, there is now accumulating evidence that alterations in brain function occur beyond the initial anesthetic administration (Hudson and Hemmings, 2011). Longer-term alterations in brain function, and consequently behavior, may occur following general anesthesia. Infants, young children and the elderly are particularly vulnerable populations (Hudson and Hemmings, 2011). In children, behavioral changes may include night terrors, increased anxiety, bedwetting, altered cognition, and altered sleep-wake cycles (Keaney et al., 2004; Perouansky and Hemmings, 2009). Moreover, although controversial, some studies have found in children an association between exposure to general anesthesia early in life and adverse long-term neurodevelopmental outcomes (for review see: Brambrink et al., 2012b; Lei et al., 2014; Lin et al., 2014; Loepke and Soriano, 2012). In adults, particularly the elderly, a state of confusion, memory loss, and alterations in executive functions are common days after surgery/anesthesia and may persist for months (Monk and Price, 2011; Silverstein and Deiner, 2013). Such postoperative behavioral disturbances have led to the concern that general anesthesia may induce long-term alterations in brain function, which may be more pronounced in vulnerable populations (Boxs 1 and 2, ). Neuronal cell death, morphological changes and neurocognitive

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impairments after general anesthesia have been unequivocally demonstrated in laboratory animal models (Vutskits and Xie, 2016). This public health concern has prompted the Food and Drug Administration to issue a Drug Safety Communication “warning that repeated or lengthy use of general anesthetic and sedation drugs during surgeries or procedures in children younger than 3 years or in pregnant women during their third trimester may affect the development of children’s brains” (Food and Drug Administration, 2016). Preclinical animal studies have indicated that anesthetic agents can be neurotoxic by inducing central nervous system (CNS) changes at both the cellular and systemic levels in the juvenile and adult animal brain. Neurotoxicity is defined as structural or functional alteration in the nervous system resulting from exposure to a chemical, biological, or physical agent (Bittner et al., 2011). Alterations in dendritic complexity (sprouting or pruning) is a newer concept of the effects of anesthetics on brain structure and function, and is analogous to how antidepressants may alter neural connectivity (Castrén and Hen, 2013; Hudetz, 2012). Such alterations would form a basis of altered synaptic and therefore circuit function (Meyer, 2015). Recently, advances in brain neuroimaging have provided an opportunity to evaluate and define changes in brain function resulting from anesthesia. In this article, we review the current evidence for the effects of exposure to general anesthesia on the developing and adult/aging brain in animals and humans. First, we briefly present the scope of the problem and the current debate in the literature regarding the accumulating experimental evidence for an association between exposure to general anesthesia and long-term deleterious effects on the CNS. In the section that follows, we review Anesthetic effects


on neuronal, glial, dendritic, and synaptic function, and the current understanding underlying mechanisms of anesthetic agents on brain systems. In the section Anesthetic effects across age, we attempt to critically evaluate the available preclinical and clinical data on brain systems in two groups children and adults. We also present the potential confounding factors in the clinical studies that have evaluated the long-term effect of general anesthesia on brain functions. In the last section on Brain measures: insights into anesthetic effects on brain structure and function, we present the recent advances in brain neuroimaging that provide an opportunity to evaluate how anesthetic drugs act on the brain to alter level of consciousness and cognition at the short and long-term level. 2. Scope of the problem: a cause for alarm? The use of anesthesia to prevent pain and produce unconsciousness during surgery has been a major advance in medicine (Mashour et al., 2005). While the mechanism of action of many anesthetics is still not completely understood, the potential for these agents to induce longer-term alterations in brain function, particularly in vulnerable populations, warrants evaluation. Vulnerable populations, aside from the very young (Loepke and Soriano, 2008) and the elderly (Monk and Price, 2011), include those with deficits in brain function (e.g., autism, dementia) undergoing surgery and general anesthesia (Seitz et al., 2013; Short and Calder, 2013; Silverstein and Deiner, 2013). Although controversial, the changes observed in humans after general anesthesia are corroborated by a large body of evidence in animal studies demonstrating that the young and old brain of a large variety of species are affected by almost all the anesthetic agents used in clinical practice, with persistent deficits in learning and memory (Brambrink et al., 2012b; Loepke and Soriano, 2008).

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Research in adults has identified relatively strong evidence of cognitive and/or behavioral changes after surgery and general anesthetics, with persistence in some patients (Bittner et al., 2011; Crosby and Culley, 2011; Monk and Price, 2011). Postoperative cognitive changes categorized as delirium or postoperative cognitive dysfunction (POCD) have a higher prevalence in the elderly (>65 years) than in younger patients (Bittner et al., 2011; Monk and Price, 2011). Preclinical data suggest also a potential role of general anesthesia in the promotion of neurodegenerative diseases (Bittner et al., 2011; Seitz et al., 2013), although no strong evidence has been found in humans to date (Vanderweyde et al., 2010). In children, multiple general anesthesia may increase the risk for development of behavioral and emotional disturbances (Bakri et al., 2015; DiMaggio et al., 2011; Sprung et al., 2012; Stargatt et al., 2006). A recent meta-analysis suggested that multiple exposures to general anesthesia before the age of 4 years may be linked to neurodevelopmental deficits later in life (Wang et al., 2014). Although general anesthesia does not appear to lead to more cognitive or behavioral problems in patients with autistic spectrum disorders (ASD) than in other children, for some autistic patients an unpredictable regression in skill and behavior has been described (Short and Calder, 2013). Children with ASD and biochemical and metabolic abnormalities might be at higher risk for behavioral changes following general anesthesia (Short and Calder, 2013) (Box 1). Despite investigations in humans, a causal link between general anesthesia/surgery and long-term behavioral and/or cognitive alterations in children and the elderly is not yet established and is still a matter of debate in the literature (Avidan and Evers, 2016). Nevertheless, the current data support the notion that anesthetic agents might alter normal brain function in individuals, and that the effects may be short lived in some patients but persist in others.

Box 1. Anesthesia and Autism – The Sensitive Brain Patient Group ASD are a group of complex disorders of brain development (Jeste and Geschwind, 2014). Patients with ASD are particularly vulnerable to cognitive and behavioral dysfunction following minor stressors, including anesthesia, and therefore the disease may provide a useful model to evaluate the effects of anesthetics on brain function. The prevalence of childhood autism is reportedly 1 in 88 children (http://www.cdc.gov/ncbddd/autism/data.html), and occurs in all racial, ethnic, and socioeconomic groups. Brain Changes in Autism Patients with ASD have well described structural abnormalities in the brain. The most common include abnormalities in the prefrontal cortex, amygdala, basal ganglia, and reward processing areas. Functional imaging studies have reported alterations in brain structure (Chen et al., 2011; Ecker et al., 2015; Ha et al., 2015) and brain functional connectivity (Assaf et al., 2010; Gotts et al., 2012). Furthermore, studies report alterations in brain mitochondrial function (which play a role in apoptosis) in ASD, with some regions (viz., cerebellum and the frontal and temporal regions) seemingly more affected than others (Chauhan et al., 2011). Anesthesia and Autism Risk An increased risk of developing ASD following early exposure to anesthetic agents has been suggested. An old study in 1991, reported significantly more cases of ASD (0.2%) in a group of children exposed to anesthetics during delivery as compared to a group of children not exposed during delivery (0.09%) (Hattori et al., 1991). More recently, a higher incidence of ASD has been noticed in neonates delivered by cesarean section with general anesthesia in comparison to neonates delivered vaginally (Chien et al., 2015). Curran et al. reported that children born by cesarean section are 20% more likely to be diagnosed with ASD later in life. However, when compared to sibling controls, the association was no longer significant, indicating potential confounding genetic and/or environmental factors (Curran et al., 2015). A recent retrospective matched-cohort study found no difference in the incidence of ASD between children exposed to general anesthesia before the age of 2 years and non-exposed matched-controls (Ko et al., 2015). This is in agreement with the results of a meta-analysis which concluded that there is no evidence for an association between anesthesia and risk of development of ASD (Gardener et al., 2011). Post Anesthesia Behavioral Changes in ASD ASD patients have challenging behavioral problems that may be exacerbated by stressors, including altered sleep, and anxiety (Jeste and Geschwind, 2014). Surgery and anesthesia are known stressors and may produce a regression in motor, social and cognitive behaviors (Short and Calder, 2013). Insight ASD represents a group of patients with altered brain structure and function who may be more susceptible to the potential effects of anesthesia. Alterations in energy metabolism (viz., mitochondrial function) in ASD (Chauhan et al., 2011) may further increase such susceptibility, impacting neuronal networks and consequently behavioral function following anesthesia.

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3. Anesthetic effects on neuronal, glial, dendritic, and synaptic function Anesthetics affect neuronal (Sloan, 2002) and glial (astrocytes and oligodendrocytes) cell function (for a review see Jiang et al., 2015). Glia, supporting elements of neurons, plays an important role in structural (e.g., myelination) and biochemical (e.g., excitatory amino acid sumps) processes. Brain circuits are susceptible to external stressors, including anesthesia. The normal development of brain circuits has been reviewed by Tau and Peterson (Tau and Peterson, 2010). Synapse formation confers a significant effect on brain circuits, since functions such as activitydependent processes may enhance neural circuit formation, while processes that diminish activity or kill neurons may result in perturbations from the norm (Colón-Ramos, 2009). Such plasticity partly resides in the dendritic spines which are the site of mostexcitatory processes (Harms and Dunaevsky, 2007). Thus, the cellular integrity of neurons and their supporting processes (glia), as well the nature of the state of synaptic connectivity provide the underlying basis for brain circuit function that can be disrupted by anesthetic agents. This process may have direct cytotoxic effects on these structural elements or indirect effects through inflammation, stress, or opioids as part of the surgical/ anesthetic state. Below, we briefly review the effects of anesthetics on brain neurons, glial cells, dendrites, synapses, and secondary processes (Fig. 1). 3.1. Neuronal effects 3.1.1. Developing brain In the developing brain, a period of higher vulnerability to anesthetic agents has been found during the period of rapid synaptogenesis (Jevtovic-Todorovic et al., 2003b; Yon et al., 2005) associated with the establishment of new synaptic connections and the maturation of axonal and dendritic outgrowth (Fredriksson et al., 2007; Jevtovic-Todorovic et al., 2003b; Rice and Barone, 2000). The timing of this period in mammals varies according to species. In rodents, this period occurs around the first two weeks of life with a peak on postnatal day 7 (Bayer et al., 1993; Rice and Barone, 2000). In humans, rapid synaptogenesis extends from midfetal gestation to several years after birth (Brambrink et al., 2012b; Rice and Barone, 2000). However, the peak vulnerability for anesthesia-induced neuroapoptosis could vary among brain regions, which may be vulnerable at distinct times of brain development. For example, neuronal cell death following exposure to isoflurane in young adult mice (post-natal day 21 and 49) were noted in brain regions of continued neurogenesis (e.g. dentate gyrus, olfactory bulb)(Deng et al., 2014; Hofacer et al., 2013) (see Section 3.1.3). Vulnerability to anesthesia may depend on the age of the neurons themselves rather than the age of the animal (Deng et al., 2014; Hofacer et al., 2013). While the exact molecular mechanisms by which general anesthetics act are not fully understood, most of the presently used anesthetics are believed to act by two principal mechanisms: (1) by potentiating inhibitory neuronal activity involving y-aminobutyric acid (GABA) receptors, and/or (2) by attenuating excitatory activity through N-methyl-D-aspartate (NMDA) glutamate receptors (Loepke and Soriano, 2012). Researchers in Olney’s laboratory discovered that transient blockade of NMDA glutamate receptors and/or GABA receptors can cause widespread apoptotic changes (defined as: “a genetically directed process of cell self-destruction that is marked by the fragmentation of nuclear DNA, is activated either by the presence of a stimulus or removal of a suppressing agent or stimulus, and is a normal physiological process eliminating DNAdamaged, superfluous, or unwanted cells” www.merriam-webster. com) in the animal developing brain (Ikonomidou et al., 1999,

2001; Jevtovic-Todorovic et al., 2003b). Since these initial studies, preclinical research has demonstrated a relationship between exposure to general anesthetics and structural and functional brain changes, resulting in persistent neurodegeneration in the developing brain as well as impairment in neurocognitive abilities in a wide range of species (Jevtovic-Todorovic et al., 2003b; Loepke and Soriano, 2008; Paule et al., 2011) (Table 1). Moreover, neuroapoptosis is triggered by exposure to most of the routinely used anesthetics and observed in various parts of the cortex (JevtovicTodorovic and Olney, 2008) as well as other brain regions (e.g., cerebellum; basal ganglia) (Deng et al., 2014) and even the spinal cord (Sanders et al., 2008). Several studies have also described chronic attenuation of long-term potentiation in the hippocampal circuits after exposure to multiple anesthetic agents in neonatal animals (Jevtovic-Todorovic et al., 2003b; Kato et al., 2013), suggesting a mechanistic basis for altered brain functions changes. 3.1.2. Aging brain In the aging brain, the potential of general anesthesia to induce neuronal cell death has not received as much attention as that reported in the developing brain. Studies in adult mice and rats have reported neuronal cell death or indication of neurodegeneration (e.g. activation of caspase-3) following exposure to isoflurane in the cortex and hippocampus as well as decreased neuronal density in the region cornu ammonis 1 (CA1 region) of the hippocampus (Ge et al., 2015; Kong et al., 2013, 2015; Lin and Zuo, 2011; Valentim et al., 2010; Xie et al., 2008). Given that the hippocampus is thought to play an important role in memory and cognition (Shohamy and Turk-Browne, 2013); cognitive alterations associated with anesthesia exposure may thus be linked to altered function as a result of neuronal apoptosis in the hippocampus (Kong et al., 2013, 2015) (Table 6). 3.1.3. Neurogenesis Anesthetic agents may also interfere with the proliferation and differentiation of immature neurons in the young and adult/aging brain, in particular in the dentate gyrus of the hippocampus (Erasso et al., 2013; Wagner et al., 2014), an important component of the hippocampal circuit involved in cognition (Shohamy and Turk-Browne, 2013). Neuronal activity could play an important role in the regulation of adult neurogenesis that may be vulnerable to anesthesia exposure (Ge et al., 2007; Tashiro et al., 2006). Although conflicting results have also been reported in adult/aged rats (Stratmann et al., 2010; Zhu et al., 2010), impairments on neurogenesis have been observed after exposure to multiple anesthetic agents, including isoflurane, propofol, and sevoflurane in the young and adult brain (Erasso et al., 2013; Fang et al., 2012; Krzisch et al., 2013). In 7 day-old rats, exposure to sevoflurane for 4 h decreased progenitor proliferation and increased cell death in the dentate gyrus for at least 4 days after the exposure. Interestingly, spatial reference memory was not impaired 2 weeks after the exposure, whereas a significant impairment was found at 6 weeks (Fang et al., 2012). Another group described the effect of 3 h exposure to propofol or isoflurane on nascent cells in the dentate gyrus in young (3-month-old) and adult (20-month-old) rats. Although neither drug affected new cell proliferation, propofol decreased the number of differentiating neurons and increased the number of astrocytes in the young rats but not the old, while isoflurane caused the same effects in the adult rats but not the young rats (Erasso et al., 2013). Moreover, in 8 to 10 weekold mice exposed to propofol for 6 h, impairment in the survival and maturation of adult-born hippocampal neurons was found in neurons of 17 days at the moment of anesthesia but not 11 days, suggesting an age-dependent sensitivity to propofol (Krzisch et al., 2013) (Tables 1, 2, and 6).


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Fig. 1. Altered Brain Networks in Anesthesia – Acute and Chronic Effects. The figure shows three concepts related to anesthetic effects on the brain and behavior: (1) Anesthesia affects a number of brain regions, some, for example the hippocampus (Kodama et al., 2011; Liang et al., 2010; Loepke et al., 2009) seem more susceptible to the effects of these drugs; (2) The acute effects of anesthesia on the brain are usually short lived or subclinical and recovery is probably associated with normalization of brain circuits. (3) Chronic effects of anesthesia are associated with brain changes (aberrant circuits) as a result of a number of brain directed effects including apoptosis and altered dendritic density.

3.2. Anesthetic agents and glial effects It is now well recognized that glial cells have an important role in the development and maturation of neurons. For example, astrocytes support neuronal function via a variety of biochemical and structural processes, and oligodendrocytes provide the axonal myelin sheath (Jiang et al., 2015). An expanding field of research demonstrates that both astrocytes and oligodendrocytes can contribute to neural function and behaviors such as learning and memory (Fields et al., 2013; Pearson-Leary et al., 2016). Exposure to anesthesia may lead to microglia activation, such activation is normally in response to even minor immunological stimuli, toxin, or injury (Kreutzberg, 1996) and, as such, may play an important role in the CNS defense mechanisms (Kreutzberg, 1996). In animals, activation of microglia following anesthesia exposure may vary according to the dose, the anesthetic, and the age at the moment of the exposure (see below and section 3.4.2). 3.2.1. Developing brain A number of anesthetic agents used in clinical practice may have a toxic effect on glial cells, in particular on astrocytes and oligodendrocytes (Fig. 2 and Table 2). In the context of brain development, Lunardi et al. (2011) first described impairment in the development of pup astrocytes in cultures following 24 h of exposure to 3% isoflurane. These impairments resulted in a disruption of the actin cytoskeleton and delayed astrocyte maturation. Evidence for the toxic effects of isoflurane on astrocytes was confirmed by later studies (Culley et al., 2013; Erasso et al., 2013; Ryu et al., 2014). These data suggest that

exposure to isoflurane impairs the cytoskeleton during the development of astrocytes. Moreover, 3-h exposure to propofol of nascent cells in the rat dentate gyrus led to a significant increase in astrocytes 24 h after the exposure (Erasso et al., 2013). In addition, 6 day-old mice exposed to 3% sevoflurane for 2 h at multiple time points (3 days) demonstrated microglia activation in the hippocampus (Shen et al., 2013). By contrast, activation was found neither when mice were exposed to a single dose of sevoflurane, multiple dose of desflurane, nor when mice were exposed during early adulthood (60 day-old) (Shen et al., 2013). Evidence for apoptosis of rhesus monkey fetal or neonatal oligodendrocytes after exposure to isoflurane or propofol was found at least 3 h after the exposure (Brambrink et al., 2012a; Creeley et al., 2013, 2014; Noguchi et al., 2016). The onset of vulnerability of oligodendrocytes seems to occur when the myelination of axons begins (Creeley et al., 2014). 3.2.2. Aging brain Activated microglia express increased levels of some proinflammatory cytokines (e.g., tumor necrosis factor-a (TNF-a), interleukin-6 (IL-6), interleukin-1 (IL-1)) that are hypothesized to play a role in cognitive impairment and neurodegenerative diseases (Patanella et al., 2009; Perry, 2004; Schuitemaker et al., 2009). Following exposure to 1.4% isoflurane for 2 h or to pentobarbital sodium anesthesia for a surgical procedure (appendectomy), microglia activation is observed in the hippocampus of 18 month-old mice or adult mice vs. control mice at 3, 14, and 28 days (Wang et al., 2015, 2016). Similar findings were discovered in a mouse model of orthopedic surgery and isoflurane anesthesia:

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Table 1 Apoptosis, Behavioral Changes and Anesthesia in young animals. Anesthetic + Mechanisms of Action

Species

Isoflurane GABAA – agonist

Rat slice cultures at 0.75, 1, or 2 MAC 7d for 6 h

Increased neurotoxicity after 1 MAC Hippocampal CA1, CA3 exposure

Brosnan and Bickler (2013)

Isoflurane

Mice (7 d)

0.75% for 4 h

Increased neuroapoptosis

Cattano et al. (2008a)

Isoflurane

Mice (7 to 49 d)

1.5% for 6 h

Increased neuroapoptosis in 7 d and Neocortex, caudate/putamen, in a lesser extent in 21 and 49 d. hippocampal CA1, cerebellum, olfactory bulb, dentate gyrus

Isoflurane

Mice (7, 27, 49 d)

1.5% for 6 h

Forebrain, DGC, olfactory bulb Hofacer et al. 7 d: neuronal loss in several forebrain structures, in the olfactory (2013) bulb, but not in DGCs; 21d: mostly apoptosis in DGCs, olfactory bulb; 47d: apoptosis in DGCs, olfactory bulb

Isoflurane

Mice (7 or 8 d)

1.5% for 6 h (0.55–0.6 MAC)


Superficial neocortex layers II, Istaphanous III et al. (2011)

Isoflurane

Rats (7 d)

0.75, 1, 1.5% for 6 h

Dose-dependent increased neuroapoptosis

LD, AV thalamic nuclei, parietal cortex (layer II)

JevtovicTodorovic et al. (2003b)

Isoflurane

Mice C57BL/6 (6 d)

2% for 3 or 6 h

Increased neuroapoptosis after 6h but not 3 h (less than with equipotent dose of desflurane), impaired long-term memory in adulthood

Parietal cortex (layer II), sensory cortex (layer IV), dorsal hippocampal commissure, retrosplenial cortex

Kodama et al. (2011)

Isoflurane

Mice (7 d)

0.75% for 6 h

Increased neuroapoptosis higher than with equipotent dose of sevoflurane

Hippocampal region CA1, cerebral cortex

Liang et al. (2010)

Isoflurane

Mice (7 d)

1.5% for 6 h

Increased neuroapoptosis. In adulthood, no decrease in neural density and, no learning deficits

Neocortex, thalamus, dentate gyrus, hippocampal CA1/2/3 regions

Loepke et al. (2009)

Isoflurane

Rats (7 d)

0.75% for 6 h


Hippocampus

Ma et al. (2007)

Isoflurane

Rats (7 d)

1 Mac for 4 h

Deficits in long- and short- term memory in early adulthood

Isoflurane

Rats (gestational day 21)

Chemomyelotomy under In fetus and newborns, alteration of 2.5%/L/min in 100% O2 behavior

Ronca et al. (2007)

Isoflurane

Rats (7 d) and hippocampal slice cultures

0.75% for 6 h + saline or dexmedetomidine (1, 10, or 25 mg/kg) at 0, 2 and 4 h.

Increased neuroapoptosis with Cortex, thalamus, isoflurane (reduced with hippocampus dexmedetomidine). At 40 d-old, long-term memory impairment after isoflurane (attenuated by dexmedetomidine). Comparable results in hippocampal slice cultures.

Sanders et al. (2009)

Isoflurane

Rats (7 d)

0.75% for 6 h + saline or dexmedetomidine (25, 50, or 75 mg/k) at 0, 2 and 4 h.

Increased neuroapoptosis following isoflurane exposure (reduced with dexmedetomidine)

Isoflurane

Mice (6 d)

2% for 2 h/day during 3 days

Impaired spatial learning & memory 3 weeks later

Tao et al. (2016)

Isoflurane

H4 human cells

2% for 6 h

Increased apoptosis and amyloid b protein levels

Xie et al. (2006)

Isoflurane

Rats (1 to 14 d)

0.75, 1, or 1.5% for 2 to 6 h Increased neuroapoptosis in 1 and 3 LD & AV thalamic nuclei d with 1.5% isoflurane

Yon et al. (2005)

Isoflurane

H4 human cells & 1% for 6 h primary neurons of

# Anesthetic challenges

Brain neurotoxicity

No increased neuroapoptosis

Brain Region

Cerebral cortex, Caudate/ Putamen

Reference

Deng et al. (2014)

Ramage et al. (2013)

Cerebral cortex


Zhen et al. (2009)


127

Table 1 (Continued) Anesthetic + Mechanisms of Action

Species


Brain neurotoxicity

Brain Region

Reference

naïve mice (gestational day15) Isoflurane

Mice (7 d)

0.75% for 4 h/3 days

Hippocampal CA1 Abnormal hippocampal histone acetylation, memory impairments 3 months later

Zhong et al. (2015)

Isoflurane

Rats (14 and 60 d) Mice (14 d)

1.7% 35 min/4 days

14 d rats and mice: Persistent impaired learning and memory + reduced neurogenesis in hippocampus, but no effect on cell death in hippocampus

Zhu et al. (2010)

Isoflurane

Monkeys (5 or 6 d)

1% isoflurane for 8 h


Ether NMDA-antagonist

Rats (gestational day 21)

Chemomyelotomy under In fetus and newborns, alteration of 50 mL behavior

Desflurane GABAA – agonist

Mice C57BL/6 (6 d)

4 or 8%, 3 or 6 h

Dose-dependent increased neuroapoptosis after 6 h, impaired long-term memory and working memory in adulthood

Parietal and sensory cortex, dorsal hippocampal commissure, retrosplenial cortex

Desflurane

Mice (7 or 8 d)

7.4% for 6 h (0.55- 0.6 MAC)


Superficial neocortex layers II Istaphanous and III et al. (2011)

Desflurane

Mice (6 d)

8% for 2 h/day during 3 days

No effect on learning and memory 3 weeks later.

Tao et al. (2016)

Desflurane

H4 human naïve cells + H4-APP cells

12% for 6 h 12% + hypoxia for 6 h

With hypoxia, effect on H4-APP cells: increased neuroapoptosis and amyloid b generation, altered APP processing

Zhang et al. (2008)

Sevoflurane GABAA – agonist

Rats (6 d)

Between 3 to 5% for 6 h


Bercker et al. (2009)

Sevoflurane

Rat slice cultures at 1 or 2 MAC for 6 h 7d

Increased neurotoxicity after 1 MAC Hippocampal CA1, CA3, exposure dentate gyrus


Sevoflurane

H4 cells: 4.1% for 6 h; H4 human cells Mice (5 to 9 month- Mice: 2.5% for 2 h old)

Increased neuroapoptosis and Mice: prefrontal cortex amyloid b protein levels production

Dong et al. (2009)

Sevoflurane

Rats (7 d)

3–5% for 4 h

Increased neuroapoptosis, decreased Hippocampal dentate gyrus progenitor proliferation, impaired memory 6 weeks later

Fang et al. (2012)

Sevoflurane

Mice (7 or 8 d)

2.9% for 6 h (0.55–0.6 MAC)


Superficial neocortex layers II Istaphanous and III et al. (2011)

Sevoflurane

Rats (7 d)

1 or 2% for 2 h

Suppression of long-term potentiation with 2%

Hippocampus

Sevoflurane

Mice C57BL/6 (6 d)

3% for 3 or 6 h

Parietal and sensory cortex, Increased neuroapoptosis after 6 h (less than with desflurane), impaired dorsal hippocampal long-term memory in adulthood commissure, retrosplenial cortex


Sevoflurane

Mice (7 d)

1.1 % for 6 h

Less neuroapoptosis than with isoflurane

Liang et al. (2010)

Sevoflurane

Rats (7 d)

1 Mac for 4 h

Impaired long-term memory in early adulthood

Sevoflurane

Mice (6 d)

3% for 6 h

Increased neuroapoptosis, impaired learning in adulthood, abnormal social behaviors

Caudate/Putamen, retrosplenial cortex, dorsal hippocampal commissure, neocortex

Satomoto et al. (2009)

Sevoflurane

Rats (7 d)

2.5% for 4 h

No significant neuronal loss, decreased expression of PSD-95,

Hippocampus

Wang et al. (2013)

Hippocampus

Zou et al. (2011) Ronca et al. (2007)

Cerebral cortex


Kato et al. (2013)

Ramage et al. (2013)

128



Species


Brain neurotoxicity

Brain Region

Reference

impaired spatial learning and memory 7 weeks later Nitrous oxide NMDA-antagonist

Rats (7 d)

50, 75, or 150% for 6 h



Nitrous oxide

Rats (7 d)

75% for 6 h


Ma et al. (2007)

Nitrous oxide

Rats (1 to 14 d)

50, 75, or 150% for 2, 4, or No increased neuroapoptosis 6h

Yon et al. (2005)

Nitrous oxide

H4 human cells & 70% for 6 h primary neurons of naïve mice (gestational day15)


Zhen et al. (2009)

Nitrous oxide

Monkeys (5 or 6 d)


Zou et al. (2011)

Xenon NMDA-antagonist

Rat slice cultures at 0.75, 1, or 2 MAC for 6 h 7d

Increased neurotoxicity after 1 or 2 MAC exposure

Hippocampal CA1, CA3, dentate gyrus


Xenon

Mice (7 d)

70% for 4 h



Cattano et al. (2008a)

Xenon

Rats (7 d)

75% for 6 h


Propofol GABAA – agonist

Rats (6 d)

Cumulative 90 mg/kg for Increased neuroapoptosis, impaired 4.5 h learning 7 weeks later

MD, LD thalamus, subiculum

Bercker et al. (2009)

Propofol

Mice (5 to 7 d)

25 to 300 mg/kg

Increased neuroapoptosis with doses 50 mg/kg


Cattano et al. (2008b)

Propofol

Mice (10 d)

10 or 60 mg/kg

Increased neuroapoptosis with high dose, no effect on motor activity or learning.

Olfactory bulb, stria terminalis

Fredriksson et al. (2007)

Propofol

Rats (6 d)

30 mg/kg every 90 min for 3 h

Increased neuroapoptosis, minor behavioral changes at 30 d

Cortical and thalamic area

Karen et al. (2013)

Propofol

Rats (7 d)

75 mg/kg 75 mg/kg per day/7 days

Hippocampal CA1 region Increased neuroapoptosis (dosedependent effect) Repeated doses: decreased neuronal density, impaired spatial learning & memory 2 weeks later

Yu et al. (2013)

Midazolam GABAA – agonist

Rats (7 d)

3, 6 or 9 mg/kg



Midazolam

Mice (10 d and 20 d)

50 mg/kg

20 d mice: acute reduced locomotor activity at 3 and 24 h post-exposure but not anymore after 72 h.

Xu et al. (2009)

Midazolam

Rats (1 to 14 d)

3 to 9 mg/kg


Yon et al. (2005)

Midazolam

Mice (7 d)

9 mg/kg

Halothane GABAA agonist

Rats (from conception to 6 d)

8 to 12 ppm 8 h/5 days/weeks

Superior parasagittal cerebral Quimby Increased neurodegeneration with 10 ppm, learning deficits weeks post- cortex et al. (1974) exposure

Ketamine NMDA-antagonist

Rats (7 d)

75 mg/kg with or without 25 mg/kg dexmedetomidine (1/ day/3 days)

Neuroapoptosis, impaired learning and memory after ketamine exposure. Effects attenuated by dexmedetomidine

70% for 8 h

1


Ma et al. (2007)


CA1 region of hippocampus and dentate gyrus

Young et al. (2005)

Duan et al. (2014)


129


Species


Brain neurotoxicity

Brain Region

Reference

Ketamine

Mice (10 d)

50 mg/kg

Increase dneurodegeneration, behavioral and learning deficits 2 months later.

Parietal cortex


Ketamine

Mice (10 d)

25 mg/kg

No increased neuroapoptosis, disrupted spontaneous activity & learning deficits in young adulthood


Ketamine

Rats (7 d)

25, 50, 75 mg/kg 1, 25 mg/kg 1, every 90 min for 9 h

Repeated doses: Temporal & parietal cortex, increased neuroapoptosis (24 h after hippocampus CA1, dentate initial injection) gyrus, thalamus, VM ypothalamus, BM amygdala.

Hayashi et al. (2002)

Ketamine

Rats (7 d)

7 20 mg/kg

Increased neurodegeneration

Ikonomidou et al. (1999)

Ketamine

Rats (7 d)

5 20 mg/kg for 6 h or 5 10, 25, 50 mg/kg of dexmedetomidine for 6 h or combined Ket + Dex with various doses

At clinical dose, dexmedetomidine is neuroprotective. High cumulative dose and concentrations induce neuroapoptosis. Ketamine alone and combines with DEX induced neuroapoptosis

Liu et al. (2016)

Ketamine

Monkeys (5 or 6 d)

20 mg/kg + 20–50 mg/ (kg/h) for 24 h.

Long-lasting deficits in brain functions (months after exposure)

Paule et al. (2011)

Ketamine

Rats (7 d)

20 mg/kg 10 or 20 mg/kg every 90 min (7 doses)

Mainly DL thalamus Repeated high doses: increased neurodegeneration (24 h after initial dose)

Scallet (2004)

Ketamine

Monkeys (122 d of gestation, 5 and 35 d)

20 mg/kg + 20–50 mg/ (kg/h) for 3 or 24 h.

Increased neuroapoptosis after 24 h in gestational and 5 day

Frontal cortex (layers II and III)

Slikker et al. (2007)

Ketamine

Mice (7 d)

10–40 mg/kg

Increased neuroapoptosis (dosedependent)


Young et al. (2005)

Ketamine

Monkeys (5 or 6 d)

20 mg/kg + 20–50 mg/ (kg/h) for 3, 9, or 24 h.

Increased neuroapoptosis after 9 and Frontal cortex (layers II and 24 h III)

Zou et al. (2009)

Thiopental GABAA – agonist

Mice (10 d)

5 or 25 mg/kg

No increased neuroapoptosis and no effect on motor activity or learning.


Diazepam GABAA – agonist

Mice (10 d)

5 mg/kg

Increased neurodegeneration

Isoflurane + Nitrous oxide

Rats (7 d)


1

CA1 hippocampus, dentate gyrus, thalamus, subiculum, caudate, hypothalamus, thalamus, frontal, parietal, cingulate, retrosplenial cortex.

LD thalamus


0.75% + 35 or 75% for 6 h Increased neuroapoptosis (more than isoflurane alone)

Hippocampus

Ma et al. (2007)

Rats (7 d)

0.75% + 75% for 6 h


Spinal cord


Isoflurane + Nitrous Oxide

Monkeys (5 or 6 d)

1% + 70% for 8h

Increased number of caspase 3 after Frontal cortex, temporal 8h gyrus, hippocampus

Zou et al. (2011)

Isoflurane + Midazolam

Rats (7 d)

0.75% + 9 mg/kg for 6 h


LD, AV thalamic nuclei, Parietal cortex (layer II)


Ketamine + Diazepam

Mice (10 d)

50 mg/kg + 5 mg/kg

Increased neurodegeneration, behavioral and learning deficits 2 months later

Parietal cortex, LD thalamus


Ketamine + Thiopental

Mice (10 d)

25 mg/kg + 5 mg/kg

Increased neuroapoptosis, disrupted Olfactory bulb + stria spontaneous activity & learning in terminalis young adulthood


Ketamine + Propofol

Mice (10 d)

25 mg/kg + 10 mg/kg

Increased neuroapoptosis, disrupted Olfactory bulb + stria spontaneous activity & learning in terminalis young adulthood


130



Species


Ketamine + Midazolam

Mice (7 d)

40 mg/kg

Isoflurane + Xenon

Mice (7 d)

0.75% + 70% for 4 h

Isoflurane + Xenon

Rats (7 d)

0.75% + 30 or 60% for 6 h No increased neuroapoptosis

Isoflurane + Midazolam + Nitrous Oxide

Rats (7 d)

0.75% + 9 mg/kg + 75 vol% Increased neuroapoptosis, long-term for 6 h hippocampus potentiation suppression, impaired spatial learning & memory weeks after.

Thalamus, parietal cortex, widespread effect in other regions



Rats (7 d)

0.75% + 9 mg/kg + 75% for Disturbance in brain-derived 2, 4, or 6 h neurotrophic factor-activated apoptotic cascade

Cerebral cortex and anterior thalamus

Lu et al. (2006)


Rats (7 d)

0.75% + 9 mg/kg + 75% for Significant neuronal loss later in life Cerebral cortex and anterior 6h (30 day-old) thalamus

Nikizad et al. (2007)


Rats (1 to 14 d)

0.75% + 9 mg/kg + 75 vol% Age-dependent increased for 6 h neuroapoptosis

Anterior thalamus, cingulate, occipital, parietal cortex, subiculum

Yon et al. (2005)


Rats (7 d)

0.75% + 9 mg/kg + 75% for Increased neuroapoptosis (reduced 2, 4, or 6 h when melatonin added)

Cerebral cortex, anterior thalamus

Yon et al. (2006)


1% + 70% for 6 h H4 human cells (naïve and H4-APP) & primary neurons of naïve mice (gestational day15)

Increased neuroapoptosis in H4 naïve cells and primary neurons naïve mice, increased B-amyloid protein levels in H4-amyloid precursor protein cells

Zhen et (2009)

Morphine

Mice (from 5 to 9 d) 2 mg/kg 2times/day, 5 days

Alteration of cognitive functions in adulthood

Boasen et al. (2009)

Morphine

Rats (gestational days 11–18)

3 times 5 mg/ kg + repeated 10 mg/kg, 2times/day for 7 days.

Hippocampus Offspring at 22–31 d: impaired spatial memory and dentate synaptic plasticity

Niu et al. (2009)

Morphine

Rats (gestational days 11–18)

3 times 5 mg/ kg + repeated 10 mg/kg, 2times/day for 7 d.

Gender-dependent impaired learning and memory in adulthood

Šlamberová et al. (2001)

1

+ 9 mg/kg

1

Brain neurotoxicity

Brain Region

Reference


Cerebral cortex and caudate/ putamen

Young et al. (2005)

Increased neuroapoptosis (less than isoflurane alone)


Cattano et al. (2008a) Ma et al. (2007)

al.

Key: mg = milligram; kg = kilogram, d = day, LD = laterodorsal, AV = anteroventral, MD = mediodorsal, VM = ventromedial, BM = basomedial, DL = dorsolateral, DGC = dentate granule cell.

significant morphological changes of microglia reactivity were recorded in the hippocampus of young adult mice (3 month-old) 24 h and 3 days after the surgery, with a return to the baseline by 7 days (Cibelli et al., 2010) (Table 2). 3.3. Dendritic effects and synaptic connectivity effects 3.3.1. Developing brain Changes in neuronal or glial processes may modify dendritic complexity. Maladaptive changes in dendritic complexity are key to altered brain connectivity (Segal, 2005; Wang and Zhou, 2010). Briner et al. (2010) observed an increase in dendritic spine density in the medial prefrontal cortex in young rats 6 h after exposure to isoflurane, sevoflurane, or desflurane. The same group reported also development stage-dependent modifications in dendritic spine density in the medial prefrontal cortex with propofol (Briner et al., 2011). Moreover, the triple exposure of neonatal rats to midazolam, nitrous oxide, and isoflurane, a commonly administered drug combination, for 6 h led to reduced synaptic density and

impaired neuropil structure in the subiculum persisting for at least 2 weeks after the exposure (Lunardi et al., 2010). Changes in dendritic spine morphology with decreased density has also been observed in rodent hippocampal slices (Platholi et al., 2014) (Table 3). General anesthesia may also interfere with axonal and dendritic growth, disrupting axon guidance, or causing degeneration of axons or dendrites (for review, see Wagner et al., 2014). Using rodent cortex brain slices and isolated neuronal assays, Mintz et al. (2013) reported a disruption of axon guidance mechanisms by a broad range of anesthetics that could affect normal brain development. Moreover, isoflurane has been shown to block actin-based motility in dendritic spines and fibroblasts in developing hippocampal neuron cultures of rodents (Kaech et al., 1999) (Table 3). Taken together, these findings suggest that exposure to general anesthesia may have a lasting effect on the formation and maintenance of dendritic spines and synapses, neuronal interconnectivity, and consequently circuit function (Fig. 2). The integrity of


131

Table 2 Anesthetic Effects on Glia. Anesthetic + Mechanisms of Species Action


Brain neurotoxicity

Brain Region

Reference

Isoflurane GABAA agonist

Monkeys (6 d)

0.7–1.5% for 5 h

Significant neuroapoptosis and loss 6.3% of total myelinating oligodendrocytes 3 h post-exposure

Throughout the brain

Brambrink et al. (2012a)

Isoflurane + buprenorphine

Mice (3 months)

Anesthesia for orthopedic surgery or anesthesia/ analgesia alone

Changes of microglia reactivity 24 h and 3 d after Hippocampus surgery, increased plasma concentration of IL-6 and IL-1b in the hippocampus 6 h after the surgery, memory impairment. With anesthesia alone, less changes observed.

Cibelli et al. (2010)

Isoflurane

Monkeys (120 d gestational age)

Intermediate surgical plane of anesthesia for 5 h

Increased apoptosis of neurons and oligodendrocytes 3 h post-exposure

Neuroapoptosis: cerebellum, caudate, putamen, amygdala, several cerebro-cortical regions Oligodendrocyte apoptosis: diffusely over many white matter regions

Creeley et al. (2014)

Isoflurane

Culture astrocytes from rats embryos of 18 d

1.4% for 4 h

Decrease in glial fibrillary acid protein expression and in alpha tubulin levels for at least 2 d

Cerebral cortex

Culley et al. (2013)

Isoflurane

Rats (3 and 20 months)

1.5% for 3 h

3 months: no effect, 20 months: increased number Dentate gyrus of astrocytes

Erasso et al. (2013)

Isoflurane

Rats primary astroglia cultures 1 to 2 d

3% for 24 h

Disruption of actin cytoskeleton and delay astrocytes maturation

Cerebral cortex

Lunardi et al. (2011)

Isoflurane

Monkeys (6 d)

1.5 to 3% for 5 h

Increased apoptosis of neurons and oligodendrocytes

Mainly in thalamus, cortex, Noguchi et al. corpus callosum, corona radiata (2016)

Isoflurane

Astrocytes from 1 to 1.2, 2.4, or 3.6% for 3 d mice co-cultured 5 h with unexposed neurons

Isoflurane

Adult mice (3 months) or aged mice (18 months)

Isoflurane

Decreased ability of astrocytes to support neuronal Cerebral cortex growth at and above 2.4% exposure, persistent up to 24 h

Ryu et al. (2014)

1.4% for 2 h, or pentobarbital sodium for surgery (appendectomy)

Aged mice: impaired spatial learning memory, Hippocampus hippocampal microglia activation, enhanced IL-1b, TNF-a, interferon-g following exposure to isoflurane or surgery

Wang et al. (2015)

Aged mice (35–40 gr)

1.4% for 2 h, or pentobarbital sodium for surgery (appendectomy)

Following isoflurane or surgery: Enhanced protein Hippocampus levels of TNF-a, IL-1b, interferon-g, microglia marker Iba–1 + dysregulated protein levels of IL-4, IL-10. Impairment of acquisition/learning of spatial task. Attenuated by pretreatment Minocycline (iso/ surgery)

Wang et al. (2016)

Desflurane GABAA – agonist

Mice (6 d and 60 d)

9% for 2 h/3 days

No cognitive impairment, no indiceof neuroinflammation

Shen et al. (2013)


Mice (6 d and 60 d)

Brain tissues and 3% for 2 h or 3% for Young mice – repeated exposure: cognitive 2 h/3 days impairment 3 weeks later and neuroinflammation hippocampus (increased IL-6 level & TNF-a) in brain tissues and increased amount of IBA1 positive cells in hippocampus

Propofol GABAA agonist

Moderate plane of Monkeys (120 d gestational age) and surgical anesthesia 6d for 5 h

Increased apoptosis of neurons and oligodendrocytes 3 h post-exposure

Creeley Fetus: subcortical and et al. caudal brain regions. Neonate: neocortical (2013) regions and caudal regions

Propofol


3 months: increased number of astrocytes, 20 months: no effect

Dentate gyrus

Key: mg = milligram; kg = kilogram; d = day.

35 mg/kg for 3 h

Hippocampus

Shen et al. (2013)


132


Fig. 2. Examples of Anesthesia-induced changes in Neural Tissue. Left Panel: Isoflurane-induced changes in the primate brain. The figure shows both apoptotic changes in neurons (red) and glia (green) after 5 h of isoflurane vs. no anesthesia (control). Note the distribution of the changes – high levels of apoptosis in neurons in the thalamus and cortex and high levels in white matter (e.g., corpus callosum). (Adapted from Noguchi et al., 2016,http://creativecommons.org/licences/by/4.0/). Right Panel: Acute changes in dendritic spine density following isoflurane anesthesia in hippocampal cultures. Isoflurane alters dendritic spine morphology and reduces dendritic spine density within 20 min of exposure. (Adapted from Platholi et al., 2014, http://creativecommons.org/licences/by/4.0/). These processes may be transient with the acute effects of isoflurane exposure in most subjects, but may confer long-term changes in non-resilient subjects.

neurons is clearly a factor in their ability to maintain or evolve dendritic spines. Indeed, the modulation of dendritic spine plasticity may contribute to the immediate effects of anesthesia (Meyer, 2015). 3.3.2. Aging brain We are unaware of any study reporting specifically the effect of general anesthesia in mature neuronal synaptic connectivity. However, changes in neuronal morphology are observed with aging and characterized by small region-specific changes in dendritic branching and spine density (Burke and Barnes, 2006). Such changes are relevant since impairment of cognitive functions associated to normal aging has been linked to specific and small synaptic alterations mainly in the hippocampus and the prefrontal cortex (Morrison and Baxter, 2012). 3.4. Secondary processes in anesthesia as contributors to altered brain function Surgery itself involves a number of processes that may contribute to overall changes in brain function (Fig. 3). These include the surgical stress response, neuroinflammation, pain, and opioids (Borsook et al., 2010). 3.4.1. Surgical stress response During the perioperative period, acute stress can produce lasting alterations in cerebral structure and function. Although the potential neurotoxicity of anesthetic drugs contributes to perioperative stress, anxiety, pain and the surgical stress response are also potential contributors (for a review see Borsook et al., 2010). The stress response to surgical procedures is characterized by neurohumoral, immunologic, and metabolic alterations, and the magnitude of the response is usually proportional to the degree of surgical injury. The stress response may be modulated by the choice of the anesthetic technique (Gruber et al., 2001; Kostopanagiotou et al., 2010; Ledowski et al., 2005; Marana et al., 2003, 2013; Philbin et al., 1990). Volatile anesthetics suppress the stress response the least (Kostopanagiotou et al., 2010; Ledowski et al., 2005), with sevoflurane and desflurane impacting the response differently (Marana et al., 2013). Total intravenous anesthesia with propofol suppresses the cortisol response and production of catecholamines to a greater degree than balanced anesthesia with a volatile agent (Ledowski et al., 2005). High dose opioid anesthesia does not prevent a hormonal or metabolic (Gruber et al., 2001) or hemodynamic (Philbin et al., 1990) response to stress.

3.4.2. Neuroinflammation Surgical trauma leads to an inflammatory response at the site of injury. The inflammatory response is influenced by different factors including preoperative health status, biological variation, duration and extent of the surgical procedure, drug management, pain, anxiety, and the anesthetic agents used (Schneemilch et al., 2004). The increase in inflammatory mediators may influence neuronal functioning via the production of pro-inflammatory cytokines and activation of microglia (Hovens et al., 2014). Increased of pro-inflammatory cytokines (e.g. TNF-a, IL-6, IL-1) in neurons and microglia, indicators of neuroinflammation, are hypothesized to play a role in cognitive impairment and neurodegenerative disease (Patanella et al., 2009; Perry, 2004; Schuitemaker et al., 2009). Activation of glia and pro-inflammatory cytokines may also enhanced pain signaling and may be involved in persistent postsurgical pain (Block, 2016; Mika et al., 2013; Wieseler-Frank et al., 2005). Anesthetic-induced neurotoxicity may also be linked to neuroinflammation. There is for example a clear link between inflammation and apoptosis (Haanen and Vermes, 1995); such a link may be a primary or secondary effect of anesthesia. Therefore anesthetics may disrupt the immune response independently of the surgical effects (Yuki and Eckenhoff, 2016); and they may accentuate sensitized neuroplastic processes (e.g., synaptic connections). Moreover, many studies have also found an association between exposure to anesthetic agents and neuroinflammation in the young and aging brain (see below). 3.4.2.1. Developing brain. Anesthesia may induce changes in cytokine expression in the CNS. For example, repeated exposures of 6-day-old mice to 3% sevoflurane for 2 h daily for 3 days (but not a single exposure to the agent or exposure to desflurane), resulted in an increase in the levels of IL-6 and TNF-a in brain tissue, and increased expression of IBA-1 positive cells (marker of microglia activation) in the hippocampus (Shen et al., 2013). By contrast, 6 h of propofol anesthesia exposure showed no major impact on the expression of pro-inflammatory cytokine in the hippocampus and prefrontal cortex of rat pups assessed 6 and 24 h after the exposure (Kargaran et al., 2015). Taken together, these results suggest that the impact of anesthesia on proinflammatory cytokine expression profiles in the CNS might depend on the anesthetic used (Tables 2 and 4). 3.4.2.2. Aging brain. Neuroinflammation is also hypothesized to be involved in the potential relationship between the onset and progression of neurodegenerative disease and exposure to general


133

Table 3 Anesthetic Effects on Dendrites. Anesthetic + Mechanisms of Species Action


Brain neurotoxicity

Brain Region

Reference

Isoflurane GABAA agonist

Rats (16 d)

1.5% for 30 min, 1, or 2h

Time-dependent changes in dendritic spine architecture 6 h post-exposure (both apical and basal) No increase in neuroapoptosis

Medial prefrontal cortex

Briner et al. (2010)

Isoflurane

Embryos rats hippocampal neurons 15.3 mM cultures at 19 d

Blockage of actin-based motility in dendritic Hippocampus spines and fibroblasts

Kaech et al. (1999)

Isoflurane

Embryos mice cells culture at 18 d, 1.2% for 8 h rats cells culture at 3 d

Disruption of axon guidance mechanisms

Mintz et al. (2013)

Isoflurane

Embryos rats hippocampal neurons 2 Vol% cultures at 18 d

Alteration dendritic spine morphology and reduced density within 20 min of exposure

Hippocampus

Platholi et al. (2014)

Desflurane GABAA agonist

Rats (16 d)

7% for 1/2, 1, or 2 h

After 2 h exposure, altered dendritic spine architecture 6 h post-exposure (both apical and basal). No increase in neuroapoptosis



Sevoflurane GABAA agonist

Rats (16 d)

2.5% for 1/2, 1, or 2 h

Time-dependent changes in dendritic spine architecture 6 h post-exposure (both apical and basal), no increase in neurodegeneration or neuroapoptosis



Propofol GABAA agonist

Rats (5 to 30 d)

40 mg/kg + 4 doses (20 mg/kg) for 6 h 50 mg/kg

Decreased dendritic spine density and synapse density in 5 and 10 d rats, persisting in adulthood, Increased dendritic spine density in 15, 20, 30 d rats, Persistent changes in young adulthood



Midazolam + Nitrous Oxide + Isoflurane

Rats (7 d)

9 mg/kg + 75% + 0.75% for 6 h

Reduced synaptic density, Impaired neuropil structure at least 2 weeks post-exposure

Subiculum

Lunardi et al. (2010)

Key: mg = milligram; kg = kilogram ; d = day.

anesthesia may be a basis for disease exacerbation (Box 2) (Hussain et al., 2014; Yang and Fuh, 2015). A number of anesthesia-related studies have focused on the hippocampus because of its association with neurocognition. A potential association between an increase in pro-inflammatory cytokines in the hippocampus and transient neurocognitive decline has been suggested in adult rodents (Terrando et al., 2010; Wan et al., 2007). In a mouse model of orthopedic surgery utilizing isoflurane anesthesia, increased plasma concentrations of IL-6 and IL-1b were found in the hippocampus of 3 month-old mice 6 h after the surgery (Cibelli et al., 2010). Increased levels of pro-inflammatory cytokine IL-1b were also observed in the hippocampus of 20 month-old rats exposed to 1.3% isoflurane for 4 h (Kong et al., 2013, 2015). By contrast, no effect on the levels of IL-6 and TNF-a were reported, and no further effect was observed 2 weeks later (Kong et al., 2013, 2015). Shorter exposure time can also lead to an increased level of pro-inflammatory cytokines. For example, Lin and Zuo (2011) exposed 4 month-old rats to 1.2% isoflurane for only 2 h and found a similar increase of IL 1-b in the hippocampus 6 h later. With the same dose of anesthetic and duration, comparable increase of IL-6 and IL-1b were found in the hippocampus of older rats (18 month-old) (Cao et al., 2012). Furthermore, exposure of 20 month-old rats to isoflurane or propofol in a surgery model (right carotid artery exposure) induced a similar degree of neuroinflammation (increase of IL-1b and Iba-1) with one or the other drug in the cerebral cortex and hippocampus (Zhang et al., 2015). In contrast, propofol reportedly attenuates the effect of sevoflurane-induced neuroinflammation (Tian et al., 2015), suggesting differences in anesthetic agents may be an issue. In most of these studies, alteration of behavior and cognitive functions (spatial memory) were also present (see Section 4.2.1) (Tables 2, 4, and 6).

3.4.3. Pain Nociception is common in most, if not all, surgeries either during or post procedure, even with anesthesia (Kehlet et al., 2006). The association of pain/nociception may be an additional contributor to the surgical event. Although some volatile and intravenous anesthetics have analgesic properties (e.g. ketamine, nitrous oxide), most of them are not analgesics, so that nociceptive input in the perioperative period and pain can induce alterations in gene expression and rapid neuronal sensitization (central sensitization) (Latremoliere and Woolf, 2009). Acute postoperative pain may lead to persistent pain (Kehlet et al., 2006; Perkins and Kehlet, 2000). Multiple studies have suggested that pain and pain-related stress can be deleterious for the CNS in the young and adult brain (see below). 3.4.3.1. Developing brain. Nociception may enhance anestheticinduced neuroapoptosis in neonatal rats (Shu et al., 2012). Enhanced neuroapoptosis is also seen in neonate rats exposed to repeated inflammatory pain, and these effects were mitigated by the administration of low-dose of ketamine (Anand et al., 2007), presumably due to its anti-inflammatory (Hirota and Lambert, 2011) or effects on attenuating pain. Similarly, enhanced neuroapoptosis following exposure of neonatal rats to ketamine for 6 h was attenuated when a peripheral noxious stimulation was performed after the first injection of ketamine (Liu et al., 2012a). Importantly, pain in the neonatal period may have long-term effect – peripheral inflammatory pain in rat pups (Ruda et al., 2000) or mice (Blom et al., 2006), or repeated painful stimulation in neonatal rat pups (Anand et al., 1999, 2007) have been associated with long-term changes in nociceptive pathways in adulthood. Similar findings were found in children: for example, in children born very preterm, a higher number of neonatal pain experiences

134


Fig. 3. Complex interactions (stressors) occurring during the Anesthetic State. The figure shows the complex interactions of factors in the pre-anesthetic, anesthetic and post anesthetic status that can potentially modulate the anesthetic effects on the brain and behavior. Pre-anesthetic status: (1) the vulnerability to anesthetic agents may vary according to gender (Jevtovic-Todorovic et al., 2001; Xie et al., 2015); (2) or age (Jevtovic-Todorovic and Carter, 2005; Yon et al., 2005) or may depend on (3) the number of previous exposures (e.g., Wang et al., 2014); and (4) on the health status before the surgery (for a review in elderly humans see: Avidan and Evers, 2011), etc. Anesthetic status: The overall changes in brain function are not only related to the anesthetic agents. The peri-surgical period involves other processes that may also contribute: stress response, neuroinflammation, pain, opioids (Borsook et al., 2010). Post Anesthetic Status: The complex interaction of the factors in the pre-anesthetic and anesthetic status could partly explain why brain functions and behavior are transiently altered in some patients, while in others, or in more vulnerable population (unstable brain circuits), such alterations may persist.

has been associated with higher pain intensity ratings at 7.5 years of age (Valeri et al., 2016). Persistent changes on the nociceptive system after early pain and stress experiences in animals and humans have been reviewed elsewhere (Lidow, 2002; Loepke and Soriano, 2012; Walker et al., 2016) (Tables 4 and 5). 3.4.3.2. Aging brain. Pain increases with age and thus the population exposed to anesthesia for surgery may have pain as a major comorbidity that contributes to a ‘pre-sensitized state’ for the potential effects of anesthesia. In aged rats (24–25 months) exposed to laparotomy and isoflurane anesthesia (1.2%) for 2 h, postoperative pain has been suggested to contribute to the development of memory deficits following the surgery and anesthesia via up-regulation of hippocampal NMDA receptors (Chi et al., 2013). Similarly, acute nociception following a surgical incision in the hindpaw of adult mice is linked to age-dependent hippocampus-independent fearing learning impairment (Zhang et al., 2013). In humans, postoperative pain is a potential contributor to the development of POCD. In one study, for example, patients of 65 years of age or older were tested for POCD within 48 h after a noncardiac surgery: postoperative pain was associated to the risk for the onset of POCD (Wang et al., 2007). Similarly, the risk of early cognitive dysfunction after joint arthroplasty may relate to the efficacy of pain control (Zywiel et al., 2014) (Tables 4 and 7). 3.4.4. Opioids Opioid analgesics are routinely administered to provide analgesia and reduce dose requirements for anesthetic and sedative agents, they could mitigate anesthesia-induced brain neurotoxicity (Ward and Eckenhoff, 2016; Ward and Loepke, 2012).

However, rather than mitigating anesthesia-induced neurotoxicity, recent data has implicated opioids as causing significant effects on brain structure and function in human adult (Lin et al., 2015; Upadhyay et al., 2010; Younger et al., 2011) and to have deleterious effects on the developing brain (Ranger et al., 2014). Opioid use may also result in postoperative hyperalgesia (Kim et al., 2014). Moreover, in-vivo and in-vitro animal studies have shown effects of these drugs on dendritic spine synaptic plasticity (Liao et al., 2005; Miller et al., 2012; Robinson and Kolb, 1999) and development of neuroinflammation (Wang et al., 2012b) with subsequent changes in adult and neonate brain plasticity (BeltránCampos et al., 2015). Prenatal and neonatal exposure of rodents to morphine has been shown to alter cognitive functions later in life (Boasen et al., 2009; Niu et al., 2009; Šlamberová et al., 2001) (Table 1). 3.5. Neuroprotective factors The concomitant administration of anesthetics, sedatives, other drugs (lithium, dexmedetomidine, xenon), or natural hormones (estradiol, melatonin), or maintaining the whole body in hypothermia may mitigate anesthesia-induced neurodegeneration in animals (for review see Bajwa et al., 2015; Loepke and Soriano, 2012). Dexmedetomidine, a selective alpha-2-adrenergic agonist, has been found, for example, to decrease neurotoxicity in neonatal rats exposed to isoflurane or ketamine (Duan et al., 2014; Sanders et al., 2009, 2010). However, a recent study found that high cumulative doses and concentrations of dexmedetomidine induce neuroapoptosis in pup rats and fetuses (Liu et al., 2016). In the aging rat brains, local anesthetic blockade with lidocaine during the isoflurane exposure attenuated the deleterious effect (viz.,


135

Table 4 Secondary processes: Neuroinflammation and Pain. Anesthetic + Mechanisms of Species Action


Brain neurotoxicity

Isoflurane GABAA – agonist

Rats (18 months) and Mice (10 weeks)

1.2% (rats) or 1.4% (mice) for 2 h

Rats: impairment of acquisition/learning information Hippocampus, Cao et al. cerebral cortex (2012) but not the recall, neuroinflammation (increased level of IL-1b, IL-6), caspase 3 activation. Mice: neuroinflammation (increased level of IL-1b), cognitive deficits (in wild type mice but not IL-1b deficient mice)

Isoflurane

Rats (24–25 months)

1.2% for 2 h, with or without laparotomy, with or without analgesia

Postoperative memory deficits in the anesthesia + surgery group but not in anesthesia alone or with analgesia; higher level of NMDA receptor 2 subunits in the hippocampus in anesthesia + surgery group

Isoflurane

Mice (5–8 months) Wild mice/AD transgenic mice

1.4% isoflurane for 2 h

Increased levels of proinflammatory cytokines TNF-a, IL-6, and IL-b in brain tissues of both type of mice

Wu et al. (2012)

Isoflurane

Primary 2% isoflurane for 6 h neurons + glial cells of mice (1 or 15 d)

Increased levels of proinflammatory cytokines TNF-a in primary neurons

Wu et al. (2012)

Isoflurane

Mice (3 and 9 months)

1.4% – 3 min for a surgical incision in Older mice: At 3 and 7 d (but not at 30 d) after the the hindpaw surgery, hippocampus-independent fearing learning impairment. Local anesthetic cream at the place of incision attenuated the effect.

Isoflurane + buprenorphine

Rats (20 months)

Right carotid artery exposure + 3%, follow by 1.8% for 2 h


Naïve mice and 2.1% or 3% for 2 or 6 h AD transgenic mice (6 d)


Rats (10 d or 20 40 mg/kg d) for 6 h

Propofol + buprenorphine

Brain Region

Hippocampus

Reference

Chi et al. (2013)

Zhang et al. (2013)

Cerebral Neuroinflammation, learning and memory impairments 2 weeks after surgery. Equivalent effect cortex, hippocampus than with propofol

Zhang et al. (2015)

Increased caspase activation and apoptosis, altered amyloid precursor protein processing (APP) and increased b-amyloid protein level after 6 h exposure. More deleterious effect on AD mice than naïve mice. Increased of TNF-a in brain tissues of AD transgenic mice.

Lu et al. (2010)

each hour

Prefrontal In 10 d-old, transient increased of TNF mRNA cortex and expression in prefrontal cortex and transient increased of chemokine mRNA expression pattern hippocampus Ccl2 and Ccl3 in prefrontal cortex and hippocampus

Kargaran et al. (2015)

Rats (20 months)

Right carotid artery exposure + 15 mg/kg 1, followed by 1 mg/kg 1 min 1 for 2 h

Neuroinflammation, impaired learning and memory Cerebral cortex, 2 weeks after surgery. Equivalent effect than with hippocampus isoflurane

Zhang et al. (2015)

Propofol + Sevoflurane

Human neuroglioma H4 cells

100 nM + 4.1% for 6 h or propofol/ sevoflurane alone

Attenuation of apoptosis, AB accumulation and inflammation induced by sevoflurane when propofol is added

Tian et al. (2015)


Rats (1 d, 7 d, 14 d)

Single or repeated inflammatory pain (formalin injection in the forepaw) with or without ketamine (2.5 mg/kg 2)

Enhanced neuroapoptosis (attenuated by ketamine injection). At adulthood, short and long term memory alterations and increased pain latencies (attenuated by ketamine)

Anand et al. (2007)

Ketamine

Rats (7 d)

5 2 mg/ml each 90 for 6 h with or without peripheral noxious stimulation (CFA)

Increased neuroapoptosis with ketamine alone, attenuated when concurrent noxious stimulation was applied.

Liu et al. (2012b)


Rats (7 d)

0.75% + 70% for 6 h alone/with or without formalin/with or without surgery incision

Anesthesia alone: significant neuroapoptosis, longterm deleterious effect on cognition. Effects accentuated with nociceptive stimulation and surgery + increased IL-1b in the cortex 6 h later

1

+ 20 mg/kg

1

Cortex, hippocampus, thalamus, spinal cord.

Shu et al. (2012)

Repeated pain (needle prick Rats (from 0 d vs cotton-tipped swab) to 7 d)

Needle prick vs cotton tip rub were Repetitive pain as neonate may lead to an altered applied 1, 2, or 4 times/day from 0 d development of pain system (e.g. decreased pain to 7 d. thresholds)

Anand et al. (1999)

Peripheral inflammatory pain

Single injection of CFA at 1 and 14 d Age-dependent hyperalgesia at adult age (left hindpaw) + same injection at 12 weeks of age

Blom et al. (2006)

Mice (1 d and 14 d)

136


Table 4 (Continued) Anesthetic + Mechanisms of Species Action Peripheral inflammatory pain


Rats (0, 1, 3, or Single injection of CFA in the left 14 d) hindpaw in neonates

Brain neurotoxicity Long-lasting consequences of peripheral inflammation on the nociceptive pathways assessed in adult life (at 8 to 12 weeks old)

Brain Region

Reference Ruda et al. (2000)

Key: mg = milligram; kg = kilogram; d = day, CFA = Complete Freund’s adjuvant.

cognitive impairment) of isoflurane on the brain (Lin et al., 2012). Similar findings have been found with the pretreatment of animals with minocycline (Kong et al., 2013) or GTS-21 (a selective agonist of alpha 7 nicotinic acetylcholine receptor) (Kong et al., 2015), before isoflurane exposure (Tables 1 and 6). 4. Anesthetic effects across different age groups Recent electroencephalographic studies suggest distinct brain responses in different age groups to the same anesthetic agents (Akeju et al., 2014; Cornelissen et al., 2015; Purdon et al., 2015a) and across different anesthetics (Akeju et al., 2015; Purdon et al., 2015b). Such changes reflect the underlying effects of drugs and the state of brain development or function (sensitivity and response to the drug). Below we review anesthetic-induced changes in young children and adults, with a focus on integration of preclinical and clinical research. 4.1. Anesthesia in children: preclinical and clinical evidence of altered brain biology Studies have indicated potential neurotoxicity in young animals (Table 1) and children (Table 5) and the topic has been reviewed extensively (Brambrink et al., 2012b; Davidson et al., 2015; Lin et al., 2014; Loepke and Soriano, 2008, 2012; McCann and Soriano, 2012). While ongoing questions exist as to the translation of animal studies to the humans, the issue of alteration in brain systems (observed as changes in behavioral state) continues to be a problem because of lack of sensitive measures (in those cases that are not obvious). Prior brain states clearly contribute to the anesthetic itself (e.g., more anxious individuals may require increased levels of anesthesia). Here we review both animal and human data in the context of alterations of behavior. 4.1.1. Behavioral changes in animal models Animal models provide a way to evaluate the independent effects of anesthesia and surgery. Anesthetic agents can cause brain injury, affecting brain systems and development. Deleterious effects that have been shown in animal studies included learning impairment and memory (Table 1), and abnormal social behaviors that can resemble autism spectrum disorder (Satomoto et al., 2009). The behavioral changes may be related to a number of factors including: 4.1.1.1. Age. The most sensitive period for anesthetic exposure induced behavioral changes appear to be during early development, as differences across age have been observed in rodent models (Yon et al., 2005). Maternal anesthesia exposure also affects behavior in fetal and newborn rats. Indeed, in the study of Ronca et al., prenatal maternal exposure to ether (an older volatile anesthetic no longer available for administration in humans) more than isoflurane resulted in fewer spontaneous fetal movements and first postpartum nipple attachments (Ronca et al., 2007).

4.1.1.2. Drug type, dose, and combination. Drug dosing in anesthesia relates to the dose itself, the duration of exposure, and the number of exposures. Many anesthetic drugs are known to act on both GABAA and NMDA receptors (Campagna et al., 2003). Exposure of young animals during brain development to predominantly GABAA agonists like benzodiazepines, barbiturates, propofol, and volatile agents have led to deleterious effects on brain structure and neurological functions (for review see: Istaphanous et al., 2010; Loepke and Soriano, 2008, 2012). In some studies of volatile anesthetic agents, desflurane reportedly has more immediate deleterious effects than isoflurane than sevoflurane (Kodama et al., 2011; Liang et al., 2010; Ramage et al., 2013). In 6-day-old mice exposed to equivalent doses of desflurane, isoflurane, or sevoflurane, desflurane was associated with a greater deficit in working and long-term memory (Kodama et al., 2011). Exposure to isoflurane triggers greater levels of neurodegeneration (apoptosis) than sevoflurane in neonatal mice (Liang et al., 2010) and greater memory deficits in rats (Ramage et al., 2013). However, Istaphanous et al. (2011) found a similar extent and distribution of neuroapoptosis in neonatal mice exposed for 6 h to isoflurane, desflurane, or sevoflurane. In contrast, xenon has some neuroprotective qualities in the immature brain (Ward and Loepke, 2012): no increase in neuroapoptosis was found in mice exposed to xenon alone or isoflurane and xenon for 6 h (Ma et al., 2007). However, in a more recent in-vitro study, Brosnan and Bickler (2013) found neurotoxicity in hippocampal slice cultures from 7-day-old rats exposed to xenon. Similarly, Cattano et al. (2008a) reported neuroapoptosis in neonate mice following exposure to xenon. Interestingly, when administered with isoflurane, a decrease in isoflurane-induced neuronal apoptosis was observed. Among intravenous anesthetic agents, low dose midazolam (3– 9 mg/kg) in neonatal rats did not lead to increased neuroapoptosis (Jevtovic-Todorovic et al., 2003b; Yon et al., 2005), although with similar doses neuronal apoptosis was found in neonatal mice (Young et al., 2005). These results suggest differences in vulnerability among species. Exposure to propofol has also shown mixed results with a dose-dependent effect (Bercker et al., 2009; Cattano et al., 2008b; Fredriksson et al., 2007; Yu et al., 2013). Repeated exposure to anesthetic agents may have a more deleterious effect on brain systems, and can vary between agents. Memory impairment with repeated exposure to isoflurane has been found in rodents (Zhong et al., 2015; Zhu et al., 2010). Multiple exposures of neonatal mice to isoflurane, but not desflurane, led to impairment in spatial learning and memory at about 31 days of age (Tao et al., 2016). With propofol, the reported findings are mixed. Single or repeated exposures to propofol has resulted in significant neuroapoptosis in the hippocampal region in pup rats but persistent learning/memory deficits were only found following repeated exposures (Yu et al., 2013). Significant neurodegeneration and persistent learning deficits have also been found with a cumulative dose of 90 mg/kg over 4.5 h in pups rats (Bercker et al., 2009). In contrast, with repeated injections of propofol over 3 h in the same species, Karen et al. (2013) found significant neuronal degeneration but no persistent learning deficits. Multiple doses of


137

Table 5 Anesthesia and Neurotoxicity in Children. Study Design

Study group


Control group

Brain neurotoxicity

Retrospective cohort study

From a cross-sectional MRI database with 5 to 18 y children

Surgery with anesthesia before 4 y (documented in medical record)

Unexposed matched children (no evidence of anesthesia in their medical record)

Backeljauw Before 18 y, exposed group has lower listening comprehension and et al. (2015) performance IQ associated with lower gray matter density in occipital cortex and cerebellum. 75% of children were exposed only once

Prospective study

Children from 1.5 to 5 y Already exposed to 2 surgeries under GA and planned for a third

Healthy matched children attended vaccination clinic

Risk to become depressed, anxious, Bakri et al. and to have sleep and attentional (2015) problem (ADHD) in the weeks postsurgery in exposed group


Young-Netherlands Twin Register of twin born between 1986 and 1995.

Anesthesia exposure reported by parents before 3 y and before 12 y

Unexposed monozygotic twin pairs and unexposed co-twin

At 12 y, no difference in learning performance (achievement tests, teacher ratings) in comparison to unexposed co-twins.

Retrospective cohortstudy

Birth databases in Taiwan between 2004 and 2007

Cesarean delivery under GA or RA

Vaginal delivery

During a mean follow-up of 4.3 Chien et al. years, incidence ASD higher in (2015) cesarean delivery with GA than RA or vaginal delivery. But no difference between RA and vaginal delivery.


Swedish Medical Birth and National Patient Registers

Unassisted/assisted vaginal delivery, Sibling controls elective/emergency cesarean delivery

Elective/emergency cesarean Curran et al. delivery associated with ASD. But (2015) the association does not persist when compared to siblings controls

Prospective randomized controlled trial

International Planned for inguinal herniorrhaphy Children undergoing the same multicenter study with before 1 y under 1 h sevoflurane GA procedure with awake RA children < 60 weeks postmenstrual age when included

Davidson At 2 y, no difference between the et al. (2016) two groups in terms of cognitive development (assessed with Bayley Scale Infant and Toddler development III)


New-York State Medicaid with children born between 1999 and 2001

ICD-9 procedural code for Inguinal Hernia before 3 y

Children from the cohort matched for age, without procedural code for inguinal hernia (could had been exposed for other reasons)

DiMaggio At 4 y follow-up, higher risk of et al. (2009) behavioral or developmental disorders in the study group (ICD-9 code for unspecified delay or behavioral disorder, mental retardation, autism, language problem). Mean age: 30 month-old


New-York State Medicaid from 1999 to 2005.

ICD-9 procedural code for anesthesia/surgery before 3 y without code for preexisting behavioral or developmental diagnosis (68% with one exposure, 23% with two, 8% with 3 and more)

Siblings not exposed to surgery/ anesthesia before the age of 3 y and without recorded history of developmental or behavioral diagnosis before 10 months.

Before 4 y: higher risk for DiMaggio et al. (2011) developmental or behavioral disorders when at least 2 exposures (ICD-9 code for behavioral or learning abnormalities)

Prospective selfcontrolled study

Children from 4 to 7 y

Strabismus correction under sevoflurane GA


Mayo cohort with children born between 1976 and 1982

Surgery/anesthesia before 2 y

Retrospective matchedcohortstudy

Manitoba Population Health Research Data Repository & Early Development Instrument (EDI)

Single or multiple exposures to GA Matched unexposed controls from Children exposed between 2 and 4 before 4 years of age. Completion of the same database years of age have significant EDI between 5 and 6 years of age decreased performance in EDI results (mostly communication/ general knowledge, language/ cognition). No greater deficit was found for multiple exposure

Graham et al. (2016)


Danish birth cohort with Single exposure to inguinal hernia children born between repair before 1 y (identified with 1986 and 1990 ICD-8 code). Children with additional surgeries were not excluded

Aged-matched children from the cohort without exposure to inguinal hernia repair before 1 y. Children with additional surgeries were not excluded

No difference in academic performance at 15 or 16 y (general test of academic achievement and average teacher rating)

Hansen et al. (2011)


Danish birth cohort with Single exposure to surgery for Matched-children from the cohort No difference in academic children born between pyloric stenosis before 3 month-old without exposure to surgery for performance at 15 or 16 y (general 1986 and 1990 (identified with ICD-8 code). pyloric stenosis before 3-month

Hansen et al. (2013)

Reference

Bartels et al. (2009)

Fan et al. WPPSI-III before, 1 month and 6 months post-surgery: no difference (2013) between the 3 measurements

Matched-children from the cohort Increased risk for learning deficits without exposure with multiple exposures but not single exposure (evaluation before 19 y using school records)

Flick et al. (2011)

138


Table 5 (Continued) Study Design

Study group


Control group

Brain neurotoxicity

Children with additional surgeries were not excluded

old. Children with additional surgeries were not excluded

test of academic achievement and average teacher rating)

Reference


49 children born between 1975 and 1984 with developmental disorder

Children born under GA (hospital A) vs. non-anesthetics deliveries (hospital B and C)


Raine cohort with children born between 1989 and 1992

ICD-9 code for anesthesia/surgery before 3 y

Children from the cohort without Higher risk of language and abstract Ing et al. ICD-9 code for anesthesia/surgery reasoning deficits at the age of 10 y (2012) before 3 y than controls with single or multiple exposures


Raine cohort with children born between 1989 and 1992

ICD-9 code for anesthesia/surgery before 3 y

Children from the cohort without At 10 y, increased risk of ICD-9 code for anesthesia/surgery neurodevelopmental outcomes before 3 y when assessed with neuropsychological testing or the ICD-9 codes in the exposed group, but no difference on academic achievement scores.

Ing et al. (2014)

Randomized, doubleblind controlled study

Children from 2 to 7 y scheduled for surgery

Oral premedication midazolam (0.5 mg/kg)

Oral premedication placebo (acetaminophen 15 mg/kg)

Fewer negative behavioral changes when premedicated with midazolam one week postoperatively but not at 2 weeks after

Kain et al. (1999a)

Longitudinal study

Children from 1 to 7 y scheduled for surgery

GA for surgery: otolaryngology (34%), minor general surgery (13%), lower genitourinary procedures (11%)

Cohort divided in 3 groups: low, medium, high anxiety at anesthesia induction (Modified Yale preoperative Anxiety Scale)

High level of anxiety during induction is associated with more negative behavioral changes postoperatively. The frequency of negative behavioral changes decreased with time after surgery and is influenced by the surgical procedure.

Kain et al. (1999b)

Longitudinal prospective randomized controlled trial

120 children planned for Daycase surgery under sevoflurane a daycase surgical or halothane GA (without previous procedure exposure to GA)


National Health insurance research database in Taiwan

Prospective study

Children from 6 months Myringotomy surgery under to 6 y halothane or sevoflurane, with or without midazolam premedication

Prospective randomized study



Population-database Single or multiple exposures to GA Matched unexposed control from Children > 2 years of age at the time O’Leary from Institute for et al. (2016) before EDI completion at 5 to 6 years the same database of first surgery have an increased Clinical Evaluative of age. odds of early developmental Sciences, Canada & Early vulnerability. The risk is not Development increased by the frequency of Instrument (EDI) exposure.



Unexposed-children from the Single or multiple surgery/ anesthesia before 2 y (via the cohort same cohort records)

Increased risk of ADHD at 19 y when Sprung et al. (2012) exposed to multiple surgery and anesthesia

Prospective study


GA for any procedure

Stargatt Significant behavioral changes in 24% patients 3 d post-exposure and et al. (2006) in 16% at 30 d post-exposure. When compared with sibling controls: significantly more changes at 3 d but not at 30 d.

More cases of children with ASD in hospital A using anesthetics for deliveries

Sevoflurane group more distressed Keaney than halothane group in the et al. (2004) immediate postoperative period but no difference at long-term

Exposure to GA and surgery before 2 Matched unexposed control from No difference in the incidence of y (ICD-9 code) the same database ASD between the two groups.

Midazolam premedication 0.5mh/ kg-1, surgery under halothane and nitrous oxide

Hattori et al. (1991)

Ko et al. (2015)

Lapin et al. Faster recovery times for sevoflurane group. (1999) Sevoflurane without midazolam premedication leads to higher incidence of postoperative agitation in the recovery room Placebo premedication, surgery under halothane and nitrous oxide

Siblings controls at comparable age without exposure to GA

During induction, midazolam children were quieter, but with an increase of postoperative behavioral changes

McGraw and Kendrick (1998)

E. Colon et al. / Progress in Neurobiology 153 (2017) 121–160 Matchedcohort study

Sibling-matched cohort Sibling pair exposed vs. unexposed from 4 Universityto GA for inguinal hernia repair affiliated hospitals (USA) < 3 years of age

Retrospective study

Preterm-born children (24 to 32 weeks gestational age)

Neonatal pain: number of invasive procedures from birth to termequivalent age



Surgery before 4 y (via the cohort records)

Prospective randomized study

Healthy children from 9 to 11 y

Elective hernia surgery under GA with propofol or sevoflurane

Siblings unexposed

139 No difference in global cognitive functions (IQ) and specific neurocognitive functions assessed around 10 y.

Sun et al. (2016)

Valeri et al. Greater number of invasive procedures as neonates associated (2016) with higher pain intensity (selfratings) after blood collection at age 7.5 y Wilder et al. Children from the cohort without Significant risk for learning exposure disabilities when multiple (2009) exposures before 4 y (performance IQ and achievement test before 19 y) Yin et al. Wechsler Memory Scale before, 7 days, and 3 months post-surgery: (2014) short-term memory impaired in the propofol group 7 days post surgery but recovery at 3 months

Key: GA = general anesthesia, RA = regional anesthesia, ASD = Autism Spectrum Disorder, y = year.

ketamine up to 75 mg/kg led to deleterious effects on the structure of the brain of neonatal rodents, whereas with a single dose many studies did not find significant effects (Table 1). Similar findings were found in a rhesus monkey model with a time-dependent effect (Paule et al., 2011; Slikker et al., 2007; Zou et al., 2009). Earlier stages of development in rhesus monkeys (122 days of gestation or post-natal day 5 and 6) appear to be more sensitive to ketamine-induced neuronal cell death than later stages (post-natal day 35) (Slikker et al., 2007). Moreover, less than 3 h exposure to ketamine in newborn appeared not enough to produce significant neurotoxic brain changes, while exposure up to 9 or 24 h produced significant cell deaths in the frontal cortex (Zou et al., 2009), and 24 h exposure during this sensitive period led to cognitive impairments more than 2 to 3 years later (Paule et al., 2011). Combination of anesthetic agents may result in more significant brain changes. The combined administration of midazolam, isoflurane, and nitrous oxide at a dose sufficient to maintain a surgical plane of anesthesia for 6 h caused a significant increase in apoptotic degeneration in the newborn rat brain (JevtovicTodorovic et al., 2003b; Lu et al., 2006; Nikizad et al., 2007; Yon et al., 2005, 2006), as well as an attenuation in the long-term potentiation of the hippocampal circuits (Jevtovic-Todorovic et al., 2003b). A similar increase in the rate of brain cell degeneration has been shown, with dose-dependent effects, in rodents following exposure to ketamine and diazepam (Fredriksson et al., 2004), ketamine and thiopental, or ketamine and propofol (Fredriksson et al., 2007). When used alone, exposure to nitrous oxide triggers little or no neuronal apoptosis (Jevtovic-Todorovic et al., 2003b; Ma et al., 2007). However, when nitrous oxide is combined to isoflurane (Ma et al., 2007), or with isoflurane and midazolam (Jevtovic-Todorovic et al., 2003b), anesthesia-induced neuronal apoptosis is increased. Similar results were found in newborn rhesus monkeys. Combined isoflurane and nitrous oxide exposure for 8 h led to significant neuroapoptosis in the frontal cortex, temporal gyrus, and hippocampus but not the exposure to one anesthetic at a time (Zou et al., 2011). Translation of animal data to humans (biological and behavioral) has made the utility of animal studies undergo more scrutiny. In addition to species differences, doses typically used in animal models are often higher than those routinely used in humans, even when differences in body size and metabolic rates among species are taken into account (Lin et al., 2014). It is not clear if the period of vulnerability between animals and humans is comparable, because development of the human brain does not follow the exact same pathway as the animal brain (Istaphanous et al., 2010). Nonetheless, insights into brain regions affected, as measured by apoptotic

and behavioral changes can be used to inform human studies (Psaty et al., 2015; Rappaport et al., 2015). 4.1.2. Behavioral changes in children The potential neurotoxicity of anesthetic agents on the developing brain is of great importance as millions of infants and preschool children receive general anesthesia every year to facilitate surgical and medical procedures (Reddy, 2012). As previously described, a large body of evidence from animal studies has demonstrated that early exposure to virtually all anesthetic agents used in clinical practice may affect brain development, with persistent deficits in learning and memory (Jevtovic-Todorovic et al., 2003b; Lee et al., 2015; Loepke and Soriano, 2008). In susceptible children (e.g., autism spectrum disorders), the postanesthetic effects seem more pronounced (e.g., behavioral regression), supporting the notion that these agents may indeed affect neural circuits in a profound manner and may provide a model of the effects of anesthesia in children and children with autism (Box 1). Despite mounting evidence of neurotoxic effects following exposure to anesthetic agents in animals, similar evidence in early childhood is still a matter of debate. Indeed, while some studies have found an association between exposure to general anesthetics and adverse long-term neurodevelopmental outcomes, others failed to reach this conclusion (for review see: Brambrink et al., 2012b; Lei et al., 2014; Lin et al., 2014; Loepke and Soriano, 2012). Moreover, ethical concerns have been raised regarding the conduct of studies in children necessary to reach definitive conclusions. For example, randomization of children to undergo painful procedures with or without anesthesia and analgesia is unacceptable. Furthermore, in comparison to animal studies, exposure to anesthetics in humans cannot be dissociated from a surgical or medical indication (Istaphanous et al., 2010). Until recently, most of the clinical evidence in children derives from case reports, retrospective postoperative behavioral studies (Flick et al., 2011; Wilder et al., 2009), or retrospective epidemiological analyses (Table 5). Consequently, in most studies, factors such as underlying pathologies, perioperative factors, surgery, and secondary processes (stress response, pain, neuroinflammation, opioids) confound any inference of causality. Data to support alterations in behaviors (a surrogate for brain changes) following general anesthesia in children is summarized below. 4.1.2.1. Deficits in cognition. Many studies have shown that early exposure to general anesthesia is associated with an increased risk

140


of developing learning and behavioral disabilities later in life (Table 5). In children exposed to anesthesia before 4 years of age, subtle structural abnormalities, mainly in the cerebellum, but alsoin the occipital cortex and right frontal lobe, have been associated with a decrease in performance IQ and language comprehension later in life (Backeljauw et al., 2015). Moreover, recent epidemiological analyses of large data sets from two separate Canadian registries examined the impact of anesthesia exposure on educational domains. In the two studies, the Educational Development Instrument (EDI) was compared in a populationbased cohort between children receiving surgery before 5 years of age and naïve cohorts. In one of the study, children greater than two years had slim but increased odds of developmental vulnerability (O’Leary et al., 2016). The second retrospective cohort study from Manitoba examined the impact of anesthesia exposure before 2 years or between 2 and 4 years of age. This report compared 3850 exposed children with 13,586 naïve controls. They also found increased vulnerability in patients exposed to anesthesia at the age of 2 to 4 years with no greater risk for multiple than single exposure (Graham et al., 2016). However, some studies have found no differences in cognitive development and academic performance following exposure to anesthesia (Bartels et al., 2009; Davidson et al., 2016; Fan et al., 2013; Hansen et al., 2011, 2013) (Table 5). 4.1.2.2. Emotional and behavioral disturbance. General anesthesia has been linked to an increased risk for attention deficit hyperactivity disorder (ADHD) later in life in children exposed multiple time before 2 years of age (Sprung et al., 2012), or to a higher risk of developing anxiety, depression, behavioral changes, sleep difficulties, and attentional problems (Kain et al., 1999a,b; Stargatt et al., 2006). The observed behavioral changes are dependent on a number of factors: 4.1.2.3. Repeated exposures. Multiple early exposures to general anesthesia may be more deleterious than a single exposure, because repeated exposures have been more often linked to subsequent learning disabilities and emotional disturbances (Bakri et al., 2015; Flick et al., 2011; Ing et al., 2012; Sprung et al., 2012; Wilder et al., 2009). Such findings are corroborated by the results of a recent meta-analysis suggesting that undergoing a surgical procedure with anesthesia before the age of 4 years may be associated with later neurodevelopmental deficits (cognitive and behavioral). The number of exposures rather than the time at exposure before 4 years of age may be the more significant factor (Wang et al., 2014). Accordingly, a single exposure to general anesthesia before the age of 1 year for inguinal herniorrhaphy (Hansen et al., 2011) or before 3 months of age for pyloromyotomy (Hansen et al., 2013) was not associated with subsequent differences in academic performance in comparison to unexposed children. These results are in line with the recently published PANDA trial that analyzed the impact of a single general anesthesia during inguinal hernia surgery before the age of 3 years on exposed and naïve sibling pairs. The investigators reported no difference in mean IQ scores or in performance in specific neuropsychological domains (e.g. memory, language, motor functions, attention) between the sibling pairs (Sun et al., 2016). Nevertheless, in some studies, single exposure has been associated with developmental or behavioral disorders (Backeljauw et al., 2015; DiMaggio et al., 2009; Ing et al., 2012). 4.1.2.4. Drug type. The anesthetic drug may produce different effects, as evidenced by the following examples. Sevoflurane and halothane have been associated with emergence agitation and negative behavioral changes (NBCs). Although children

anesthetized with sevoflurane were more distressed on emergence from anesthesia, there was no difference in NBCs at 1, 7, and 30 days postoperatively (Keaney et al., 2004). Emergence agitation was greater with sevoflurane than halothane (Lapin et al., 1999), but postoperative agitation was reduced when children who received sevoflurane were premedicated with midazolam. Fewer negative behavioral changes were also found within the first postoperative week with midazolam premedication (Kain et al., 1999a). However, increased NBCs in children after midazolam premedication have also been found (McGraw and Kendrick, 1998). Acute short-term memory impairment has been found in young children after anesthesia with propofol but not sevoflurane (Yin et al., 2014). 4.1.3. Evaluating the pediatric data: potential confounding factors An important consideration in evaluating an association between early exposure to anesthesia and neurodevelopmental outcome is the study design and measures used to assess learning and cognitive abilities (Beers et al., 2014; Block et al., 2012; Flick and Warner, 2012). The majority of studies are observational with evaluation of the association between anesthetic exposure and cognitive outcomes using indirect measures such as tests of academic achievement or International Classification of Disease codes (ICD-9). Objective evaluation of neurodevelopment outcomes requires the use of standardized neuropsychological instruments (Beers et al., 2014; Sun, 2010). Therefore, the use of different outcomes measured across studies may explain in part the divergent results. The association between early exposure to anesthesia and neurocognitive disorders later in life does depend on the outcome measures used (Ing et al., 2014). Ing et al. (2014) evaluated cognitive function at the age of 10 years in children exposed to anesthesia before the age of 3 years with three different measures: neuropsychological testing, ICD-9 codes, and tests of academic achievement, and found an increased risk of cognitive deficits with the neuropsychological testing and ICD-9 codes but not with the achievement scores. Another issue is that the tools available for assessment of neurocognitive functions in young children have a relatively weak predictive value for subsequent assessments and long-term outcome (Lei et al., 2014). An important limitation of many human studies is their nonrandomized design which limits the ability to differentiate neurotoxicity caused by anesthetics from the effects of surgery (stress, pain, or inflammation), which are also potent modulators of brain development (Istaphanous et al., 2010). The condition that necessitated anesthesia, hospitalization, coexisting diseases, and genetic predispositions are also potential confounding factors (Lei et al., 2014). Moreover, in most of these studies, information regarding conduct and duration of anesthesia, anesthetic drugs administered, and surgical procedures were not accounted for. Consequently, the evidence in favor of neurocognitive alterations (cognitive and learning deficits) cannot be causally linked to the exposure to anesthesia, and longitudinal randomized controlled studies are needed. The first randomized controlled trial to evaluate the effect of general anesthesia in infancy on neurodevelopmental outcome is ongoing (Davidson et al., 2016). In this multicenter trial, infants 75%, no difference between males and females

Retrosplenial cortex

JevtovicTodorovic et al. (2001)

Nitrous Oxide

Rats (280–320 gr)

From 0 to 150-Vol% for 4 to 16 h

Short exposure to 150-Vol% causes reversible vacuole reaction. Prolonged exposure ( 8 h) causes cell death reaction.

Posterior cingulate, retrosplenial cortex

JevtovicTodorovic et al. (2003a)

Nitrous Oxide

Rats (6, 18, or 24 From 0 to 175% for 3 Dose-dependent neurotoxic reactions equivalent in young and months) h aging rats


JevtovicTodorovic and Carter (2005)



35 mg/kg for 3 h

Propofol

Mice (8–10 weeks)

Hippocampus 100 mg/kg first and Impairment of survival and maturation of adult-born 5 50 mg/kg/h for 6 hippocampal neurons of 17 day-old at the time of anesthesia but h not 11 day-old.

Propofol

Rats (18 months)

Average 0.6 0.1 mg/kg min 1 for 2 h

Propofol

Rats (17 months)



Brain neurotoxicity

Brain Region

3 month-old: decreased number of maturing neurons in nascent Dentate gyrus cells 24 h post-exposure + increased number of astrocytes. 20 month-old: no effect

Reference


Krzisch et al. (2013)

No lasting impairment in spatial memory task

Lee et al. (2008)

0.6 0.1 mg/kg1/ min 1

No cognitive impairment

Wang et al. (2012a)

Rats (320 g)

From 0 to 80 mg/ kg 1 for 3 h

Dose-dependent neurotoxic brain reactions



Ketamine

Rats (8 d and 1, 2, 3, or 8 months)

From 0 to 80 mg/ kg 1

Age-dependent increase in vacuolization of neurons in females Retrosplenial cortex at lower doses and increased reaction at each succeeding doses. Increase in vacuolization of neurons in males mainly when exposed at 8 month-old


Ketamine

Rats (6, 18, or 24 From 20 to 100 mg/ months) kg 1

Dose-dependent neurotoxic effect. Aging animal more sensitive Posterior cingulate, than young retrosplenial cortex




1.2% + 70% for 2 h

Impaired spatial learning in 18 months rats 8 weeks postexposure, but improved performance in 6 months rats



Rats (18 months)

1.2% + 70% for 2 h

Impaired spatial memory at least 2 weeks after exposure

Culley et al. (2004a)

1

/


143

Table 6 (Continued) Anesthetic

Species


Brain neurotoxicity



1.2% + 70% for 2 h

Impaired learning performance


Rats (18 months)

1.2% + 70% for 4 h

Persistent changes in hippocampal gene expression (2 d postexposure)

Analyses in Hippocampus


Ketamine + Nitrous Oxide

Rats (320 g)

From 0 to 80 mg/ kg 1 + 0 to 120% for 3h

Enhanced dose-dependent neurotoxic brain reaction (in comparison to each agent alone)



Ketamine + Nitrous Oxide

Rats (6, 18, or 24 From 0 to 30 mg/ months) kg + 50 150% for 3h

Increased neurotoxic reaction. At low doses, higher sensitivity in aging animals




1.8% + 70% for 4 h

Hippocampus, 18 months: spatial learning impairment at 3 months postanesthesia + reduction in phosphor-ERK1/2 and increased NR2 B cortex protein expression in hippocampus and cortex.

Mawhinney et al. (2012)

Propofol or Pentobarbital

Mice (18 months)

Impaired cognitive performance and hyperphosphorylation of 50 mg/kg or 100 mg/kg. tau. Partial recovery of cognitive impairment and no tau Repeated (same hyperphosphorylation when body temperature was controlled dose twice a week/2 weeks

Fentanyl + Droperidol Rats (90 d)

20 mg/kg + 500 mg/ kg for splenectomy or no surgery

Brain Region

Rats exposed to anesthesia + surgery: transient impairment in spatial learning, glial activation, increased proinflammatory cytokines

Reference Culley et al. (2004b)

Xiao et al. (2013)

Hippocampus

Wan et al. (2007)

Key: mg = milligram; kg = kilogram; d = day.

humans and the potential link with surgery/anesthesia are still highly debated (Avidan and Evers, 2016), and experimental preclinical studies have tried to better understand the underlying pathophysiology. In the following sections, we review the preclinical and clinical evidence of brain changes following general anesthesia in adult and old animals and the elderly. 4.2.1. Behavioral changes in animal models Animal models used to evaluate the behavioral effects of anesthesia and surgery. Such studies have provided support for the notion that anesthetic agents may have adverse effects in a dosedependent manner on older brains. In addition such effects have the potential to impair cognitive functions over the long-term (Bittner et al., 2011; Brambrink et al., 2012b; Hudson and Hemmings, 2011). Presumably anesthetic may produce differences in behaviors based on a number of factors including sensitivity to anesthesia, age and co-morbidities such as pain (see discussion below and section 3.4) (Tables 3, 4 and 6). Data to support alterations in brain and behaviors following general anesthesia in adult/old animals is summarized below. 4.2.1.1. Drug type, dose, and combination. The effect of exposure to general anesthesia in adult/old animals may depend on the anesthetics, the dose used, and gender. For example, exposure to nitrous oxide and/or ketamine at clinically relevant concentrations produces vacuolization in the cerebrocortical neurons of adult and old rodents (Jevtovic-Todorovic et al., 2000, 2001, 2003a; JevtovicTodorovic and Carter, 2005), but the combination of ketamine and nitrous oxide has led to significant increased brain neurotoxicity in comparison to the use of a single agent (Jevtovic-Todorovic et al., 2000; Jevtovic-Todorovic and Carter, 2005). Studies have also shown that exposure to isoflurane or desflurane resulted in dosedependent brain alterations. For example, exposure to 1% of isoflurane, but not 1.5% or 2%, led to transient spatial learning impairment in 2–3 month-old mice (Valentim et al., 2010). Moreover, exposure to 1.5 but not 1 MAC desflurane for 4 h impaired the acquisition learning and memory of old rats 1 week after the exposure (Callaway et al., 2015). Moreover, sensitivity to

ketamine may be dependent on gender as significantly more vacuolization of neurons in the retrosplenial cortex of females was found following exposure to ketamine in comparison to males (Jevtovic-Todorovic et al., 2001). 4.2.1.2. Behavioral manifestations. Many evidence from experimental animal studies have demonstrated impaired cognitive performance following exposure of adult/old animals to general anesthesia, but contrasting results have also been observed (see below). Impaired cognitive effects have been found across a number of anesthetics including isoflurane (Culley et al., 2004a; Ge et al., 2015; Kong et al., 2013, 2015; Lin et al., 2012; Lin and Zuo, 2011; Liu et al., 2012b; Ni et al., 2015; Wang et al., 2012a; Wang et al., 2015, 2016), the combination of isoflurane and nitrous oxide (Culley et al., 2003, 2004a,b; Mawhinney et al., 2012), desflurane (Callaway et al., 2015), and sevoflurane (Le Freche et al., 2012). By contrast, some studies have reported no effect on cognitive processes with propofol (Lee et al., 2008; Wang et al., 2012a) or isoflurane (Stratmann et al., 2010), and improvement of cognitive processes have been also observed following exposure of 2 month-old rats to isoflurane (Stratmann et al., 2009), or exposure of young adult (2–3 month-old) and old rats (20–24 month-old) to sevoflurane (Callaway et al., 2012a). Consequently the association between general anesthesia and alterations in cognitive processes may depend on the anesthetic used. Moreover, exposure to general anesthesia can also differently affect acquisition/learning and memory processes. For example, no effect of isoflurane exposure was observed on the acquisition phase of a spatial learning/ memory task, but a small memory deficit in younger adult rats (3 month-old) 1 week after exposure and a subtle deficit in long-term memory in old rats (12 month-old) 4 weeks after the exposure were reported (Callaway et al., 2012b). Similarly, an impairment of hippocampus-dependent learning and memory fear conditioning task as well as the recall/memory of a spatial memory task but not the acquisition phase were observed following exposure to 1.2% isoflurane for 2 h (Lin and Zuo, 2011). The reverse has been also observed with impairment of the acquisition/learning phase of information but not of the recall of learnt information (Cao et al.,

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2012). The factors responsible for differences in the results of these studies are not clear yet and may be related to the task used, the anesthetic exposure, the age, the time of cognitive testing (Lin and Zuo, 2011). More studies are then needed to better understand the association between exposure of adult/old animals to general anesthesia and potential cognitive alterations. 4.2.1.3. Age. Older brain could be more vulnerable to anesthesiainduced brain alterations than the younger adult brain and such susceptibility may be a factor in differences observed with aging and anesthesia. For example, a higher dose-dependent sensitivity to ketamine and nitrous oxide is observed in aged rats (18–24month-old) than in younger adults (6–8-month-old) (JevtovicTodorovic and Carter, 2005). The effect of exposure to anesthesia on cognitive processes may be also influenced by age. Indeed, exposure to desflurane resulted in spatial memory task impairment in aged rats (20–24 month-old) but not in younger adult rats (3 month-old) (Callaway et al., 2015). Similar agedependent findings on cognitive processes in rodents were observed following exposure to the combination of isoflurane and nitrous oxide (Culley et al., 2003) or to isoflurane (Wang et al., 2015). Contrasting results have also been observed: the combination of 1.2% isoflurane and 70% nitrous oxide for 2 h gave rise to similar long-term learning/memory deficit in old (20

month-old) and younger adult rats (6 month-old) (Culley et al., 2004b). Taken together, these results suggest the age-dependent effects of anesthesia exposure may depend on the anesthetic exposure (drug type, dose, duration and number of exposures). Finally, as discussed in section 3.1.2 and section 3.4.2.2, exposure to inhaled anesthetics also produces neuronal cell death (Ge et al., 2015; Kong et al., 2013, 2015; Lin and Zuo, 2011; Valentim et al., 2010; Xie et al., 2008), and neuroinflammation (Cao et al., 2012; Kong et al., 2013, 2015; Lin and Zuo, 2011; Tian et al., 2015; Zhang et al., 2015). Altered hippocampal expression for at least 48 h after exposure (Culley et al., 2006) and changes in cytosolic protein expression up to 72 h after exposure to 3 h of desflurane (Fütterer et al., 2004) have also been demonstrated. 4.2.2. Behavioral changes in the elderly In the elderly (>65 years), postoperative delirium (POD) and cognitive dysfunction (POCD) are common following surgery and general anesthesia. Although recovery is usually rapid, some patients report lasting cognitive impairment (e.g. poor concentration, memory disruption, information processing) weeks or months after the surgery (Bittner et al., 2011; Monk and Price, 2011). Postoperative delirium occurs in at least 20% of elderly individuals who are hospitalized each year (Monk and Price, 2011). POD may be the easily recognized hyperactive or agitated state, but

Box 2. Anesthesia and Alzheimer’s Disease – The Susceptible Brain Patient Group Alzheimer’s disease (AD) is the most common form of dementia, and is thought to be due to an interaction of genetic, environmental and still unknown risk factors (Eckenhoff and Laudansky, 2013; Seitz et al., 2013). Considering the neurotoxic effects of general anesthesia on the brain, the question of whether exposure to anesthetic agents is related to an earlier onset of AD or a higher risk of AD has been widely reported in the literature (Avidan and Evers, 2011; Seitz et al., 2013). Brain Changes in Alzheimer’s Disease AD is one of the most common neurodegenerative disorders and affects 20 to 30 million people worldwide. AD neuropathogenesis includes the generation and deposition of amyloid beta peptides and neurofibrillary tangle formation, and may also include neuroinflammation (Heneka and Obanion, 2007). Typically, AD begins with mild memory deficits that is followed in later stages by a profound dementia affecting multiple cognitive and behavioral spheres (Heneka and Obanion, 2007). Animal Evidence Animal studies have observed an increased of b-amyloid plaque formation, increased tau hyperphosphorylation, and impaired memory and learning following exposure to a large variety of anesthetic agents (for review see: Whittington et al., 2013; Xie and Xu, 2013). By contrast, in animal models of AD, some studies failed to demonstrate consistent cognitive impairment following exposure to general anesthesia (for review see: Vutskits and Xie, 2016). Moreover, clinically relevant concentrations of isoflurane may induce neuroinflammation by increasing levels of the pro-inflammatory cytokines TNF-a, IL-6, and IL-b, in the primary neurons and brain tissues of wild-type and AD transgenic mice (Wu et al., 2012). In the same way, exposure of 6-day-old AD transgenic mice to 3% sevoflurane for 6 h led to significant increase of TNF-a in brain tissues, caspase activation and apoptosis, alter amyloid precursor protein processing (APP) and increase b-amyloid protein level (Lu et al., 2010). Human Evidence Exposure of human and animals tissue cultures to isoflurane, sevoflurane, desflurane with hypoxia, or nitrous oxide and isoflurane for 6 h increases b-amyloid production and cellular apoptosis (Dong et al., 2009; Xie et al., 2006; Zhang et al., 2008; Zhen et al., 2009) (see Table 1). However, several of these studies were performed in in-vitro tumor cell lines (Hudson and Hemmings, 2011). In contrast, studies performed in elderly humans have been less conclusive. An inverse association between the age of AD onset and an increased number of exposures to anesthesia and surgery before the age of 50 years has been suggested, but the same group found no evidence for an increased incidence of AD with anesthesia and surgery (Bohnen et al., 1994a,b). An association between exposure to general anesthesia for coronary artery bypass graft (CABG) and an increased risk of AD has been found in patients older than 55 years, and not when the patients were exposed to a surgery without anesthesia (Lee et al., 2005). In contrast, the same group found no evidence when patients were exposed to a prostate or hernia surgery (Vanderweyde et al., 2010). Other studies found also no evidence for an increased risk of AD (Avidan et al., 2009) or cognitive alterations (Dijkstra et al., 1998) following exposure to general anesthesia and surgery. A recent meta-analysis performed on 15 case-control studies corroborates these negative results (Seitz et al., 2011). In contrast, changes in cerebrospinal fluid (CSF) tau and b-amyloid from a normal level before the surgery to a pattern observed in patients with mild cognitive impairment or with AD pattern have been found 6 months after cardiac surgery (Palotás et al., 2010) and 48 h after a nasal surgery (Tang et al., 2011). The relationship between general anesthesia and the risk of AD is then still not clear and has been the subject of recent reviews (Arora et al., 2014; Hussain et al., 2014; Yang and Fuh, 2015). Insight Patients with AD have altered brain structure and function and may be then more vulnerable to the potential effects of anesthesia.


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Table 7 Anesthesia and Neurotoxicity in the Elderly. Study Design & Groups


Retrospective cohort study:

3 groups of patients extracted from database: (1) Incidence of long-term POCD or trajectory of no surgery, no major illness, (2) exposure to dementia not affected by anesthesia or surgery, surgery, (3) major illness without surgery. but more important cognitive decline of demented than non demented participants

- Participants from the database of the Washington University AD research center with or without dementia at enrollment

Brain neurotoxicity

Reference Avidan et al. (2009)

Retrospective case control study

Patients who developed AD between 1975 and 1984 (Medical record Minnesota) + anesthesia record from Mayo Clinic. Compared to age and gender matched control

Prior cumulative exposure to GA is not risk factor Bohnen for AD. et al. (1994a)

Retrospective case control study

Patients who developed AD between 1975 and 1984 (Medical record Minnesota) + anesthesia record from Mayo Clinic.

Cumulative exposure to GA before 50 y may be associated with an earlier age of onset for AD.

Prospective, randomized, double-blind, controlled Elderly patients planned for major non-cardiac study: surgery with BIS-guided anesthesia (value between 40–60 during maintenance) vs. routine - Mean age of patients: BIS group (68.1.7 8.2 y); care. Compared with matched-control patients routine care group (67.6 8.3 y) without surgery during the same period

Bohnen et al. (1994b)

Significantly less patients with delirium in the BIS Chan et al. group after surgery. Similar cognitive (2013) performance in all group 1 week after surgery, but lower rate of PODC at 3 months in BIS group than in routine care group

Group 1: 3 or less operations under GA (less than Cognitive performances non predicted by history Dijkstra 3 h), Group 2: at least one operation under GA of operation under GA et al. (1998) lasting 3 h or at least 4 shorter episodes under GA, - Healthy subjects from patients register of general practitioner in the Netherlands from 24 Control: healthy subjects never operate under GA to 86 y

Retrospective cross-sectional study:

Retrospective study using Danish registry of 8503 Twin patients exposed to at least one major, Preoperative cognitive trajectory and underlying middle aged and elderly twins minor, hip, knee surgery, or other within 18 to 24 y disease could be more relevant for subsequent before a cognitive examination. Control group: cognition than surgery/anesthesia twins non exposed to surgery Prospective study: - Study group: patients > 50 y - Control group: patients > 55, with joint osteoarthritis (no planned surgery in the next 12 months)

Longitudinal cohort-study: - Mean age of patients: 76.7 5.2 y

Retrospective cohort-study: - Veterans Affairs patients > 55 y

Dokkedal et al. (2016)

Study group: Coronary angiography under Higher incidence of POCD in study groups sedation, major non cardiac surgery under GA, or independently from the type of surgery or cardiac surgery under GA anesthesia at 3 months post-surgery

Evered et al. (2011)

Patients planned for non-cardiac surgery without Accelerated trajectory of cognitive aging in evidence of dementia and compared to a surgical patients with postoperative delirium nonsurgical control group

Inouye et al. (2016)

Comparison between exposure to coronary artery Increased risk for the emergence of AD following Lee et al. bypass graft (CABG) surgery or a procedure CABG surgery rather than PTCA (2005) without anesthesia (percutaneous transluminal coronary angioplasty (PTCA))

Prospective longitudinal study

Patients exposed to major non-cardiac surgery (at At discharge, significantly more cognitive least 2 h): young (18–39 y), middle-aged (40–59 impairments in all aged patients than controls. y), elderly ( 60 y), age-matched control subjects 3 months post-surgery, significantly more POCD in elderly patients than in controls. Risk factors: increasing age, lower education, previous cva, POCD at discharge

Monk et al. (2008)

Longitudinal study:

Patients planned for a coronary artery bypass surgery

Gradual cognitive decline post-surgery and changes in cerebrospinal fluid similar to what is seen in AD at least 6 months post-surgery.

Palotás et al. (2010)

Major surgery (resection of an esophageal carcinoma) in 3 groups: sevoflurane anesthesia/ methylprednisolone before sevoflurane/ intravenous propofol for maintenance.

Increased risk of POCD in the sevoflurane group at Qiao et al. least 7 days post-procedure (2015)

- Mean age of patients: 64.23 5.19 y

Prospective double blind randomized controlled trial: - Patients aged 65–75 y

Randomized controlled trial:

Patients > 60 y planned for surgery ( 60 min) with BIS-guided anesthesia or blinded BIS - Mean age of patients: BIS group (69.7 6.3 y); monitoring. Compared with matched control Bis-blinded group (70.1 6.5 y) (age-MMSE)

Lower risk of delirium after surgery in the BISguided group

Radtke et al. (2013)

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Table 7 (Continued) Study Design & Groups


Brain neurotoxicity

Randomized international multicenter study:

RA vs. GA for major non-cardiac surgery

No difference in the incidence of POCD according Rasmussen to the type of anesthesia at 3 months post-surgery et al. (2003)

Patients randomized in percutaneous coronary intervention or off-pump coronary artery bypass grafting

Neuropsychological assessment 7.5 y later: no difference between groups.

Sauër et al. (2013)

Prevalence and mean days of delirium were significantly higher in deep sedation group compared to light sedation group.

Sieber et al. (2010)

Reference

- Patients > 60 y

Follow-up multicenter randomized trial of 280 patients ( 60 y)

Patients 65 y planned for hip fracture repair (spinal anesthesia with propofol) with deep (BIS - Mean age of patients: deep sedation (81.8 6.7 50) or light (BIS 80) sedation. y); light sedation (81.2 7.6 y)

Double-blind randomized controlled trial:

Longitudinal study: - Mean age of patients: 53 6 y

Single-site substudy of the BAG-RECALL multicenter study: - Mean age of patients: Bis group (62 14 y); ETAC (61 14 y)

Randomized controlled clinical trial:

Patients planned for an idiopathic nasal leak correction with total intravenous propofol, remifentanil, or inhaled sevoflurane (depending on provider choice)

Biomarkers for neuroinflammation and AD Tang et al. pathology increase in the cerebral fluid in the days (2011) after surgery. No difference according to the anesthetic agent.

Patients planned for cardiac or thoracic surgery with intraoperative BIS-guided or end-tidal anesthetic concentration-guided depth (ETAC) of anesthesia protocols

BIS-guided group has a significantly shorter stay in Whitlock ICU than ETAC group, and marginally less delirium et al. (2014) after surgery

Epidural vs. GA for orthopedic surgery

No difference in the incidence of POCD according Williamsto the type of anesthesia at 1 week and 6 months Russo et al. post-surgery (1995)

- Patients mean age = 69 y

Study with patients 65 y old or older (patients with evidence of delirium/dementia postoperatively were excluded)

Non-cardiac surgery (hip/knee replacement, other Cognitive tests before and 1 and 2 days Wang et al. orthopedic surgery, urologic, gynecologic, postoperatively: patients with oral postoperative (2007) vascular surgery) analgesia are less likely to experience POCD.

Key: GA = general anesthesia, RA = regional anesthesia, cva = cerebral vascular accident, AD = Alzheimer disease, ICU = intensive care unit, y = year.

more commonly is a hypoactive state of fatigue and a paucity of activity (Steinmetz and Rasmussen, 2016). A recent systematic review found that POD in the adult cardiac surgical population is associated with long-term increases in the risk of mortality, decreased cognition, functional decline, and health-related quality of life (Crocker et al., 2016). Similarly, older adults who experienced delirium after non-cardiac surgery had an accelerated trajectory of cognitive aging (Inouye et al., 2016). POCD is milder and subtler, long-lasting cognitive dysfunction that can reduce quality of life, increase stress of family members, and reduce survival (Bittner et al., 2011; van Harten et al., 2012). Approximately 20–30% of patients develop POCD within the first week after surgery regardless of age, but cognitive alterations more commonly persist in elderly patients (Monk and Price, 2011). The belief that surgery/general anesthesia could have a long-term deleterious effect on cognitive function in the elderly has given rise to the hypothesis that exposure to surgery/general anesthesia could be a risk factor in the development or exacerbation of neurodegenerative disease and in particular Alzheimer disease (Seitz et al., 2013) (Box 2). The mechanisms of POD and POCD have not been fully elucidated, and could be linked to the surgical procedure itself (cardiac and major procedures), general anesthesia, perioperative stress (inflammation, hypotension, hypoxemia), pain, medical comorbidities (risk factors for cerebrovascular disease), preoperative cognitive function, or a combination of these factors (Avidan and Evers, 2011; Krenk et al., 2010; Monk and Price, 2011; Steinmetz and Rasmussen, 2016; van Harten et al., 2012). However, advanced age and preoperative cognitive function appear to be major risk factors (Monk et al., 2008; Steinmetz and Rasmussen, 2016).

Significantly, several studies have suggested that the incidence of POCD is independent of the type of surgery or anesthesia, particularly at 3 to 6 months after exposure (Evered et al., 2011; Rasmussen et al., 2003; Williams-Russo et al., 1995). Evered et al. (2011) found a higher incidence after cardiac than non-cardiac surgery, and with general anesthesia over light sedation. However, these differences were no longer significant 3 months after surgery. Two other prospective studies have evaluated cognitive outcomes following exposure to regional or general anesthesia for a major surgery (Rasmussen et al., 2003; Williams-Russo et al., 1995). In both studies, the type of anesthesia had no impact on the development of POCD 6 months after the surgery. Studies comparing POCD following exposure to general anesthesia or regional anesthesia, and between different general anesthetic techniques, have found contradictory findings, which may be explained by the variability in study methodology and definitions of POCD (Monk and Price, 2011; Qiao et al., 2015). Moreover, a deeper level of anesthesia has been associated with an increased incidence of POD but not early POCD (Chan et al., 2013; Radtke et al., 2013; Sieber et al., 2010; Whitlock et al., 2014). It should be noted that the potential link between surgery/ anesthesia and the risk to develop persistent POCD is still a matter of debate (Avidan and Evers, 2016) and some recent studies have challenged the view that anesthesia may contribute to POCD. For example, in a multicenter trial in the Netherlands, 280 patients were randomized to a percutaneous coronary intervention (i.e. procedure with no surgery or general anesthesia), or to off-pump coronary artery bypass grafting. The two groups of patients had no difference in cognitive performance at 7.5 years follow-up (Sauër et al., 2013). Consistent with these results, a systematic review centered on patients older than 65 years and who had


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Box 3. Effect of Ketamine on Brain Structure and Function The Drug A number of drugs from different classes alter brain structure and function including opioids, antidepressants (Castrén and Hen, 2013), and ketamine (Li and Walter, 2016). Ketamine is an intravenous agent with analgesic effect already at subanesthetic doses. This drug is well known to cause cognitive impairment, schizophrenic-like psychoses, or depersonalization at subanesthetic doses (Rogers et al., 2004). Mechanism of Action Ketamine is a NMDA receptor antagonist with fast-acting antidepressant effect (Phoumthipphavong et al., 2016), as well as antinociception, anti-inflammatory and neuroprotective effects (for review see Hirota and Lambert, 2011). Alterations in Brain Structure More directly related to anesthetic drugs, ketamine is of interest because through a putative glutamatergic neuroexcitatory mechanism it is thought to decrease gray matter volume in the superior and medial frontal gyrus (Liao et al., 2011). Changes in white matter in bilateral frontal and left temporoparietal cortices have been associated with chronic use of ketamine (Liao et al., 2010). The drug also induces apoptosis (Mak et al., 2010) and alters dendritic morphology (Phoumthipphavong et al., 2016). In the latter, both short term and long term changes (distal branches and dendritic spine density) have been observed. Such effects may be more severe in immature nervous systems (Vutskits, 2006). Alterations on Brain Function Ketamine disrupts frontal and hippocampal processing in memory (Honey et al., 2005). Using resting-state fMRI, increased connectivity in the cerebellum and visual cortex in relation to the medial visual network and decreased connectivity in the auditory and somatosensory network in relation to regions involved in pain perception and affective processing of pain were observed following exposure to low dose of ketamine (Niesters et al., 2012). The acute and chronic effect of ketamine on brain structure and function has been reviewed in Li and Walter (2016). Synopsis Ketamine thus provides an example of an anesthetic that alters brain systems through mechanisms that include mechanisms of apoptosis and dendritic spine changes, and these seem to be more dramatic in the young brain. Furthermore, in keeping with behavioral changes, ketamine disrupts frontal and hippocampal processing in memory (Honey et al., 2005).

undergone different surgical cardiac procedure (coronary carotid revascularization, cardiac valve procedures, ablation for atrial fibrillation) reported that intermediate and long-term cognitive impairment attributable to a cardiac procedure may be

uncommon in older adults (Fink et al., 2015). The association between exposure to surgery/anesthesia and long-term cognitive outcomes has also been evaluated in a Danish cohort of 8503 middle aged and elderly twins. In this study, only a negligible

Fig. 4. Evaluating short- and long-term effects of Anesthesia on Brain Systems. Neuroimaging techniques (fMRI, EEG, fNIRS, PET) may be used to evaluate the short and longterm effect of anesthetic agents on brain system. Different brain responses across different anesthetics have been suggested with neuroimaging techniques (Akeju et al., 2014; Purdon et al., 2015b). Besides the direct effect of anesthetic agents, other perioperative stressors (pain, brain temperature, inflammation, stress response) may also affect brain circuits. Neuroimaging techniques can help to evaluate these direct and indirect effects using pharmacological MRI (direct effect of the drugs), magnetic resonance spectroscopy (chemical changes), fNIRS, fMRI, or PET (functional changes), fMRI and DTI (morphological changes).

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lower level of cognitive functioning was observed to be associated with a history of major surgery. Moreover, the authors reported that the preoperative cognitive trajectory and underlying diseases were more relevant for the subsequent cognition changes in midand late life than the surgery and anesthesia (Dokkedal et al., 2016) (Table 7).

drugs in the young (e.g. Akeju et al., 2015; Backeljauw et al., 2015; Cornelissen et al., 2015; Makaryus et al., 2015) and adult/aging brain (e.g. Becerra et al., 2016; Deiner et al., 2015; MacDonald et al., 2015; Purdon et al., 2013; Song and Yu, 2015; Yücel et al., 2015).

4.2.3. Evaluating the data: potential confounding factors There is accumulating evidence from animal studies that exposure to anesthetic agents can be toxic for the aging brain. However, the causal link between anesthesia and the development of POCD or Alzheimer’s disease in humans remains difficult to prove. As discussed above, in the elderly, factors other than surgery and anesthesia can also influence cognitive function (Avidan and Evers, 2011; Krenk et al., 2010; Monk and Price, 2011; van Harten et al., 2012). Even in animal studies, factors other than anesthesia can produce cognitive dysfunctions. In an adult rat study using Fentanyl and Droperidol, transient neurocognitive decline was temporally associated with glial activation and increased proinflammatory cytokines in the hippocampus. However, neither cognitive dysfunction nor neuroinflammatory changes were found when the rats were only subjected to anesthesia without surgery (Wan et al., 2007). Moreover, it has been suggested that POCD after anesthesia could be explained by tau hyperphosphorylation resulting from hypothermia consequent to anesthesia (Planel et al., 2007; Run et al., 2009). Accordingly, Xiao et al. (2013) showed that maintenance of body temperature during anesthesia could partly reverse anesthesia-induced cognitive impairment in older mice (Xiao et al., 2013). Other important limitations of studies evaluating the mechanism of POCD in the elderly relate to study design and methodology (for a review see: Avidan and Evers, 2011). Importantly, there are no universally accepted criteria for the assessment and diagnosis of POCD. Furthermore, unlike in animal studies, in humans it is difficult to isolate each factor that may account for POCD (Steinmetz and Rasmussen, 2016). Finally, preoperative cognitive decline and reduced cerebral cognitive reserve may mask diagnosis of POCD, resulting in underestimation of the true incidence (Monk and Price, 2011). Well-designed randomized controlled trials in the elderly (and children) are needed to elucidate the link between general anesthesia, surgery, and long-term cognitive outcomes (Avidan and Evers, 2011; Monk and Price, 2011; Steinmetz and Rasmussen, 2016).

Using PET, dose-dependent changes in cerebral blood flow (CBF) and cerebral metabolic rate of glucose consumption (GMR) have been demonstrated for the majority of anesthetics with a certain degree of regional heterogeneity characteristic to the anesthetic agents (for review see: Song and Yu, 2015; Uhrig et al., 2014). During sedation with midazolam, PET showed significant changes in CBF in distinct neuronal areas related to arousal, memory, and attention involving the prefrontal cortex, the superior, middle, and medial frontal gyri, the cingulate gyrus, parietal and temporal association areas, the insular cortex, and the thalamus (Bagary et al., 2000; Heinke and Schwarzbauer, 2002; Reinsel et al., 2000; Veselis et al., 1997). Similarly, in propofolinduced unconsciousness, a decrease in blood-flow was seen in the medial thalamus, orbitofrontal cortex, cuneus and precuneus, and the posterior cingulate and right angular gyri, while decreased GMR occurred in the thalamus, cerebral cortex, and hippocampus brain regions also involved in arousal and associative functions (Song and Yu, 2015). With volatile anesthetics, PET has also shown modulation of glucose metabolism. One MAC of sevoflurane produced the greatest decreases in regional GMR in the occipital lobe (including the lingual gyrus) and the thalamus, and the smallest decreases in the putamen, caudate nucleus, and cingulate and frontal cortices (Schlunzen et al., 2010). Fig. 5 shows an example of the effects of propofol and isoflurane on cerebral metabolism (Alkire et al., 1995, 1997). At the level of unconsciousness, isoflurane and sevoflurane have very similar effects on CBF, with increases in the anterior cingulate and insula regions, and decreases in the cerebellum, thalamus, cuneus, and lingual gyrus (Schlunzen et al., 2004, 2006). In contrast, ketamine produces an increase in CBF and GMR. Subanesthetic doses (sedation, analgesia) of ketamine induced a global increase in rCBF in a dose-dependent manner. The greatest increases were seen in the anterior cingulate cortex, thalamus, putamen, and frontal cortex (Långsjö et al., 2003). Also at subanesthetic doses, ketamine produced a global increase in regional GMR with the highest increases in the thalamus, the frontal and parietal cortices, and to a lesser extent in the cerebellum, caudate nucleus, and cingulate gyrus (Långsjö et al., 2004).

5. Brain measures: insights into anesthetic effects on brain structure and function

5.1. Positron emission tomography (PET)

5.2. Functional magnetic resonance imaging (fMRI) Although molecular targets for most anesthetic drugs have been identified, less is known on the effects at a system level regarding the neural mechanisms by which anesthetic agents impact brain circuits to produce unconsciousness (Meyer, 2015). Recent advances in neuroimaging methods, specifically positron emission tomography (PET), magnetic resonance imaging (MRI), near-infrared spectroscopy (NIRS), and electroencephalogram (EEG), have begun to improve our understanding of how anesthetic drugs act on the brain to alter level of consciousness and cognition (MacDonald et al., 2015) (Box 3). Moreover, NIRS and EEG can be used clinically to help identify changes in brain during sedation and anesthesia. Effects of anesthetic drugs may be evaluated in terms of (1) direct pharmacological effects of the drug using pharmacological MRI (phMRI) (Nathan et al., 2014); (2) Chemical changes short- and long-term using magnetic resonance spectroscopy (MRS); (3) Functional changes using fNIRS, fMRI, and/or PET; (4) Morphological changes using MRI with diffusion tensor imaging (DTI) (Fig. 4). These imaging techniques have been used in the perioperative period to evaluate to effect of anesthetic

Functional MRI measures of anesthesia have recently been reviewed (Tang and Ramani, 2016). Anesthesia disrupts connections between higher and lower order networks (Boveroux et al., 2010). Higher order networks (default mode network, executive control network, salience network) are more sensitive to anesthetic agents (Kim et al., 2012; Tang and Ramani, 2016). Fig. 6 shows an example of large-scale network connectivity during propofol unconsciousness. General anesthesia is a brain process of “disruption” of higher-order cortical information integration (Hudetz, 2012). While acute alterations in brain function can be observed with fMRI (and PET), differences between short- and long-term effects of anesthetic drugs on behavior are likely a consequence of these initial effects. 5.2.1. Short-term changes MRI techniques have been used extensively to study the neuronal correlates of anesthetic agents. Most studies have investigated the changes in responsiveness of neuronal networks


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Fig. 5. PET Measures of Anesthesia – Effects of Anesthesia on Decreasing Cerebral Metabolism. Note for both drugs, the effects show a profound decrease in metabolism. Top: Decreasing cerebral metabolism under propofol anesthesia. The left image depicts the glucose metabolism measurement (mmoles/100 g 1/min 1) of one subject in the awake condition. The right image depicts the glucose metabolism measurement of the same subject in the anesthetized condition. A reduction of 57% whole-brain metabolism was observed (Adapted from Alkire et al., 1995, with permission, © 1995 American Society of Anesthesiologists, Inc. J.B. Lippincott Company). Bottom: Decreasing cerebral metabolism under isoflurane anesthesia. The left image depicts the awake baseline resting brain metabolic state of one subject. The right image depicts the brain metabolism of the same subject under isoflurane anesthesia. Unresponsive behavior appeared at a concentration of 0.4% expired isoflurane. The scale represents glucose utilization (mg/ 100 g 1/min 1) (Adapted from Alkire et al., 1997, with permission, © 1997 American Society of Anesthesiologists, Inc. Lippincott-Raven Publishers).

during processing of sensory stimuli. During sedation and anesthesia, studies have shown that sensory information processing in the cortex is impaired in a dose-dependent manner (for a review see: Hudetz, 2012; MacDonald et al., 2015). During the transition from consciousness to unconsciousness, the majority of studies have observed an initial decline in cortical activation in higher-order cortical association areas (i.e. memory, word processing, cognition), followed by attenuation and sometimes complete eradication of responses in the primary cortex (MacDonald et al., 2015). For example, with lower doses and a lighter level of anesthesia, auditory activations in response to words or sounds were preserved in a large part of the cortical regions activated in the awake state (Dueck et al., 2005; Kerssens et al., 2005). At deeper stages of anesthesia, BOLD auditory-related activations were attenuated (Kerssens et al., 2005), or significantly reduced (Dueck et al., 2005) in the primary cortex. In contrast, studies assessing more complex aspects of language processing have suggested that complex auditory processing is already impaired during sedation or a light level of anesthesia (Adapa et al., 2014; Davis et al., 2007; Heinke, 2004; Plourde et al., 2006). Antognini et al. (1997) found no significant cortical activation to innocuous (tactile, mild electric shock) somatosensory stimulation at low (0.7%) and moderate (1.3%) concentrations of isoflurane. In contrast, responses to a supramaximal electrical stimulus were preserved in the caudate nucleus and thalamus at the low dose but abolished at the moderate dose of isoflurane (Antognini et al., 1997). Similarly, Hofbauer et al. (2004) have revealed with PET that cortical activity in response to a painful thermal stimulus is partly preserved with propofol-induced unconsciousness.

Functional MRI has also been used to study the effects of anesthetics on functional and effective connectivity. Findings depend on the specific anesthetic agent, dosage, and the network studied (Hudetz, 2012). Consistent with studies investigating changes in neuronal activity in response to sensory inputs, higherorder functional networks appear to be more sensitive to anesthesia than lower-order sensorimotor networks (Hudetz, 2012; MacDonald et al., 2015). In a recent resting-state study, loss of responsiveness induced by propofol was mostly linked with decreased connectivity in the frontal lobe (Guldenmund et al., 2016). 5.2.2. Long-term changes Only a few studies have evaluated the long-term effects of general anesthesia with fMRI. In a recent study of healthy children exposed to surgery and anesthesia before 4 years of age, Backeljauw et al. (2015) found an association between lower performance IQ and language comprehension with lower grey matter volume in posterior brain regions. In an animal model, Makaryus et al. (2015) used proton magnetic resonance spectroscopy (1HMRS) on pup rats exposed to sevoflurane for 5 h on postnatal day (PND) 7 or 15. 1HMRS spectra were acquired in the thalamus 24 and 48 h after the exposure. Whereas an increase in Nacetylaspartate (NAA) was found 24 and 48 h later in unexposed PND 7 rats, in exposed pups N-acetylaspartate concentration did not change and there was a 25% decrease in the concentrations of glycerophosphorylcholine and phosphorylcholine. In contrast, NAA increased in both unexposed and exposed PND 15 rats. Magnetic resonance spectroscopy therefore might be a useful tool

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Fig. 6. fMRI Measures of Anesthesia – Differential Effects on components of Brain Resting Networks. The left panel represents the resting-state network connectivity during normal awake conditions and the right panel represents the resting-state network connectivity in the same network but during deep sedation with clinical unconsciousness: default network (A, E), right executive control network (B, F), left executive control network (C, G), anticorrelations between default-mode network and lateral frontoparietal cortices (D, H). (Adapted from Boveroux et al., 2010, with permission, © 2010, the American Society of Anesthesiologists, Inc. Lippincott Williams & Wilkins.).

to evaluate changes in neuronal metabolites following exposure to general anesthesia.

5.3. Functional near-infrared spectroscopy (fNIRS) fNIRS is a non-invasive optical technique that can be used clinically to measure activity within the cortical areas of the human brain. This method measures changes in the concentrations of oxygenated (HbO), de-oxygenated (HbR), and total hemoglobin (HbT) as part of the hemodynamic response to alterations in neuronal activity in the cerebral cortex (Boas et al., 2014; Hernandez-Meza et al., 2015). Because it is safe, non-invasive, portable, and has a high temporal resolution, fNIRS can be used to study changes in depth of anesthesia (for review see: Boas et al., 2014; Hernandez-Meza et al., 2015) or to record brain activation to noxious stimulation or pain under general anesthesia (Becerra et al., 2016; Kussman et al., 2016) (Fig. 7). 5.3.1. Short-term changes fNIRS can be used to differentiate brain responses to innocuous versus noxious somatosensory stimuli with noxious stimuli

eliciting activation in the contralateral somatosensory cortex and deactivation in the frontal cortex (Yücel et al., 2015). Accordingly, fNIRS has recently been used to measure the response to a noxious stimulus known to be painful in patients under general anesthesia or sedation (Becerra et al., 2016; Kussman et al., 2016). In a study investigating cortical hemodynamic responses to catheter ablation of arrhythmias in patients under general anesthesia, a pattern of change in HbO concentration, temporally related to the cardiac ablative lesion, was found in the frontal cortex (Kussman et al., 2016). This pattern was very similar to those found in awake healthy volunteers in response to a noxious stimulus (Yücel et al., 2015), and in healthy patients moderately sedated for colonoscopy (Becerra et al., 2016). Thus fNIRS could be used to objectively measure noxious stimulation during painful clinical procedures and to assist with the management of pain in populations unable to communicate (e.g., neonates, infants, general anesthesia, stroke, dementia). 5.3.2. Long-term changes To our knowledge, fNIRS has not been used to evaluate any longterm changes of anesthetics on brain function and structure. Although studies exist in which intraoperative NIRS was associated


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Fig. 7. NIRS Measures of Pain Under different levels of consciousness – Pain processing still present in Anesthesia. fNRIS can be used to measure responses to a painful stimulus. A. Changes in HbO in healthy awake subjects as a response to an innocuous stimulus (blue) and noxious stimulus (red) in the frontal region (Adapted from Yücel et al., 2015, http://creativecommons.org/licences/by/4.0/). B. Average changes in HbO in healthy anesthetized subjects during rest (dashed lines) and during cardiac ablations (nociceptive condition – solid line) across all frontal channels (Adapted from Kussman et al., 2016, http://creativecommons.org/licences/by/4.0/). C. Average changes in HbO in sedated healthy subject during rest (dashed lines) and insufflation of the bowel for a colonoscopy (nociceptive condition – solid line) across the prefrontal cortex (Adapted from Becerra et al., 2016, with permission, © 2015 by the International Association for the Study of Pain. Wolters Kluwer Health, Inc).

with early (Austin et al., 1997; Murkin et al., 2007; Plomgaard et al., 2016) and late (Andropoulos et al., 2013; Kussman et al., 2010; Slater et al., 2009) neurological morbidity, the associations have been attributed to the effects of reduced cerebral oxygenation and the NIRS monitors were not designed to assess brain function and structure. The relationship between intraoperative NIRS and longterm neurologic outcomes are mostly associations without demonstration of causality. 5.4. Electrophysiological methods (EEG) Electroencephalography (EEG) can be used to monitor changes in brain activity during exposure to anesthetic agents. All anesthetic agents induce changes in brain electrical activity (Brown et al., 2010). For example, maintenance of general anesthesia with propofol or sevoflurane in adults leads to simultaneous slow (

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