Biological complexity and adaptability of simple mammalian olfactory memory systems.

Neuroscience and Biobehavioral Reviews 50 (2015) 29–40

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Neuroscience and Biobehavioral Reviews journal homepage: www.elsevier.com/locate/neubiorev

Review

Biological complexity and adaptability of simple mammalian olfactory memory systems P. Brennan a , E.B. Keverne b,∗ a b

School of Physiology and Pharmacology, University of Bristol, Bristol, UK Sub-Department of Animal Behaviour, University of Cambridge, Cambridge, UK

a r t i c l e

i n f o

Article history: Received 5 March 2014 Received in revised form 20 August 2014 Accepted 22 October 2014 Available online 31 October 2014 Keywords: Olfactory Vomeronasal Learning Memory Receptors Neurogenesis

a b s t r a c t Chemosensory systems play vital roles in the lives of most mammals, including the detection and identification of predators, as well as sex and reproductive status and the identification of individual conspecifics. All of these capabilities require a process of recognition involving a combination of innate (kairomonal/pheromonal) and learned responses. Across very different phylogenies, the mechanisms for pheromonal and odour learning have much in common. They are frequently associated with plasticity of GABA-ergic feedback at the initial level of processing the chemosensory information, which enhances its pattern separation capability. Association of odourant features into an odour object primarily involves anterior piriform cortex for non-social odours. However, the medial amygdala appears to be involved in both the recognition of social odours and their association with chemosensory information sensed by the vomeronasal system. Unusually not only the sensory neurons themselves, but also the GABA-ergic interneurons in the olfactory bulb are continually being replaced, with implications for the induction and maintenance of learned chemosensory responses. Crown Copyright © 2014 Published by Elsevier Ltd. All rights reserved.

Contents 1. 2. 3.

4. 5. 6. 7.

Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . Stereotyped versus learned olfactory responses . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . Main olfactory learning and memory . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3.1. Non-social odour learning in the main olfactory system . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3.2. Short-term social recognition in rodents . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3.3. Lamb odour recognition in sheep . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . Pheromonal learning in the vomeronasal system. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 4.1. The medial amygdala and social recognition . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . Neurogenesis and olfactory learning . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . Vomeronasal receptor neuron turnover . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . Conclusions . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

1. Introduction The chemosensory systems are well established in small brain mammals providing information on predators, gender, offspring and sexual status. All of these features require a process of

∗ Corresponding author at: Sub-Department of Animal Behaviour, University of Cambridge, High Street, Madingley, Cambridge CB23 8AA, UK. Tel.: +44 01223 741816; fax: +44 01223 741802. E-mail address: [email protected] (E.B. Keverne). http://dx.doi.org/10.1016/j.neubiorev.2014.10.020 0149-7634/Crown Copyright © 2014 Published by Elsevier Ltd. All rights reserved.

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recognition based on learning and memory. Across very different phylogenies the mechanisms for pheromonal and olfactory learning have much in common. They all engage important and necessary processing at the first relay (accessory olfactory and main olfactory bulb) where intrinsic GABA-ergic neurons are integral to the synchronisation of the output from projection neurons which also play an integral role in the recognition process by these trilaminar circuits (Brennan and Keverne, 1997). The mammalian vomeronasal chemosensory system (VNO) connects primarily with the amygdala and hypothalamus, regions of

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P. Brennan, E.B. Keverne / Neuroscience and Biobehavioral Reviews 50 (2015) 29–40

the brain concerned with neuroendocrine responses and primary motivated behaviour. These sub-cortical brain regions regulate hormonal changes that are relevant to sexual behaviour, maternal care and fear responding. In these contexts, recognition of mates and offspring are sustained, contributing to reproductive success and offspring survival. The first relay from the VNO receptors is the accessory olfactory bulb (AOB) which, in the context of pregnancy block, has itself the capacity to form and retain long term memories (Brennan et al., 1990). Chemosensory neurons are exposed to the vagaries of the external environment and have a relatively short lifespan. Consequently, these receptor neurons undergo regenerative turnover, which continues throughout life. Turnover of receptor neurons can provide adaptability to new environments by selection for different receptor types and the formation of new memories (Nissant and Pallatto, 2011). This raises the question as to how the balance between retaining “old” memories and prioritising new memories occurs. Integral to this is the added complication of adult regeneration of intrinsic bulbar GABA-ergic neurons arriving to this front-line processing circuitry from the sub-ventricular rostral flow of neural progenitor cells. Mammalian chemosensory neurons are characterised by a large number of genes coding for the receptors that respond to chemical cues and pheromones. This is illustrated by 1400–1700 olfactory receptors, 300-170 vomeronasal class 1 receptors (V1Rs) and 280220 vomeronasal class 2 receptors (V2Rs) in the mouse and rat (Ibarra-Soria et al., 2014). In Old world primates and apes (including humans) the vomeronasal receptors are virtually all non-coding pseudogenes, while the main olfactory receptors are reduced to a third of the number found in rodent species (Kambere, 2007). The mammalian evolutionary trend for this diminished representation of VNO chemoreception in particular has paralleled the increase in mammalian neocortical size and the switch from nocturnal to diurnal lifestyles (Nei et al., 2008). Nevertheless, the basic principles for coding and initial processing of chemosensory cues and pheromones have much in common across the vomeronasal and main olfactory systems. The vomeronasal system has a vital and complementary role to the main olfactory system. In mammals vomeronasal receptors are located in the vomeronasal organ, a blind-ended tubular structure in the nasal septum that is connected to the nasal and/or oral cavities (Døving and Trotier, 1998). The main distinction between the function of the two systems appears to be that the main olfactory system is adapted to detect airborne odourants, whereas stimulus access to the vomeronasal organ depends on pumping relatively involatile stimuli such as peptides and proteins into the vomeronasal organ following physical contact with the stimulus source (Meredith and O’Connell, 1979). The mammalian vomeronasal system connects primarily with the amygdala and indirectly to the hypothalamus, regions of the brain concerned with neuroendocrine responses and primary motivated behaviour. These sub-neocortical brain regions regulate coordinated behavioural, autonomic and hormonal responses in contexts such as sexual behaviour, parental care and aggressive/defensive behaviour. The main olfactory and vomeronasal systems play complementary roles in both innate and learned responses to chemosensory signals. This review aims to highlight the different neural mechanisms involved in learning social and non-social olfactory cues and the enhanced plasticity provided by neurogenesis in olfactory systems.

2. Stereotyped versus learned olfactory responses It used to be thought that the main olfactory system mediated a flexible, learned response to odours, whereas the vomeronasal

system mediated relatively stereotyped innate responses to pheromones, However, this has given way to a more complex picture in which both the main olfactory and vomeronasal systems can mediate sterotyped responses to chemosignals, and the responses via both systems can be influenced by learning. The two systems have interrelated functions often providing complementary information about the same stimulus source. For instance, both the main and vomeronasal systems can mediate an avoidance response and freezing behaviour in mice. Predators produce involatile urinary chemosignals belonging to the major urinary protein (MUP) family, which stimulate the V2r receptor class of vomeronasal receptor to elicit freezing and avoidance behaviour in mice (Papes et al., 2010). Different predators produce different MUP variants in their urine potentially enabling mice to discriminate chemosignals from different species of predator (Ben-Shaul et al., 2010). The main olfactory system can also mediate responses to predator chemosignals, such as the volatile odourant trimethlytoluine (TMT), which is a component of fox faeces. Class I olfactory receptor proteins are expressed by olfactory sensory neurons (OSNs) in the dorsal zone of the olfactory epithelium and project to glomeruli in the D1 region of the main olfactory bulb (MOB). OSNs expressing Class II olfactory receptor proteins are located in the ventral zone of the olfactory epithelium and project to the D2 region and to the ventral region of the MOB (Kobayakawa et al., 2007). Wildtype lab mice have an aversion to TMT, which is lost in mice with a largescale genetic deletion of OSNs expressing class II olfactory receptors that project to the D2 domain of the MOB. This loss of stereotyped aversion to TMT odour was not due to the inability to detect TMT, as the mice could still learn to either approach or avoid TMT in appetitive and aversive conditioning paradigms via class II-expressing OSNs that projected to the ventral region of the MOB (Kobayakawa et al., 2007). These findings demonstrate that there are populations of mitral cells in D2 domain of the MOB that are “hardwired” to regions that generate aversive responses to predator odours. The activation of the medial aspect of the bed nucleus of the stria terminalis (BNST) by TMT in wild-type but not D2 deleted mice identifies this pathway as mediating innate responses to predator chemosignals (Kobayakawa et al., 2007). Interestingly, mice also show innate avoidance responses to odours that are typical of spoiled foods, such as 2-methylbutyric acid and isoamylamine, which appear to be mediated by the D1 region of the MOB (Kobayakawa et al., 2007) but do not activate the medial aspect of the BNST. These findings suggest that different sub-regions of the MOB mediate innate responses in different behavioural contexts and that these differ from the MOB subregions that convey learned odour responses (Fig. 1). Both the main olfactory system and the vomeronasal system also mediate pheromonal responses to chemosignals from members of the same species. For instance, the main olfactory system of rabbits mediates the response to the rabbit mammary pheromone 2-methylbut-2-enal (Schaal et al., 2003). This airborne pheromone is produced by the skin surrounding the doe’s nipples and induces robust arousal and stereotyped nipple search behaviour in rabbit pups that enables successful nipple location, attachment and suckling (Distel and Hudson, 1985). The vomeronasal system mediates the pheromonal effects such as those of exocrine secretory peptide 1 (ESP-1). This is a peptide pheromone produced in male mouse tear secretions that is sensed by the V2Rp5 vomeronasal receptor and increases the proportion of females showing lordosis in response to mounting attempts by males, via a sexually dimorphic pathway to the ventromedial hypothalamus (Haga et al., 2010). Another exocrine secreting peptide found in mouse tear secretions, ESP-22, is only produced by juvenile mice inhibits mounting attempts by males (Ferrero et al., 2013). Interestingly MUPs from male mice that are from the same family as predator MUPs are sensed by V2Rs to


(a) 2MB and IA-amine (bad food odour) MOE

D1 D2 V

BST LD

Innate odour avoidance

MOB

BLA

Conditioned odour avoidance/fear

(b)

BST MA

TMT (predator odour) D1 D2 MOE V

Innate fear response (freezing, stress)

BST LD

Innate odour avoidance?

BLA

Conditioned odour avoidance/fear

MOB

Fig. 1. Separate pathways through the main olfactory bulb (MOB) mediate innate and learned responses to aversive odours in mice. (a) Off-food odours, such as 2-methylbutyric acid (2MB) and isoamylamine (IA-amine) sensed by the main olfactory epithelium (MOE), elicit innate avoidance behaviour, via glomeruli in the D1 region of the main olfactory bulb (MOB) and the lateral division of the bed nucleus of the stria terminalis (BST LD). (b) The predator odour trimethyltoluene (TMT), sensed by the MOE, elicits innate fear responses, such as freezing and activation of the hypothalamic-pituitary-adrenal axis, via glomeruli in the D2 region of the MOB and the medial aspect of the bed nucleus of the stria terminalis (BST MA). Learned responses to both off-food and predator odours can be mediated by a separate pathway involving the ventral (V) region of the MOB and non-BST pathways, such as the basolateral amygdala (BLA).

generate male aggressive behaviour in the appropriate behavioural context (Chamero et al., 2007). Not all responses mediated by the main olfactory or vomeronasal systems are hardwired to elicit innate responses. This is particularly true regarding the recognition of individuality, where a certain profile, of volatile and non-volatile chemosignals, only acquires meaning following association with a particular individual. Particularly robust mechanisms of learning of individuality chemosignals have evolved, which play vital roles in the recognition of mates and offspring contributing to reproductive success and offspring survival (Brennan and Kendrick, 2006). 3. Main olfactory learning and memory 3.1. Non-social odour learning in the main olfactory system OSNs in the main olfactory epithelium express only one olfactory receptor type. The mouse olfactory bulb contains roughly 2000 glomeruli, each glomerulus receiving projections from OSNs expressing the same receptor type, with approximately 2 glomeruli for each receptor type in each olfactory bulb (Mombaerts et al., 1996). The olfactory receptor type expressed by the neuron determines the location of its glomerular convergence on the MOB. This self-organising developmental process leads to a spatial mapping of input from different receptor types (Imai et al., 2006), which is broadly consistent within and between individual mice and rats (Soucy et al., 2009). There appears to be some aspect of localisation of responses to different odourant classes across broad regions of the MOB (Mori and Sakano, 2011). However, no topographic mapping has been found at a finer scale, in that neighbouring glomeruli have been found to respond to completely different odourant types (Soucy et al., 2009). Each olfactory receptor type responds

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differently, but with overlapping ranges to structurally-related odourant molecules. This is therefore mirrored in the range of odourants that elicit responses in the glomerulus receiving input from that receptor type. Imaging glomerular responses using Ca2+ sensitive dyes across a variety of mammalian species has revealed that both individual odourants and naturally occurring odours, which are complex mixtures of odourants, activate a number of glomeruli distributed across the MOB (Mori et al., 2006). Nevertheless, using optogenetics to selectively activate a single glomerulus, it has been shown that each glomerulus can transmit odour information using identity, intensity and temporal codes (Smear et al., 2013). External tufted cells and mitral cells in the main olfactory bulb sample information from a single glomerulus, and therefore receive information about activation of an individual olfactory receptor type. They convey this information in their highly distributed projections to central olfactory brain areas, including the anterior olfactory nucleus, olfactory tubercle, cortical amygdala, piriform cortex and entorhinal cortex (Mori et al., 1999). However the pattern of mitral cell activity in response to an odour does not depend solely on the pattern of glomerular input, but also on inhibitory regulation by local inhibitory interneurons, as well as centrifugal feedback from central olfactory areas. There are two main classes of local inhibitory interneuron in the MOB. Periglomerular cells are dopaminergic and/or GABAergic and modulate glomerular input from the spatial pattern of mitral cell activity (Li and Cleland, 2013). Granule cells are the predominant class of interneurons and are thought to mediate long-range lateral interactions among mitral cells and play an important role in synchronising their activity (Li and Cleland, 2013). A unique feature of these interactions between mitral cells and the periglomerular and granule cell interneurons is that they occur at reciprocal dendrodendritic synapses (Isaacson and Strowbridge, 1998). Glutamatergic input from mitral cells depolarises the periglomerular/granule cell dendritic spine, which in turn releases GABA to inhibit the mitral cell. This tightly coupled form of inhibitory feedback control of projection neurons is equivalent to the axonally-mediated interneuronal inhibition of projection neurons found in other cortical areas. Sufficient glutamatergic input to granule cells will trigger spiking of the granule cell, releasing GABA onto the lateral dendrites of connected mitral cells (Isaacson and Strowbridge, 1998). These reciprocal synapses thus have the potential to mediate both self- and lateral-inhibition of mitral cell activity in addition to mediating centrifugal feedback to mitral cells from cortical areas, such as anterior olfactory nucleus and piriform cortex, that terminate on granule cells (Boyd et al., 2012; Markopoulos et al., 2012). The inhibitory interneurons effectively shape the spatial pattern of mitral cell activity that represents an odour. This is not a static pattern of activity but a pattern that evolves during the odour response (Niessing and Friedrich, 2010; Spors and Grinvald, 2002), in which the precise timing of mitral cell activity within the sniff cycle can be discriminated (Smear et al., 2011). Furthermore, the spatiotemporal patterns of mitral cell activity are modulated by learning. Changing the meaning of an odour, by changing the reward contingency of the odour, changes the mitral cell responses to the odour (Doucette et al., 2011; Doucette and Restrepo, 2008; Freeman and Schneider, 1982; Kay and Laurent, 1999). The MOB thus appears to function as a pattern separator, in which learning acts to decorrelate similar patterns of odour input that have specific significance for the animal. In addition to enhancing the discriminability of odours that have different meanings for the animal, the olfactory system has to be able to generalise similar patterns of odour input that have similar meaning. This is vital, as the odourant profile of an odour is likely to vary from one presentation to the next and so the olfactory system

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has to be able to perform pattern completion to generalise to the same output from a degraded input. This appears to be an important function of the anterior piriform cortex, which has recurrent connectivity typical of pattern completing neural networks (Haberly, 1985). Mitral cells send highly divergent projections to the piriform cortex, with each mitral cell input distributed across a wide region of piriform cortex. The pyramidal cells of anterior piriform cortex receive weak synaptic input on their primary dendrites from mitral cells from across the MOB, along with inputs from anterior olfactory nucleus and recurrent connectivity from anterior piriform cortex pyramidal cells (Haberly, 1985). Optical uncaging of glutamate to activate individual glomeruli in the MOB has shown that activation of a single MOB glomerulus is normally insufficient to fire spikes in anterior piriform cortex pyramidal cells of anaesthetised mice. Instead, synchronous activation of at least 4 glomeruli are required for pyramidal neuron spiking, and individual pyramidal cells are activated by different patterns of glomerular activation (Davison and Ehlers, 2011). Therefore the spatiotemporal pattern of mitral cell activation that represents an odour at the level of the MOB is represented as a sparse and distributed pattern of pyramidal cell activation at the level of the anterior piriform cortex (Stettler and Axel, 2009). Ca2+ -sensitive dye imaging of pyramidal cell responses to odours shows this broad distributed representation, where cells responsive for different odours are intermingled without any apparent topographic representation of odours (Stettler and Axel, 2009). Anterior piriform cortex pyramidal cells thus function as a pattern recognition system in which long-term potentiation strengthens co-incident inputs, associating the combination of odour features that represent an odour object. Indirect evidence for the ability of the piriform cortex to recognise learned patterns of mitral cell input comes from cross adaptation studies in anaesthetised rats. Cross adaptation of MOB mitral cell responses was found for aliphatic aldehydes differing by 2 carbon atoms in chain length, whereas little cross adaptation was found between pyramidal cell responses to the same odourants in the anterior piriform cortex (Wilson, 2000). This suggests that the MOB is representing individual odourant features, whereas the anterior piriform cortex is associating odourant features into a unitary representation of the mixture of odourant features – as odour objects. In addition, a pattern completion function of anterior piriform cortex is supported by studies of mixtures of odourants in which a ten-component (10c) odourant mixture is more difficult for rats to discriminate from a similar mixture in which one of the components has been removed (10c − 1) compared to a similar mixture in which one component has been replaced. This behavioural generalisation of the 10 and 10 − 1 component mixtures is mirrored by a higher level of correlation between the patterns of activity of anterior piriform cortex pyramidal neurons compared to MOB mitral cell responses (Barnes et al., 2008). Furthermore, training the rats to discriminate the 10 vs 10 − 1 mixtures results in decreased correlation of the piriform cortex responses that mirrors the acquisition of the behavioural discrimination (Chapuis and Wilson, 2012). These proposed roles of the MOB in pattern separation and the anterior piriform cortex in pattern completion and recognition may be analogous to similar roles performed by the dentate gyrus and CA3 regions of hippocampus (Yassa and Stark, 2011). In contrast to the pattern recognition role of anterior piriform cortex, the connectivity of posterior piriform cortex, suggests that it functions as higher-order association cortex, performing multimodal associations (Haberly, 1985). The olfactory system also has major input to the hippocampus via direct and indirect projections of the MOB to entorhinal cortex. However, simple associative odour learning has been shown to be unaffected by hippocampal lesions (Eichenbaum, 1998). Instead, hippocampal lesions have been found

to disrupt configural learning of odour cues, such as the sequence of odour presentation (Fortin et al., 2002). Neurons in orbitofrontal cortex have been found to respond to the reward contingencies of the odour and the neurons reversed their responses to odours when the reward contingencies changed (Rolls et al., 1996; Schoenbaum and Eichenbaum, 1995b). Furthermore orbitofrontal neurons recorded from awake rats performing an 8-odour discrimination task, in which there were predictable sequences of odours, were found to encode multiple aspects of the task. In addition to responding to the odour presented and its reward contingency, firing of orbitofrontal neurons was also modulated by whether the odour had been predicted by a previous odour in the sequence and whether it predicted that a rewarded odour would be presented next (Schoenbaum and Eichenbaum, 1995a). However, these complex response properties are not solely a feature of orbitofrontal cortex. There are extensive reciprocal connections between anterior piriform cortex and orbitofrontal cortex via dorsomedial thalamus and recordings of pyramidal cell activity in anterior piriform cortex of awake behaving rats found that their response properties were qualitatively indistinguishable to those found in orbitofrontal cortex (Schoenbaum and Eichenbaum, 1995a). Thus different aspects of non-social odour learning involve plasticity across multiple olfactory brain regions, and the reward contingencies of an odour modulate neuronal activity across all levels of olfactory information processing from MOB to orbitofrontal cortex. 3.2. Short-term social recognition in rodents The ability to discriminate between and to learn to recognise other individuals of the same species is vital for mammalian social behaviour. In the majority of mammalian species this is achieved by the recognition of specialised “individuality” chemosignals and/or general body odour. The standard test for short-term social recognition in rodents, such as mice, is to introduce an unfamiliar juvenile to the subject, which elicits an intense bout of olfactory investigation, without the complication of eliciting sexual or aggressive behaviours. If the juvenile is removed and subsequently reintroduced to the subject, the amount of investigation is reduced, as it is recognised as familiar. This habituation of investigation time on reintroduction of a previously investigated animal is a measure of the ability of the subject to recognise the test animal and to discriminate it from unfamiliar individuals that have not been previously encountered and that elicit high levels of investigation. This form of social recognition in rats normally has a duration of around 90 min, but has been found to be enhanced by peripheral administration of the neuropeptides oxytocin and vasopressin (Dantzer et al., 1987). Moreover, social recognition was prevented in oxytocin-knockout mice and could be restored by intracerebroventricular oxytocin infusions, or replicated in wild-type mice by intracerebroventricular infusions of an oxytocin antagonist prior to the initial social encounter (Ferguson et al., 2000). Importantly, general olfactory ability and the habituation to non-social odour stimuli and other non-social forms of learning were not significantly affected by the lack of oxytocin. These findings imply that there are separate neural mechanisms for learning social compared to non-social odours. Subsequent experiments established that although the MOB and medial amygdala both have high levels of oxytocin receptor expression, only local infusions of oxytocin targeted to the medial amygdala and not the MOB, were effective in restoring social recognition in the oxytocin knockout mice (Ferguson et al., 2001). However there may be species differences, as both oxytocinergic and vasopressinergic transmission in the MOB promote recognition of social odours in rats (Wacker and Ludwig, 2012). Vasopressin neurons are located in the external plexiform layer of the olfactory bulb with their dendrites in the glomerular layer, which receives


the olfactory sensory input. Vasopressin neurons in both the main and accessory olfactory bulbs are intrinsic glutaminergic rather than GABA-ergic neurons that modulate mitral cell activity (Tobin et al., 2010). Pharmacological inhibition of olfactory bulb V1a receptors disrupts social recognition in rats, without affecting responses to non-social odours (Tobin et al., 2010). Also, bilateral infusion of both oxytocin and vasopressin into the olfactory bulbs lengthens the retention interval for short term social odour recognition in male rats (Dluzen et al., 1998a,b). These effects of oxytocin and vasopressin appear to be dependent on noradrenergic transmission in the olfactory bulb as they are eliminated following noradrenaline depletion or the administration of an alpha adrenergic antagonist (Dluzen et al., 2000, 1998a,b). The priming effect of initial dendritic Ca2+ elevation on subsequent dendritic vasopressin release, in the olfactory bulb, has been proposed to account for short-term social recognition (Wacker and Ludwig, 2012). According to this hypothesis, the initial, short encounter with an unfamiliar juvenile primes the dendrites of vasopressinergic neurons that are activated by the sub-population of glomeruli receiving input from the juvenile’s chemosensory signature. If the rat re-encounters the same juvenile, during the short-term primed state, then depolarisation of the primed neurons would result in local dendritic vasopressin release in the vicinity of the activated glomeruli. This enhanced vasopressin release could then inhibit mitral cell activity, supressing transmission of the chemosensory information from the familiar juvenile to central olfactory areas leading to reduced chemosensory investigation (Wacker and Ludwig, 2012). Although this is an attractive hypothesis, there is clearly a need for further research to establish the relationship between the roles of vasopressin and oxytocin in olfactory bulb function. 3.3. Lamb odour recognition in sheep Newborn lambs have much in common with respect to their appearance, vocalisations and tactile cues at suckling. Nevertheless, olfactory recognition of own lamb is crucial to ensure selective acceptance of own offspring in a flock of synchronised seasonal breeding ewes. Here the selective recognition of own and rejection of strange lambs is crucial to maternal reproductive success. After giving birth, sheep and many other species from a selective bond with their offspring based on olfactory recognition of main olfactory mediated chemosignals (Lévy et al., 1995). This reliance on main olfactory cues of individuality is likely to reflect the lack of functional class 2 vomeronasal receptors in the sheep genome (Nei et al., 2008), which are the receptor type that mediate responses to involatile peptides and proteins in the vomeronasal system. Nevertheless, the finding of sensory neurons in the main olfactory epithelium of mice that respond to MHC peptides (Spehr et al., 2006), still leaves open the possibility of responses based on genetically-determined non-volatile chemosignals sensed by the main olfactory system. Before giving birth, a ewe spends the majority of her time eating and this is reflected in the response properties of mitral cells recorded in the MOB, the majority of which respond preferentially to food odours. Notably, before birth none of the mitral cells responded preferentially to lamb odours and nor was there any significant release of MOB neurotransmitters in response to lamb odours that could be detected using microdialysis (Kendrick et al., 1992). After birth, there was an abrupt change in the response landscape of the MOB. Now, the number of mitral cells recorded from the same region of the MOB that responded preferentially to lamb odours was dramatically increased. This was associated with dramatic increases in glutamatergic, GABAergic, cholinergic and noradrenergic neurotransmitter release in the MOB in response to lamb odours (Kendrick et al., 1992). These observations suggest that

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recognition of own lamb odours is accompanied by increased activity of a subset of MOB mitral cells. Depletion of bulbar noradrenaline using 6-hydroxydopamine infusions into the olfactory stria, or local infusions of the beta adrenergic antagonist propranolol during the critical post birth period prevented the development of the selective behaviour towards own lamb (Lévy et al., 1990). The bulbar noradrenaline release is triggered by the vaginocervical stimulation at birth and induces plasticity of the bulbar circuits that respond to own lamb odour. This is likely to enhance the separation of the bulbar representation of own lamb odour from that of alien lambs, so that they can be more reliably linked to different behavioural responses via central brain areas. Parturition also induces a variety of neuroendocrine changes and, in particular, oxytocin release in the olfactory bulb. Using retro-dialysis, oxytocin was infused into the olfactory bulb and produced increases in ACh, NA and GABA release, which was less pronounced in nulliparous sheep (Lévy et al., 1993). This may partly explain why maternally inexperienced sheep require a longer period to bond with their first-born lambs. However, the overall effects of oxytocin in the MOB are likely to be complex. Oxytocin depresses spontaneous GABA inhibitory post-synaptic currents in cultured rat mitral cells and reversibly reduces the frequency of IPSCs without affecting their amplitude (Osako et al., 2000). This effect is mimicked by the OT receptor agonist in a reversible manner and is blocked by the OT receptor antagonist. OT has no effect on the membrane currents evoked by the exogenous application of GABA. These results demonstrate that OT depresses GABA receptor mediated IPSCs by a pre-synaptic mechanism that would be expected to have a disinhibitory effect on mitral cells, increasing glutamatergic input to associated granule cell synapses. This is likely to enhance long-term potentiation at the mitral to granule cell synapse, as has been demonstrated in slices of mouse AOB (Fang et al., 2008). The behaviour ensuring selective acceptance requires the olfactory sensory changes in the MOB at birth to be directed to higher order processing. In part, this processing engages the medial prefrontal cortex, as revealed by c-fos mapping (Broad et al., 2002). In the medial prefrontal cortex, levels of c-fos expression in the pyramidal output layer 5 were found to be significantly higher in ewes that rejected strange lambs. Retro-dialysis of tetracaine anaesthetic, into the medial prefrontal cortex, reduced aggressive motor responses to strange lambs and, during the period of memory formation, also prevented the normal selective expression of aggressive rejection (Broad et al., 2002). The medial prefrontal cortex appears to serve as a sensorymotor link critical for the guidance of reward-related behaviour and for setting of mood. These infusions of tetracaine to the medial prefrontal cortex did not prevent the formation of recognition memory since rejections of the strange lamb reappeared after termination of local anaesthetic infusions. Thus a functional medial prefrontal cortex is not engaged in olfactory recognition or the mediation of proactive maternal behaviour but is required for aggressive rejection responses to odour cues from strange lambs. This raises the question of whether selective lamb recognition could be explained by a selective gating of the own lamb odour signal, preventing it from eliciting aggressive behaviour and enabling the default acceptance behaviour to proceed. This theory is certainly consistent with sheep that lack any olfactory input as a result of anosmia, but which accept both own and alien lambs (Lévy et al., 1995). A similar gating of MOB mitral cell activity by oxytocinergic and vasopressinergic receptors has been proposed to account for social odour recognition in rats (Wacker and Ludwig, 2012). 4. Pheromonal learning in the vomeronasal system A similar selective gating of learned chemosignals has been proposed to explain the vomeronasal mechanisms, at the level of the

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AOB, underlying mate recognition in mice. The AOB has a relatively simple trilaminar structure similar to the MOB. Vomeronasal sensory neurons express only one receptor type from either the V1 or V2 families of receptor genes and converge their axons onto 5–20 glomeruli that receive input from the same receptor type (Belluscio et al., 1999; Rodriguez et al., 1999). The apical dendrites of mitral cell projection neurons in the AOB form complex dendrodendritic synapses with periglomerular cells. However, unlike mitral cells of the MOB, AOB mitral cells extend a branched primary dendritic tree to sample information from gomeruli receiving input from different but closely related vomeronasal (V1R) neurons (Wagner et al., 2006). This selective-heterotypic glomerular connectivity of mitral cells potentially enables them to integrate information among different vomeronasal receptor types, leading to combinatorial responses of mitral cells, such as responses recorded to specific combinations of sex and strain of anaesthetised conspecifics (Luo et al., 2003). In comparison with the extensive lateral dendritic tree of MOB mitral cells, AOB mitral cells have relatively poorly developed secondary dendrites (Larriva-Sahd, 2008; Mori, 1987). Therefore AOB mitral cells predominantly receive reciprocal dendrodendritic inhibition from GABAergic granule cells onto their primary dendritic trees, in a position suitable to selectively inhibit input from specific glomeruli. Females respond to male urinary pheromones in a variety of ways that enhance their reproductive success. These include early induction of puberty and the induction of oestrus in grouped, oestrus-suppressed females (Vandenbergh, 1969; Whitten, 1956). Such male pheromones, were they to be around after mating, have the capacity to activate those same neuroendocrine mechanisms resulting in loss of pregnancy. However, female mice from a recognition memory to the pheromones of the copulating male during a sensitive period in the few hours immediately after mating. The subsequent recognition of the familiar mating male pheromones is hypothesised to selectively gate their transmission and thereby prevent pregnancy loss (Fig. 2) (Brennan et al., 1990; Keverne and de la Riva, 1982). Pheromones from unfamiliar males for which no memory has been formed, are able to block pregnancy during a critical window that extends into the early post-implantation stages of pregnancy (Bruce, 1961). This recognition memory is contingent on mating induced noradrenaline release in the AOB (Keverne and de la Riva, 1982), and has mechanisms in common with the female’s main olfactory bulb memories for pups, and the specific recognition of own lambs in the synchronised seasonal birth of sheep (Brennan and Kendrick, 2006). Indeed vaginocervical stimulation has been shown to be a more general stimulus promoting social odour recognition. Vaginocervical stimulation prolongs the duration of social recognition in rats in the juvenile recognition paradigm and this effect depended on oestrus state and could be blocked by infusions of antagonists of oxytocin or beta adrenergic receptors in the MOB (Larrazolo-Lopez et al., 2008). The mechanism by which noradrenaline induces memory formation in the AOB is likely to be complex and to involve the activation of alpha adrenergic receptors rather than the requirement for beta adrenergic transmission often found in the MOB. Alpha 1 and alpha 2 adrenergic receptors are found on both mitral and granule cells of the AOB and have broadly opposing effects (Dong et al., 2009). Alpha 1 receptor activation increases inhibition of mitral cells (Araneda and Firestein, 2006; Smith et al., 2009), whereas alpha 2 receptor activation decreases inhibitory feedback from granule cells, disinhibiting mitral cells (Dong et al., 2009). It is likely that the alpha 2 receptors are involved with mate recognition learning, as long-term potentiation in the AOB is enhanced by alpha 2 agonists and blocked by alpha 2 antagonists (Kaba and Huang, 2005). Infusions of lignocaine locally into the mouse AOB during the sensitive period for memory formation prevent memory formation

Fig. 2. The Bruce effect (selective pregnancy failure) is one of the best known examples of olfactory imprinting in adult vertebrates. Urine from unfamiliar males (or familiar urine supplemented with unfamiliar MHC peptides) activates neuroendocrine mechanisms leading to pregnancy failure via the accessory olfactory bulb (AOB), amygdala and medial hypothalamus. The mated female learns to recognise the pheromones of the mating male, during a sensitive period around mating. Subsequent exposure to her mate’s urine activates mitral cells (M) in the AOB that are subject to enhanced inhibition from granule cell (G) interneurons. This increased inhibition disrupts the transmission of the mate’s pheromonal signal to the amygdala, preventing pregnancy block.

and subsequent male recognition. However, memory formation was not prevented when lignocaine is infused into the subsequent neural relay in the amygdala (Kaba et al., 1989). This suggests that neural activity in the AOB at the first stage of vomeronasal processing is necessary and sufficient for mate recognition memory formation. Further studies using localised drug infusions into the AOB implicate the dendrodentritic synapses, between glutamatergic mitral and GABA-ergic granule cells of the AOB to be integral to this memory formation (Brennan, 2004; Kaba and Nakanishi, 1995). Memory formation is blocked by inhibition of protein kinase C during the early phase or by inhibition of protein synthesis during a later phase 3–6 h following memory formation (Kaba et al., 1989). Additionally, memory formation is enhanced, but not dependent on nitric oxide signalling in the AOB (Brennan and Kishomoto, 1993; Okere et al., 1996). Morphological analysis of these reciprocal synapses initially found an increased post-synaptic density length of the


excitatory synapses over the first 3–5 days following memory formation (Matsuoka et al., 1998). This increase in excitatory transmission and consequent GABAergic reciprocal feedback is consistent with the increased GABA release in response to the mating male pheromones observed 2 days following mating. This switched to an increase in size of the GABA-ergic inhibitory synapses and a return to normal length for the glutamatergic synapses from 5 to 30 days (Matsuoka et al., 1998), covering the 30-day duration of the memory (Kaba et al., 1988). Both increases in the excitatory and inhibitory sides of the reciprocal synapse increase the overall inhibitory gain of the reciprocal synapse, leading to enhanced self-inhibition of the sub-population of mitral cells activated by the mate’s chemosignals. This is hypothesised to gate the transmission of the mating male’s signal to central vomeronasal areas and prevent pregnancy block. This hypothesis is supported by experiments demonstrating a suppression of activity at central sites along the pregnancyblocking pathway in response to exposure to the learned mating male chemosignals. Recordings from the medial amygdala of awake behaving mice have found that, before mating, individual neurons responded equally to urinary chemosignals from different inbred strains of male. But after mating and memory formation, twice as many spikes were fired in response to unfamiliar male pheromones compared to those of the mating male strain (Binns and Brennan, 2005). Similarly, a lower number of medial amygdala neurons express c-Fos in response to exposure to the mate’s versus unfamiliar male chemosignals (Halem et al., 2001). A lower expression of c-Fos in response to the mating male versus unfamiliar male chemosignals has also been observed in dopaminergic neurons in the arcuate hypothalamus at the neuroendocrine output of the pregnancy block pathway (Matthews et al., 2013). These findings support the hypothesis that memory formation is indeed associated with the selective gating of the familiar chemosensory signal from the mating male, preventing it from activating the neuroendocrine mechanisms of pregnancy block.

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Fig. 3. The medial amygdala (MeA) plays a vital role in social recognition, integrating individuality cues from both vomeronasal and main olfactory systems. It is mediates associative learning between main olfactory an vomeronasal inputs, enabling main olfactory cues to drive innate outputs that are solely driven by vomeronasal cues in naïve animals. The main olfactory and vomeronasal inputs to the MeA are shown, based on data from rodents. Interconnections of individual areas other than the MeA have been omitted for clarity. MOB, main olfactory bulb; AON, anteriorolfactory nucleus; PIR, piriform cortex; NLOT, nucleus of the lateral olfactory tract; ACo, anterior cortical amygdala; PLCo, posterior lateral cortical amygdala; AOB, accessory olfactory bulb; BST, bed nucleus of the stria terminalis; BAOT, bed nucleus of the accessory olfactory tract; PMCo, posterior medial cortical amygdala. Interconnections of the individual structures not involving the MeA have been omitted for clarity.

4.1. The medial amygdala and social recognition The medial amygdala has been found to be important for social recognition across a variety of species and differing behavioural contexts. For instance, local infusions of lignocaine into the cortical and medial, but not the basolateral amygdala prevent the ability of a ewe to selectively recognise and direct maternal behaviour to their own lamb and reject alien lambs (Keller et al., 2004). The medial amygdala has traditionally been regarded as the vomeronasal amygdala, as it was thought to receive direct chemosensory input solely from the vomeronasal system via the accessory olfactory bulb (AOB) (Scalia and Winans, 1975). However, more recent studies in both mice and rats have demonstrated a direct projection from the MOB to the medial amygdala (Kang et al., 2009; ProSistiaga et al., 2007), in addition to multiple indirect pathways (Fig. 3). Functional studies using c-Fos expression as a marker of neuronal activity suggest that this projection is from MOB mitral cells that respond preferentially to social odours rather than non-social odours (Kang et al., 2009). Interestingly the MOB and AOB inputs to the medial amygdala largely overlap, but terminate in distinct sublaminae, suggesting that an individual medial amygdala neuron receives direct convergent input from both main olfactory and vomeronasal systems. Recordings of neural activity from the medial amygdala of hamsters found cells that responded to stimulation of the main olfactory or vomeronasal systems, demonstrating the convergence of information from these two chemosensory systems at the level of the medial amygdala (Licht and Meredith, 1987; Meredith, 1998). Furthermore, these olfactory tract inputs from the AOB and/or MOB to the medial amygdala undergo long-term

potentiation of the postsynaptic field potential in mouse AOB slices in vitro, which is enhanced in the presence of oxytocin (PAB unpublished observations). AOB input to the medial amygdala appears to be highly distributed, with as few as 4 AOB mitral cells sending projections throughout the extent of the medial amygdala (von Campenhausen and Mori, 2000). These similarities of the highly distributed MOB projection to anterior piriform cortex make the medial amygdala a prime candidate for oxytocin-dependent association of individuality chemosignals into an olfactory construct characteristic of the individual. Not only is the medial amygdala in a position to associate the pattern of AOB mitral cells that convey the individual vomeronasal signature of a mouse, based on the profile of MHC peptides (Leinders-Zufall et al., 2004) and/or major urinary proteins (MUPs) (Hurst et al., 2001), but also to associate this with the more variable individual profile of volatile odourants sensed by the main olfactory system. Thus, whereas initial investigation of an individual is likely to involve direct contact for involatile cues of individuality to be sensed by the vomeronasal system, subsequently individual recognition can occur at a distance based on main olfactory cues alone. Such an association of main olfactory with vomeronasal inputs is likely to be involved in the context of recognition of the individuality of urine marks by female mice. Female mice have an innate attraction to investigate and spend time in proximity to male urine marks. This is due to the attractant effects of an atypical MUP named “darcin” secreted in the male mouse urine (Roberts et al., 2010). Darcin not only elicits attraction and investigation, it also

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acts as an unconditioned stimulus to induce the learning of the associated odour of urinary volatiles, presumably sensed by the main olfactory system (Roberts et al., 2010). Their association with darcin appears to confer on the individual volatile odour profile the attractive properties of darcin itself, and darcin induces a conditioned place preference for the context in which the urine mark was sensed (Roberts et al., 2012). Similarly, sexually-naïve male hamsters had deficits in mating behaviour if their vomeronasal organ had been surgically removed (Meredith, 1986). However, sexual behaviour was normal in hamsters that experienced a single mating experience before vomeronasal organ removal. The main olfactory cues from the receptive female had become associated with the vomeronasal cues at the initial mating experience and were now able to drive mating behaviour in the absence of vomeronasal cues (Meredith, 1986). Although the medial amygdala has been identified as a major hub in the social learning network, the distributed nature of olfactory learning means that there are also important roles in social odour learning for other brain regions. This is particularly the case for the olfactory bulb in contexts that could be termed “life events” in which it is vital that a robust individual discrimination occurs to match a major change in behavioural priorities.

5. Neurogenesis and olfactory learning Chemosensory neurons are exposed to the vagaries of the external environment and have a relatively short lifespan. Consequently, these receptor neurons undergo regenerative turnover, which continues throughout life. Turnover of sensory neurons can provide adaptability to new environments by selection for different receptor types and the formation of new memories (Moreno et al., 2009). Individual olfactory sensitivity is dependent on prior olfactory experience, as the activity of OSNs prolongs their lifespan. Recently, the expression of the Hist2h2be variant of histone 2B (H2BE), which is only expressed by OSNs, has been shown to be negatively regulated by neural activity. High levels of H2BE, in inactive OSNs, shortens their lifespan, whereas knockout of the H2be greatly extends the lifetime of OSNs (Santoro and Dulac, 2012). This ‘use it’ or ‘lose it’ neuronal survival strategy adapts olfactory sensitivity to the olfactory environment and may explain changes in the frequency of certain OSN receptor types with age (Rodriguez-Gil et al., 2010). This raises the question as to how the balance between retaining “old” memories and prioritising new memories occurs. In addition to the neurogenesis of OSNs, there is the added complication of neurogenesis at the initial stage of neural processing, with precursor GABA-ergic neurons migrating to the olfactory bulb with the rostral flow from the sub-ventricular zone. This has been extensively studied for the main olfactory system where, in the adult mouse, a thousand new neurons integrate daily into the MOB (Imayoshi et al., 2008). Moreover, the number of neurons migrating and surviving is increased in animals housed in an odour-enriched environment. Thus mice display improved specific olfactory performance in an odour enriched environment, without any change in their spatial learning abilities (Rochefort et al., 2002). Sensory enrichment increases adult born GABA-ergic neuron survival and dendritic development (Livneh et al., 2009) while sensory deprivation decreases survival time of these neurons, illustrating the dynamic nature of olfactory bulb circuitry dependent on active stimulation. The discrimination of perceptually similar odours improves in mice after repeated exposure and is accompanied by increased survival of newborn GABA-ergic neurons within the olfactory bulb. Blocking neurogenesis in the rostral sub-ventricular flow prevents any learned improvement in discrimination (Lepousez et al., 2013).

However, the dependence of odour learning and discrimination on neurogenesis may depend on the type of odour learning paradigm with simple odour discriminations unaffected by blockade of neurogenesis (Imayoshi et al., 2008; Sakamoto et al., 2011). Instead, neurogenesis may be more important for operant, action-based learning (Mandairon et al., 2011). It is notable that the two regions of the adult mammalian brain in which neurogenesis is most prominent, the olfactory bulb and the hippocampal dentate gyrus, each function as pattern separators. The addition of highly-plastic new neurons to these circuits during learning may enhance the ability of the circuits to generate distinct patterns of output in response to the learned stimulus, thereby minimising interference with previously learned information (Wiskott et al., 2006). How the precursor GABA-ergic neurons survive and develop following their migration to the olfactory bulb is dependent on their integration into active circuits. There appears to be a critical time window around days 14–20 during neuronal development for sensory experience to be effective in rescuing and integrating newborn neurons into MOB circuits (Yamaguchi and Mori, 2005). Ablation of the GABA␣2 subunit of migrating neuroblasts did not affect their survival, but dramatically delayed maturation of these neurons, which exhibited reduced dendritic branching and spine density (Pallotto et al., 2012). Spine loss appeared due to reduced GABAergic post-synaptic currents, illustrating the importance of neural activity through GABA-ergic signalling for structural maturation and formation of reciprocal glutamatergic synapses. Selective expression of channel-rhodopsin in these migrating neurons has been used to investigate their activity-dependent integration in the adult olfactory bulb. Improvements in difficult odour-discrimination learning and memory only occur when photo-activation is simultaneous with odour presentation (Alonso et al., 2012). Photo-activation at 40 Hz facilitates learning, a stimulus frequency that also enhances GABA-ergic inhibition in vivo. One important context for integration of adult born neurons is seen in the olfactory bulb of lactating mothers. In vivo time lapse imagining of adult born GABA-ergic neurons revealed their dendritic spines to be more stable in lactating mothers. Spine density was however lower in lactating mothers while the density of their pre-synaptic components was higher. These structural features illustrate enhanced integration of adult born GABA-ergic granule cells into the bulbar circuitry of lactating mothers (Kopel et al., 2012). Retrograde monosynaptic tracing has also revealed that adult-generated neurons first receive local connections from GABA-ergic interneurons before long-range connections become established with the anterior olfactory nucleus and piriform cortex (Deshpande et al., 2013). Parturition and interaction with lambs has also been found to affect the survival and maturation of newborn olfactory bulb neurons in post partum sheep. The post-partum period was associated with a decrease in number of neuroblasts in the MOB granule cell layer of ewes that had interacted with their lamb. However, dendritic length and the number of nodes were enhanced in granule cell layer neuroblasts by lamb interaction. This suggests that post-partum lamb odour learning, which is required for selective lamb recognition, is associated with enhanced incorporation of new inhibitory interneurons into existing bulbar circuits.

6. Vomeronasal receptor neuron turnover Since the VSNs undergo continual cell death and replacement the question arises as to how long a memory trace at the first relay in the AOB sustains female mouse recognition for the male that mated. Experiments have shown that this recognition memory is sustained at 10, 20 and 30 days after mating, but at 50 days


the memory trace is sufficiently changed as to make the familiar male that mated be now treated as strange (Kaba et al., 1988). If the female is allowed to remain pregnant (21 days) the memory duration is foreshortened and its circuitry made available to new memory formation at post-partum oestrous. This foreshortening of recognition memory by pregnancy may be duplicated by administering the hormones of pregnancy (Kaba et al., 1988). Since pregnancy increases nerve growth factors in saliva, which enters the VNO during licking and grooming, this may account for the increased neurogenesis observed in the VNO. Pregnancy is also a period when female mouse urine contains high levels of MUPs (lipocalins), which promote generation of vomeronasal (V2R) stem cells, and accelerate their turnover. This provides an example of how neural pheromone memory is adapted to ensuring reproductive success of the female by enhancing receptor turnover and providing a new template for re-establishment of recognition memory according to female endocrine state. The vomeronasal (V2R) neurons respond to non-volatile urinary pheromones which access the VNO via a pump-action caused by vasodilation of the vomeronasal blood vessels. Urine from adult male mice contains small, testosterone-dependent, volatile compounds, which are transported in urine bound to MUPs. These MUPs are present at high levels in reproductively active male mouse urine of up to 30 mg/ml (Hurst and Beynon, 2004). MUPs are sufficiently polymorphic to form an identification code for each individual, which governs territorial marking and inbreeding avoidance (Hurst et al., 2001). Moreover, male mouse urine and its MUP fraction in particular, are effective in promoting proliferation and survival of V2R neurons (Xia et al., 2006). There is no such neural activation by artificial urine or mouse urine stripped of proteins. This neural activation involves phosphorylation of neuronal Erk, Akt, and Creb signalling, and is effective at very low protein concentrations in urine, being dependent on dose of MUP proteins (Xia et al., 2006). In neural stem cells it has been shown that PI3-Akt and Erk pathways subserve distinct functions with Akt signalling being essential for progenitor survival and Erk signalling for promoting vomeronasal stem cell neurogenesis. These findings have shown that molecules signalling relevant biological information to the mature V2Rs are at the same time required for the survival and proliferation of

Male strain only

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regenerating vomeronasal receptors. This may not only sustain female pheromonal memory, but also has the potential to adapt and optimise this memory formation according to the pheromone signals in the social environment. To address this potential, newborn mice were reared for the first 18 days of life by their genetic mother but in the presence of male urine from a different strain (Broad and Keverne, 2012; Papes et al., 2010). When adult, an enhanced behavioural preference was shown for this non-kin urine, even when competing with same kin urine. This change in preference was accompanied by significant epigenetic changes in gene expression in the VNO. 765 VNO genes were expressed differentially according to strain, and this increased to 901 with early exposure to the urine of the different male strain. V1R and the olfactory receptor genes were most affected by urine from a different strain and this was most pronounced in males (Fig. 4). It is not just the receptor neurons of the vomeronasal system that undergo turnover, but the local GABA-ergic neurons of the accessory olfactory bulb are also continuously replaced by immature precursors derived from the sub-ventricular zone stem cells. Similarly to the established role of neurogenesis in olfactory learning in the MOB, mate recognition memory formation critically depends on the presence of newborn granule cells entering the AOB (Oboti et al., 2011). Exposure to male chemosignals increased the number of newborn neurons incorporating into mature AOB circuits. Moreover, inhibition of neurogenesis, by local infusion of anti-mitotics into the subventricular zone, eliminated the addition of newborn neurons to the AOB and prevented mate recognition memory formation. Interestingly, the incorporation of newborn neurons into AOB is reduced in mice with neurotoxic lesions of the medial amygdala, suggesting that the addition of new neurons to the AOB was dependent on activity in the medial amygdala following mating. This potentially conflicts with earlier findings that memory formation is unaffected by lignocaine silencing of the medial amygdala, during the sensitive period for memory formation just after mating (Kaba et al., 1989). However, it may be that the sensitive period for amygdala activation to enhance neuronal incorporation into the AOB occurs at a much later time point than that affected by the immediately post-mating lignocaine infusions.

Male strain and treatment V1r receptor genes V2r receptor genes olfactory receptor genes urinary protein genes

Female strain only

Female strain and treatment

Fig. 4. Functional categories of genes involved in olfactory detection that are differentially expressed as a function of strain (control C57BL/6J vs control 129sv) or strain and treatment (treated C57BL/6J vs treated 129sv). Data is expressed as a percentage of total genes that are differentially expressed.

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Alternatively, centrifugal input from the medial amygdala may play a long-term permissive role enabling the incorporation of new neurons into the AOB, which would be disrupted following irreversible lesioning. 7. Conclusions In most mammalian species, reproduction is largely dependent on chemosensory cues detected by exceptionally large gene families, the largest in the mammalian genome. The significant interspecies differences in reproductive strategy and the rapid rate of evolution of chemosensory receptors means that caution is needed in generalising findings from one species to other mammals. However, the basic functions of the olfactory and vomeronasal system, and their complementary involvement in both innate and learned responses, is common across mammals. Indeed, chemosensory cues are processed primarily by relatively simple paleocortical circuits. The olfactory bulb functions as a pattern separator, enhancing the discriminability of output patterns during learning that can be linked to the appropriate response by more central olfactory brain areas. There are separate brain systems for handling non-social and social odours, in which the medial amygdala plays a vital role in integrating vomeronasal and main olfactory individuality cues, enabling the recognition of individual conspecifics. Unlike the visual and auditory system in which learning occurs via modification and addition of synapses to existing neurons, the enhanced opportunities of plasticity provided by neurogenesis, at both the level of the sensory epithelium and the GABA-ergic interneurons in the olfactory bulb is vital for learning and discriminating the association of complex odour features. This neurogenesis enables olfactory systems to respond in a more adaptive, as well as deterministic way. Chemosensory learning and memory, particularly in the social context, is integral to the lifestyle of small brain mammals. It is efficient, adaptive and ensures reproductive success. While being economical in the use of neural space, chemosensory learning is extravagant in the vast deployment of chemoreceptor genes for olfactory and pheromonal responding. In primate mammals, the gene pool for olfactory receptors has diminished while most of the vomeronasal receptor genes and their specific ion channels are non-functional pseudogenes. With evolution of the neocortex, auditory and visual information have taken over as the primary sensory systems for decision making, decisions which change to the very same sensory information over developmental time. Knowledge and experience are integral to decision making and behavioural outcomes involving intelligent strategies. The sensory cues themselves are no more than a key to accessing a multitude of context and recognition memories. References Alonso, M., Lepousez, G., Sebastien, W., Bardy, C., Gabellec, M.M., Torquet, N., Lledo, P.M., 2012. Activation of adult-born neurons facilitates learning and memory. Nat. Neurosci. 15, 897–904. Araneda, R.C., Firestein, S., 2006. Adrenergic enhancement of inhibitory transmission in the accessory olfactory bulb. J. Neurosci. 26, 3292–3298. Barnes, D.C., Hofacer, R.D., Zaman, A.R., Rennaker, R.L., Wilson, D.A., 2008. Olfactory perceptual stability and discrimination. Nat. Neurosci. 11, 1378–1380. Belluscio, L., Koentges, G., Axel, R., Dulac, C., 1999. A map of pheromone receptor activation in the mammalian brain. Cell 97, 209–220. Ben-Shaul, Y., Katz, L.C., Mooney, R., Dulac, C., 2010. In vivo vomeronasal stimulation reveals sensory encoding of conspecific and allospecific cues by the mouse accessory olfactory bulb. Proc. Natl. Acad. Sci. U. S. A. 107, 5172–5177. Binns, K.E., Brennan, P.A., 2005. Changes in electrophysiological activity in the accessory olfactory bulb and medial amygdala associated with mate recognition in mice. Eur. J. Neurosci. 21, 2529–2537. Boyd, A.M., Sturgill, J.F., Poo, C., Isaacson, J.S., 2012. Cortical feedback control of olfactory bulb circuits. Neuron 76, 1161–1174. Brennan, P., Kaba, H., Keverne, E.B., 1990. Olfactory recognition: a simple memory system. Science 250, 1223–1226.

Brennan, P.A., 2004. The nose knows who’s who: chemosensory individuality and mate recognition in mice. Horm. Behav. 46, 231–240. Brennan, P.A., Kendrick, K.M., 2006. Mammalian social odours: attraction and individual recognition. Philos. Trans. R. Soc. B 361, 2061–2078. Brennan, P.A., Keverne, E.B., 1997. Neural mechanisms of mammalian olfactory learning. Prog. Neurobiol. 51, 457–481. Brennan, P.A., Kishomoto, J., 1993. Local inhibition of nitric oxide synthase activity in the accessory olfactory bulb does not prevent the formation of an olfactory memory in mice. Brain Res. 619, 306–312. Broad, K.D., Hinton, M.R., Keverne, E.B., Kendrick, K.M., 2002. Involvement of the medial prefrontal cortex in mediating behavioural responses to odour cues rather than olfactory recognition memory. Neuroscience 114, 715–729. Broad, K.D., Keverne, E.B., 2012. The post-natal chemosensory environment induces epigenetic changes in vomeronasal receptor gene expression and a bias in olfactory preference. Behav. Genet. 42, 461–471. Bruce, H.M., 1961. Time relations in the pregnancy-block induced in mice by strange males. J. Reprod. Fert. 2, 138–142. Chamero, P., Marton, T.F., Logan, D.W., Flanagan, K., Cruz, J.R., Saghatelian, A., Cravatt, B.F., Stowers, L., 2007. Identification of protein pheromones that promote aggressive behaviour. Nature 450, 899–902. Chapuis, J., Wilson, D.A., 2012. Bidirectional plasticity of cortical pattern recognition and behavioral sensory acuity. Nat. Neurosci. 15, 155–161. Dantzer, R., Bluthe, R.M., Koob, G.F., Le Moal, M., 1987. Modulation of social memory in male rats by neurohypophyseal peptides. Psychopharmacology (Berl.) 91, 363–368. Davison, I.G., Ehlers, M.D., 2011. Neural circuit mechanisms for pattern detection and feature combination in olfactory cortex. Neuron 70, 82–94. Deshpande, A., Bergami, M., Ghanem, A., Conzelmann, K., Lepier, A., Gotz, M., Berninger, B., 2013. Retrograde monosynaptic tracing reveals the temporal evolution of inputs onto new neurons in the adult dentate gyrus and olfactory bulb. Proc. Natl. Acad. Sci. U. S. A. 110, E1152–E1161. Distel, H., Hudson, R., 1985. The contribution of the olfactory and tactile modalities to the performance of nipple-search behaviour in newborn rabbits. J. Comp. Physiol. A 157, 599–605. Dluzen, D.E., Muraoka, S., Engelmann, M., Ebner, K., Landgraf, R., 2000. Oxytocin induces preservation of social recognition in male rats by activating alpha adrenoreceptors of the olfactory bulb. Eur. J. Neurosci. 12, 760–766. Dluzen, D.E., Muraoka, S., Engelmann, M., Landgraf, R., 1998a. The effects of infusion of arginine vasopressin, oxytocin, or their antagonists into the olfactory bulb upon social recognition responses in male rats. Peptides 19, 999–1005. Dluzen, D.E., Muraoka, S., Landgraf, R., 1998b. Olfactory bulb norepinephrine depletion abolishes vasopressin and oxytocin preservation of social recognition responses in rats. Neurosci. Lett. 254, 161–164. Dong, C., Godwin, D., Brennan, P., Hedge, A., 2009. Protein kinase C alpha mediates signaling underlying an novel form of synaptic plasticity in the accessory olfactory bulb. Neuroscience 163, 811–824. Doucette, W., Gire, D.H., Whitesell, J., Carmean, V., Lucero, M.T., Restrepo, D., 2011. Associative cortex features in the first olfactory brain relay station. Neuron 69, 1176–1187. Doucette, W., Restrepo, D., 2008. Profound context-dependent plasticity of mitral cell responses in olfactory bulb. PLoS Biol. 6, e258. Døving, K.B., Trotier, D., 1998. Structure and function of the vomeronasal organ. J. Exp. Biol. 21, 2913–2925. Eichenbaum, H., 1998. Using olfaction to study memory. Ann. N. Y. Acad. Sci. 855, 657–669. Fang, L.Y., Quan, R.D., Kaba, H., 2008. Oxytocin facilitates the induction of long-term potentiation in the accessory olfactory bulb. Neurosci. Lett. 438, 133–137. Ferguson, J.N., Aldag, J.M., Insel, T.R., Young, L.J., 2001. Oxytocin in the medial amygdala is essential for social recognition in the mouse. J. Neurosci. 21, 8278–8285. Ferguson, J.N., Young, L.J., Hearn, E.F., Matzuk, M.M., Insel, T.R., Winslow, J.T., 2000. Social amnesia in mice lacking the oxytocin gene. Nat. Genet. 25, 284–288. Ferrero, D.M., Moeller, L.M., Osakada, T., Horio, N., Li, Q., Roy, D.S., Cichy, A., Spehr, M., Touhara, K., Liberles, S.D., 2013. A juvenile mouse pheromone inhibits sexual behaviour through the vomeronasal system. Nature 502, 368–371. Fortin, N.J., Agster, K.L., Eichenbaum, H.B., 2002. Critical role of the hippocampus in memory for sequences of events. Nat. Neurosci. 5, 458–462. Freeman, W.J., Schneider, W., 1982. Changes in spatial patterns of rabbit olfactory EEG with conditioning to odors. Psychophysiology 19, 4456. Haberly, L., 1985. Neuronal circuitry in olfactory cortex: anatomy and functional implications. Chem. Senses 10, 219–238. Haga, S., Hattori, T., Sato, T., Sato, K., Matsuda, S., Kobayakawa, R., Sakano, H., Yoshihara, Y., Kikusui, T., Touhara, K., 2010. The male mouse pheromone ESP1 enhances female sexual receptive behaviour through a specific vomeronasal receptor. Nature 466, 118–122. Halem, H.A., Cherry, J.A., Baum, M.J., 2001. Central forebrain responses to familiar male odors are attenuated in recently mated female mice. Eur. J. Neurosci. 13, 389–399. Hurst, J., Beynon, R., 2004. Scent wars: the chemobiology of competitive signalling in mice. Bioessays 26, 1288–1298. Hurst, J.L., Payne, C.E., Nevison, C.M., Marie, A.D., Humphries, R.E., Robertson, D.H.L., Cavaggioni, A., Beynon, R.J., 2001. Individual recognition in mice mediated by major urinary proteins. Nature 414, 631–634. Ibarra-Soria, X., Levitin, M.O., Logan, D.W., 2014. The genomic basis of vomeronasalmediated behaviour. Mamm. Genome 25, 75–86. Imai, T., Suzuki, M., Sakano, H., 2006. Odorant receptor-derived cAMP signals direct axonal targeting. Science 314, 657–661.

P. Brennan, E.B. Keverne / Neuroscience and Biobehavioral Reviews 50 (2015) 29–40 Imayoshi, I., Sakamoto, M., Ohtsuka, T., Takao, K., Miyakawa, T., Yamaguchi, M., Mori, K., Ikeda, T., Itohara, S., Kageyama, R., 2008. Roles of continuous neurogenesis in the structural and functional integrity of the adult forebrain. Nat. Neurosci. 11, 1153–1161. Isaacson, J.S., Strowbridge, B.W., 1998. Olfactory reciprocal synapses: dendritic signaling in the CNS. Neuron 20, 749–761. Kaba, H., Huang, G.-Z., 2005. Long-term potentiation in the accessory olfactory bulb: a mechanism for olfactory learning. Chem. Senses 30, i150–i151. Kaba, H., Nakanishi, S., 1995. Synaptic mechanisms of olfactory recognition memory. Rev. Neurosci. 6, 125–141. Kaba, H., Rosser, A., Keverne, E.B., 1989. Neural basis of olfactory memory in the context of pregnancy block. Neuroscience 32, 657–662. Kaba, H., Rosser, A.E., Keverne, E.B., 1988. Hormonal enhancement of neurogenesis and its relationship to the duration of olfactory memory. Neuroscience 24, 93–98. Kambere, M.B., 2007. Co-regulation of a large and rapidly evolving repertoire of odorant receptor genes. BMC Neurosci. 8, S2. Kang, N., Baum, M.J., Cherry, J.A., 2009. A direct main olfactory bulb projection to the ‘vomeronasal’ amygdala in female mice selectively responds to volatile pheromones from males. Eur. J. Neurosci. 29, 624–634. Kay, L.M., Laurent, G., 1999. Odor- and context-dependent modulation of mitral cell activity in behaving rats. Nat. Neurosci. 2, 1003–1009. Keller, M., Perrin, G., Meurisse, M., Ferreira, G., Lévy, F., 2004. Cortical and medial amygdala are both involved in the formation of olfactory offspring memory in sheep. Eur. J. Neurosci. 20, 3433–3441. Kendrick, K.M., Levy, F., Keverne, E.B., 1992. Changes in sensory processing of olfactory signals induced by birth in sheep. Science 256, 833–836. Keverne, E.B., de la Riva, C., 1982. Pheromones in mice: reciprocal interaction between the nose and brain. Nature 296, 148–150. Kobayakawa, K., Kobayakawa, R., Matsumoto, H., Oka, Y., Imai, T., Ikawa, M., Okabe, M., Ikeda, T., Itohara, S., Kikusui, T., Mori, K., Sakano, H., 2007. Innate versus learned odour processing in the mouse olfactory bulb. Nature 450, 503–508. Kopel, H., Schechtman, E., Groysman, M., Mizrahi, A., 2012. Enhanced synaptic integration of adult-born neurons in the olfactory bulb of lactating mothers. J. Neurosci. 32, 7519–7527. Larrazolo-Lopez, A., Kendrick, K.M., Aburto-Arciniega, M., Arriaga-Avila, V., Morimoto, S., Frias, M., Guevara-Guzman, R., 2008. Vaginocervical stimulation enhances social recognition memory in rats via oxytocin release in the olfactory bulb. Neuroscience 152, 585–593. Larriva-Sahd, J., 2008. The accessory olfactory bulb in the adult rat: a cytological study of its cell types, neuropil, neuronal modules, and interactions with the main olfactory system. J. Comp. Neurol. 510, 309–350. Leinders-Zufall, T., Brennan, P., Widmayer, P., Chandramani, P.S., Maul-Pavicic, A., Jäger, M., Li, X.-H., Breer, H., Zufall, F., Boehm, T., 2004. MHC class I peptides as chemosensory signals in the vomeronasal organ. Science 306, 1033–1037. Lepousez, G., Valley, M.T., Lledo, P.M., 2013. The impact of adult neurogenesis on olfactory bulb circuits and computations. Annu. Rev. Physiol. 75, 339–363. Lévy, F., Gervais, R., Kindermann, U., Orgeur, P., Piketty, V., 1990. Importance of ␤noradrenergic receptors in the olfactory bulb of sheep for recognition of lambs. Behav. Neurosci. 104, 464–469. Lévy, F., Guevara-Guzman, R., Hinton, M.R., Kendrick, K.M., Keverne, E.B., 1993. Effects of parturition and maternal experience on noradrenaline and acetylcholine release in the olfactory bulb in sheep. Behav. Neurosci. 107, 662–668. Lévy, F., Locatelli, A., Piketty, V., Tillet, Y., Poindron, P., 1995. Involvement of the main but not the accessory olfactory system in maternal behaviour of primiparous and multiparous ewes. Physiol. Behav. 57, 97–104. Li, G., Cleland, T.A., 2013. A two-layer biophysical model of cholinergic neuromodulation in olfactory bulb. J. Neurosci. 33, 3037–3058. Licht, G., Meredith, M., 1987. Convergence of main and accessory olfactory pathways onto single neurons in the hamster amygdala. Exp. Brain Res. 69, 7–18. Livneh, Y., Feinstein, M., Klein, M., Mizrahi, A., 2009. Sensory input enhances synaptogenesis of adult-born neurons. J. Neurosci. 29, 86–97. Luo, M.M., Fee, M.S., Katz, L.C., 2003. Encoding pheromonal signals in the accessory olfactory bulb of behaving mice. Science 299, 1196–1201. Mandairon, N., Sultan, S., Nouvian, M., Sacquet, J., Didier, A., 2011. Involvement of newborn neurons in olfactory associative learning? The operant or nonoperant component of the task makes all the difference. J. Neurosci. 31, 12455–12460. Markopoulos, F., Rokni, D., Gire, D.H., Murthy, V.N., 2012. Functional properties of cortical feedback projections to the olfactory bulb. Neuron 76, 1175–1188. Matsuoka, M., Mori, Y., Ichikawa, M., 1998. Morphological changes of synapses induced by urinary stimulation in the hamster accessory olfactory bulb. Synapse 28, 160–166. Matthews, G.A., Patel, R., Walsh, A., Davies, O., Martinez-Ricos, J., Brennan, P.A., 2013. Mating increases neuronal tyrosine hydroxylase expression and selectively gates transmission of male chemosensory information in female mice. PLOS ONE 8, e69943. Meredith, M., 1986. Vomeronasal organ removal before sexual experience impairs male hamster mating behavior. Physiol. Behav. 36, 737–743. Meredith, M., 1998. Vomeronasal, olfactory, hormonal convergence in the brain. Ann. N. Y. Acad. Sci. 855, 349–361. Meredith, M., O’Connell, R.J., 1979. Efferent control of stimulus access to the hamster vomeronasal organ. J. Physiol. 286, 301–316. Mombaerts, P., Wang, F., Dulac, C., Chao, S.K., Nemes, A., Mendelsohn, M., Edmondson, J., Axel, R., 1996. Visualizing an olfactory sensory map. Cell 87, 675–686.

39

Moreno, M.M., Linster, C., Escanilla, O., Sacquet, J., Didier, A., Mandairon, N., 2009. Olfactory perceptual learning requires adult neurogenesis. Proc. Natl. Acad. Sci. U.S.A. 106, 17980–17985. Mori, K., 1987. Membrane and synaptic properties of identified neurons in the olfactory bulb. Prog. Neurobiol. 29, 275–320. Mori, K., Nagao, H., Yoshihara, Y., 1999. The olfactory bulb: coding and processing of odor molecule information. Science 286, 711–715. Mori, K., Sakano, H., 2011. How is the olfactory map formed and interpreted in the mammalian brain? Annu. Rev. Neurosci. 34, 467–499. Mori, K., Takahashi, Y.K., Igarashi, K.M., Yamaguchi, M., 2006. Maps of odorant molecular features in the mammalian olfactory bulb. Physiol. Rev. 86, 409–433. Nei, M., Niimura, Y., Nozawa, M., 2008. The evolution of animal chemosensory receptor gene repertoires: roles of chance and necessity. Nat. Rev. Genet. 9, 951–963. Niessing, J., Friedrich, R.W., 2010. Olfactory pattern classification by discrete neuronal network states. Nature 465, 47–52. Nissant, A., Pallatto, M., 2011. Integration and maturation of newborn neurons in the adult olfactory bulb – from synapses to function. Eur. J. Neurosci. 33, 1069–1077. Oboti, L., Schellino, R., Giachino, C., Chamero, P., Pyrski, M., Leinders-Zufall, T., Zufall, F., Fasolo, A., Peretto, P., 2011. Newborn interneurons in the accessory olfactory bulb promote mate recognition in female mice. Front. Neurosci. 5, 113. Okere, C.O., Kaba, H., Higuchi, T., 1996. Formation of an olfactory recognition memory in mice: reassessment of the role of nitric oxide. Neuroscience 71, 349–354. Osako, Y., Otsuka, T., Taniguchi, M., Oka, T., Kaba, H., 2000. Oxytocin depresses spontaneous gamma-aminobutyric acid-ergic inhibitory postsynaptic currents in cultured mitral cells of the rat olfactory bulb by a presynaptic mechanism. Neurosci. Lett. 289, 25–28. Pallotto, M., Nissant, A., Fritschy, J.M., Rudolph, U., Sassoe-Pognetto, M., Panzanelli, P., Lledo, P.M., 2012. Early formation of GABAergic synapses governs the development of adult-born neurons in the olfactory bulb. J. Neurosci. 32, 9103–9115. Papes, F., Logan, D.W., Stowers, L., 2010. The vomeronasal organ mediates interspecies defensive behaviors through detection of protein pheromone homologs. Cell 141, 692–703. Pro-Sistiaga, P., Mohedano-Moriano, A., Ubeda-Banon, I., Del Mar Arroyo-Jimenez, M., Marcos, P., Artacho-Perula, E., Crespo, C., Insausti, R., Martinez-Marcos, A., 2007. Convergence of olfactory and vomeronasal projections in the rat basal telencephalon. J. Comp. Neurol. 504, 346–362. Roberts, S.A., Davidson, A.J., McLean, L., Beynon, R.J., Hurst, J.L., 2012. Pheromonal induction of spatial learning in mice. Science 338, 1462–1465. Roberts, S.A., Simpson, D.M., Armstrong, S.D., Davidson, A.J., Robertson, D.H., Mclean, L., Beynon, R.J., Hurst, J.L., 2010. Darcin: a male pheromone that stimulates female memory and sexual attraction to an individual male’s odour. BMC Biol. 8, 75. Rochefort, C., Gheusi, G., Vincent, J.D., Lledo, P.M., 2002. Enriched odor exposure increases the number of newborn neurons in the adult olfactory bulb and improves odor memory. J. Neurosci. 22, 2679–2689. Rodriguez-Gil, D.J., Treloar, H.B., Zhang, X., Miller, A.M., Two, A., Iwema, C., Firestein, S.J., Greer, C.A., 2010. Chromosomal location-dependent nontochastic onset of odor receptor expression. J. Neurosci. 30, 10067–10075. Rodriguez, I., Feinstein, P., Mombaerts, P., 1999. Variable patterns of axonal projections of sensory neurons in the mouse vomeronasal system. Cell 97, 199–208. Rolls, E.T., Critchley, H.D., Mason, R., Wakeman, E.A., 1996. Orbitofrontal cortex neurons: role in olfactory and visual association learning. J. Neurophysiol. 75, 1970–1981. Sakamoto, M., Imayoshi, I., Ohtsuka, T., Yamaguchi, M., Mori, K., Kageyama, R., 2011. Continuous neurogenesis in the adult forebrain is required for innate olfactory responses. Proc. Natl. Acad. Sci. U.S.A. 108, 8479–8484. Santoro, S.W., Dulac, C., 2012. The activity-dependent histone variant H2BE modulates the life span of olactory neurons. Elife 1, e00070. Scalia, F., Winans, S.S., 1975. The differential projections of the olfactory bulb and accessory olfactory bulb in mammals. J. Comp. Neurol. 161, 31–56. Schaal, B., Coureaud, G., Langlois, D., Giniès, C., Sémon, E., Perrier, G., 2003. Chemical and behavioural characterization of the rabbit mammary pheromone. Nature 424, 68–72. Schoenbaum, G., Eichenbaum, H., 1995a. Information coding in the rodent prefrontal cortex. I. Single-neuron activity in orbitofrontal cortex compared with that in pyriform cortex. J. Neurophysiol. 74, 733–750. Schoenbaum, G., Eichenbaum, H., 1995b. Information coding in the rodent prefrontal cortex. II. Ensemble activity in orbitofrontal cortex. J. Neurophysiol. 74, 751–762. Smear, M., Resulaj, A., Zhang, J., Bozza, T., Rinberg, D., 2013. Multiple perceptible signals from a single olfactory glomerulus. Nat. Neurosci. 16, 1687–1691. Smear, M., Shusterman, R., O’Connor, R., Bozza, T., Rinberg, D., 2011. Perception of sniff phase in mouse olfaction. Nature 479, 397–400. Smith, R.S., Weitz, C.J., Araneda, R.C., 2009. Excitatory actions of noradrenaline and metabotropic glutamate receptor activation in granule cells of the accessory olfactory bulb. J. Neurophysiol. 102, 1103–1114. Soucy, E.R., Albeanu, D.F., Fantana, A.L., Murthy, V.N., Meister, M., 2009. Precision and diversity in an odor map on the olfactory bulb. Nat. Neurosci. 12, 210–220. Spehr, M., Kelliher, K., Li, X.-H., Boehm, T., Leinders-Zufall, T., Zufall, F., 2006. Essential role of the main olfactory system in social recognition of major histocompatibility complex peptide ligands. J. Neurosci. 26, 1961–1970. Spors, H., Grinvald, A., 2002. Spatio-temporal dynamics of odor representations in the mammalian olfactory bulb. Neuron 34, 301–315. Stettler, D.D., Axel, R., 2009. Representations of odor in the piriform cortex. Neuron 63, 854–864. Tobin, V.A., Hashimoto, H., Wacker, D.W., Takayanagi, Y., Langnaese, K., Caquineau, C., Noack, J., Landgraf, R., Onaka, T., Leng, G., Meddle, S.L., Engelmann, M., Ludwig,

40


M., 2010. An intrinsic vasopressin system in the olfactory bulb is involved in social recognition. Nature 464, 413–417. Vandenbergh, J., 1969. Male odor accelerates female sexual maturation in mice. Endocrinology 84, 658–660. von Campenhausen, H., Mori, K., 2000. Convergence of segregated pheromonal pathways from the accessory olfactory bulb to the cortex in the mouse. Eur. J. Neurosci. 12, 33–46. Wacker, D.W., Ludwig, M., 2012. Vasopressin, oxytocin, and social odor recognition. Horm. Behav. 61, 259–265. Wagner, S., Gresser, A.L., Torello, A.T., Dulac, C., 2006. A multireceptor genetic approach uncovers an ordered integration of VNO sensory inputs in the accessory olfactory bulb. Neuron 50, 697–709. Whitten, W., 1956. Modification of the oestrous cycle of the mouse by external stimuli associated with the male. J. Endocrinol. 13, 399–404.

Wilson, D.A., 2000. Odor specificity of habituation in the rat anterior piriform cortex. J. Neurophysiol. 83, 139–145. Wiskott, L., Rasch, M.J., Kempermann, G., 2006. A functional hypothesis for adult hippocampal neurogenesis: avoidance of catastrophic interference in the dentate gyrus. Hippocampus 16, 329–343. Xia, J., Sellers, L.A., Oxley, D., Smith, T., Emson, P., Keverne, E.B., 2006. Urinary pheromones promote ERK/Akt phosphorylation, regeneration and survival of vomeronasal (V2R) neurons. Eur. J. Neurosci. 24, 3333–3342. Yamaguchi, M., Mori, K., 2005. Critical period for sensory experience-dependent survival of newly generated granule cells in the adult mouse olfactory bulb. Proc. Natl. Acad. Sci. U.S.A. 102, 9697–9702. Yassa, M.A., Stark, C.E., 2011. Pattern separation in the hippocampus. Trends Neurosci. 34, 515–525.

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