Neurobiology of Learning and Memory xxx (2013) xxx–xxx

Contents lists available at ScienceDirect

Neurobiology of Learning and Memory journal homepage: www.elsevier.com/locate/ynlme

Rapid corticosteroid actions on synaptic plasticity in the mouse basolateral amygdala: Relevance of recent stress history and b-adrenergic signaling R.A. Sarabdjitsingh ⇑, M. Joëls Department of Translational Neuroscience, Brain Center Rudolf Magnus, University Medical Center Utrecht, 3508 AB Utrecht, The Netherlands

a r t i c l e

i n f o

Article history: Available online xxxx Keywords: Stress LTP Basolateral amygdala Corticosterone Propranolol

a b s t r a c t The rodent stress hormone corticosterone rapidly enhances long-term potentiation in the CA1 hippocampal area, but leads to a suppression when acting in a more delayed fashion. Both actions are thought to contribute to stress effects on emotional memory. Emotional memory formation also involves the basolateral amygdala, an important target area for corticosteroid actions. We here (1) investigated the rapid effects of corticosterone on amygdalar synaptic potentiation, (2) determined to what extent these effects depend on the mouse’s recent stress history or (3) on prior b-adrenoceptor activation; earlier studies at the single cell level showed that especially a recent history of stress changes the responsiveness of basolateral amygdala neurons to corticosterone. We report that, unlike the hippocampus, stress enhances amygdalar synaptic potentiation in a slow manner. In vitro exposure to 100 nM corticosterone quickly decreases synaptic potentiation, and causes only transient potentiation in tissue from stressed mice. This transient type of potentiation is also seen when b-adrenoceptors are blocked during stress and this is further exacerbated by subsequent in vitro administered corticosterone. We conclude that stress and corticosterone change synaptic potentiation in the basolateral amygdala in a manner opposite to that seen in the hippocampus and that renewed exposure to corticosterone only allows induction of non-persistent forms of synaptic potentiation. Ó 2013 Elsevier Inc. All rights reserved.

1. Introduction Stress initiates a bodily reaction in response to potential threats in the environment. This physiological response essentially serves to centrally control adaptation to stress, to reinstate homeostasis and facilitate neuronal plasticity for the storage of stress-related information for future use (de Kloet, Joels, & Holsboer, 2005; McEwen, 2007; Sapolsky, Romero, & Munck, 2000). Excessive stress exposure can however seriously threaten homeostatic control, affect brain functioning and lower the threshold for the precipitation of psychopathology (de Kloet et al., 2005; Herbert et al., 2006). A hallmark of the neuroendocrine response to stress is the acute and central elevation in levels of the catecholamine noradrenaline

Abbreviations: BLA, basolateral amygdala; GR, glucocorticoid receptor; HPA, hypothalamic pituitary adrenal; LTP, long-term potentiation; MR, mineralocorticoid receptor; NA, noradrenaline. ⇑ Corresponding author. Address: Department of Translational Neuroscience, Brain Center Rudolf Magnus, University Medical Center Utrecht, Universiteitsweg 100, 3584 CG Utrecht, The Netherlands. Fax: +31 (0)88 75 69032. E-mail address: [email protected] (R.A. Sarabdjitsingh).

(NA) by tonic activation of the locus coeruleus and the nucleus tractus solitarius. This neurotransmitter induces a surge of vigilance and state of alertness, primarily mediated via the amygdala, and acts predominantly via b-adrenergic receptors (Gibbs & Summers, 2002; McGaugh, 2004; Roozendaal, McEwen, & Chattarji, 2009). Slightly later, activation of the hypothalamic– pituitary–adrenal (HPA) axis results in the increased release of corticosteroid hormones (cortisol in humans, corticosterone in rodents) by the adrenal cortex into the blood stream (Ulrich-Lai & Herman, 2009). Due to their lipophilic nature and small size, these steroid hormones readily enter the brain and exert their modulatory actions via binding to the mineralocorticoid (MR) and glucocorticoid receptor (GR). MR and GR are abundantly expressed in brain regions essential for learning and memory processes, e.g. the hippocampus and amygdala (Reul & de Kloet, 1985; Sapolsky, McEwen, & Rainbow, 1983). Mechanistic studies have delineated time-dependent and region-specific effects of corticosterone on neuronal physiology (Joels, Sarabdjitsingh, & Karst, 2012; Popoli, Yan, McEwen, & Sanacora, 2012). These are considered to occur via rapid non-genomic and slow genomic effects by MR and GR variants residing in the membrane and nuclear compartment, respectively.

1074-7427/$ - see front matter Ó 2013 Elsevier Inc. All rights reserved. http://dx.doi.org/10.1016/j.nlm.2013.10.011

Please cite this article in press as: Sarabdjitsingh, R. A., & Joëls, M. Rapid corticosteroid actions on synaptic plasticity in the mouse basolateral amygdala: Relevance of recent stress history and b-adrenergic signaling. Neurobiology of Learning and Memory (2013), http://dx.doi.org/10.1016/j.nlm.2013.10.011

2

R.A. Sarabdjitsingh, M. Joëls / Neurobiology of Learning and Memory xxx (2013) xxx–xxx

Despite their sequential release, NA and corticosteroids tightly work in concert to orchestrate the physiological response to stress and profoundly affect memory formation and cognitive performance (Joels & Baram, 2009; McGaugh, 2013; Roozendaal et al., 2009). Especially the rapid corticosterone are thought to amplify the effects of NA and are considered to promote rapid neuronal excitability and behavioral adaptation to stress (Krugers, Karst, & Joels, 2012). The underlying neurobiological processes have been rather well characterized for the hippocampus. In the basolateral amygdala (BLA) however, some questions still remain unresolved regarding the rapid and slow actions of corticosterone and how this depends on noradrenergic transmission. Earlier findings showed that a single exposure to stress steadily increases long-term potentiation (LTP) in the BLA (Kavushansky & Richter-Levin, 2006; Maroun, 2006; Rodriguez Manzanares, Isoardi, Carrer, & Molina, 2005; Sarabdjitsingh, Kofink, Karst, de Kloet, & Joels, 2012; (Vouimba, Munoz, & Diamond, 2006). Pharmacological blockage of MR or GR attenuated the induction of LTP some hours later, supporting involvement of genomic corticosteroid signaling (Sarabdjitsingh et al., 2012). These facilitating genomic actions are opposite to those previously described for GR activation in the hippocampus (Groc, Choquet, & Chaouloff, 2008; Karst et al., 2005; Martin et al., 2009). Rapid effects of corticosterone have also been described in the BLA, but so far only at the single cell level (Karst, Berger, Erdmann, Schutz, & Joels, 2010). More specifically, BLA neurons were found to respond with enhanced glutamatergic transmission to a first corticosterone pulse (or stress) but exhibited reduced glutamatergic transmission when exposed to a second pulse of corticosterone >1 h after the first; this switch in response has been referred to as ‘metaplasticity’. At present it is not clear whether these rapid effects -and metaplasticity- also persist at the level of LTP in BLA circuits. We have previously also demonstrated that b-adrenoceptors play an important role in enhancing and stabilizing synaptic plasticity in the BLA. Activation of b-adrenoceptors by isoproterenol was found to increase LTP in vitro and, conversely, blockade of this receptor with propranolol reduced the stress-induced enhancement of LTP in the BLA (Pu, Krugers, & Joels, 2009; Sarabdjitsingh et al., 2012). Yet, remarkably little is known of how corticosterone rapidly interacts with stress and the noradrenergic system to regulate neuronal excitability and synaptic strengthening. Therefore, in this study we aimed to examine (1) the rapid effect of corticosterone on LTP formation in the BLA and (2) whether this is affected by recent stress exposure. Furthermore, we extended previous studies on the role of the b-adrenoceptors in synaptic plasticity. Hereto we studied (3) whether the rapid effects of corticosterone depend on the activation of the b-adrenoceptor under control or stress conditions. 2. Materials and methods 2.1. Animals Male C57/Bl6 mice (Harlan, the Netherlands; 5–6 weeks old at the moment of arrival) were group-housed in a temperature- and humidity-controlled room with food and water available ad libitum. After their arrival, animals were left undisturbed in their room to acclimatize for at least 1 week. Mice were killed early in the morning when endogenously circulating corticosterone levels are low.

of Utrecht (Permit number 2010.I.11.236). All efforts were made to minimize suffering. 2.3. Experimental design 2.3.1. Effect of restraint stress on LTP in the BLA and CA1 region In Experiment 1, we investigated the effect of stress on LTP in the BLA and hippocampus. Earlier we have reported that restraint stress is very effective in enhancing synaptic plasticity in the BLA (Sarabdjitsingh et al., 2012). In the current study we additionally investigated the effect of restraint stress in the CA1 region of the hippocampus. Restraint stress was applied by placing mice for 20 min individually in a transparent plexiglas restraining cylinder (5 cm diameter) provided with ample air holes for ventilation. A small round-shaped lid was used to fixate the mouse inside the tube, thereby preventing movement but allowing the animal to breathe freely; control mice were left undisturbed. Directly after the stressor, the mice were taken out of the restrainer, rapidly decapitated and the brains were sliced. Field potential recordings were performed in brain slices containing either the LA-BLA pathway (as described in detail below) or in the Schaffer collateral-CA1 pathway of the hippocampus. Briefly, evoked baseline field potentials were recorded for 15–20 min after which high-frequency stimulation was applied (see below for details). After tetanization, field responses were recorded for another 60 min. 2.3.2. Rapid effects of corticosterone on BLA-LTP In Experiment 2, we aimed to investigate the rapid effects of corticosterone on LTP formation in the BLA. For this purpose, baseline synaptic field responses were monitored for 10 min, followed by perfusion with 100 nM corticosterone (Sigma–Aldrich, Germany) into artificial cerebrospinal fluid (aCSF) for 20 min. Perfusion co-terminated with a tetanic stimulation (see below), after which fEPSP responses were monitored for another 60 min. Previously, Karst et al. (2010) has shown that metaplastic responses to corticosterone exist at least at the single cell level depending on the stress history of an animal. Therefore, we also studied the effect of acutely applied corticosterone on LTP in mice that were first stressed. 2.3.3. Effect of corticosterone on BLA-LTP in propranolol-pretreated mice The effect of propranolol on BLA LTP under stress and control conditions has been reported previously (Sarabdjitsingh et al., 2012). In Experiment 3, we investigated the rapid effects of corticosterone on BLA LTP after blockade of b-adrenoceptors. To block the b-adrenoceptor, animals were pretreated intraperitoneally with propranolol (10 mg/kg i.p., Sigma–Aldrich, Germany) or vehicle (0.9% NaCl), respectively. The drug dose was chosen based on literature shown to effectively block b-adrenoceptor mediated effects (Adamec, Muir, Grimes, & Pearcey, 2007; Sarabdjitsingh et al., 2012). Thirty minutes later and similarly as described for experiment 1 and 2, animals were exposed to either 20 min of restraint stress or left undisturbed and subsequently rapidly decapitated. Similar as described above, a 10 min baseline was recorded after which 100 nM corticosterone (20 min) was perfused onto brain slices of control or stressed mice. After application of high frequency stimulation (HFS), field potential responses were monitored for another 60 min. 2.4. Electrophysiology

2.2. Ethics statement All experiments were approved and conducted according to the guidelines of the Animal Committee for Bioethics of the University

After rapid dissection, the brain was chilled in ice-cold artificial cerebrospinal fluid (aCSF) consisting of 120 mM NaCl, 3.5 mM KCl, 5.0 mM MgSO4, 1.25 mM NaHPO4, 0.2 mM CaCl2, 10 mM D-glucose,

Please cite this article in press as: Sarabdjitsingh, R. A., & Joëls, M. Rapid corticosteroid actions on synaptic plasticity in the mouse basolateral amygdala: Relevance of recent stress history and b-adrenergic signaling. Neurobiology of Learning and Memory (2013), http://dx.doi.org/10.1016/j.nlm.2013.10.011

R.A. Sarabdjitsingh, M. Joëls / Neurobiology of Learning and Memory xxx (2013) xxx–xxx

3

and 25 mM NaHCO3, gassed with 95% O2 and 5% CO2. Coronal brain slices (350 lm thick) containing either the hippocampus or the BLA were prepared using a Leica VT1000S Vibratome. All slices were collected and submerged in aCSF in a holding chamber for 1–4 h before being transferred to the recording chamber and were maintained at 32 °C. As previously described (Pu et al., 2009; Sarabdjitsingh et al., 2012; Wiegert, Joels, & Krugers, 2006), BLA or CA1 field excitatory postsynaptic potentials (fEPSP) were evoked by stimuli delivered to the afferent fibers via a bipolar tungsten electrode insulated to the tip (0.075 mm lm tip diameter). For BLA, the electrode was positioned in the lateral amygdala which supplies one of the major afferent pathways to the BLA (Fig. 1A) (Pitkanen et al., 1995), while for CA1, the stimulation electrode was placed in the Schaffer collaterals. For recording, glass microelectrodes filled with aCSF (2– 3 MX) were used. Single pulses (0.15 ms) were delivered at a rate of once per 30 s (Neurolog digital stimulator, Cambridge Electronic Design, United Kingdom) and amplified with a gain of 1000. The stimulation intensity was adjusted to produce a fEPSP of approximately 50% of the maximal amplitude. Tetanic stimulation was applied only when responses to single stimuli had remained stable for at least 20 min. Subsequently, as previously described (Pu et al., 2009; Sarabdjitsingh et al., 2012; Wiegert et al., 2006) stable non-saturated LTP was induced by applying one train of high-frequency stimulation (100 Hz, 1 s for BLA or 10 Hz, 90 s for CA1). Synaptic responses were further monitored for 60 min post-tetanus. For CA1, fEPSP the slope was calculated while the amplitude of the BLA fEPSP was calculated as (a + b)/2 with (a) being the difference between the sharp negative voltage deflection at the onset and the negative peak, and (b) the difference between the negative peak and the succeeding positive peak (Fig. 1B) (Huge, Rammes, Beyer, Zieglgansberger, & Azad, 2009; Rammes et al., 2000). Two consecutive traces were averaged to represent the mean per minute. Data were acquired, stored, and analyzed using Signal 2.16 (Cambridge Electronic Design, United Kingdom). Changes in synaptic strength were expressed relative to normalized baseline and expressed as mean ± SEM.

t-test was used for within-group comparisons (1) before vs after tetanization and (2) before and during 100 nM corticosterone perfusion where appropriate. For between-group comparisons either a two-tailed unpaired Student’s t-test, a two-way ANOVA or a 2  2  2 factor ANOVA was used with stress, propranolol and/or corticosterone perfusion as between-subject factors. When applicable, pairwise post hoc comparisons were carried out using a Tukey’s or LSD post hoc test, as indicated in the text. Probability values of p < 0.05 were considered to represent significant differences.

2.5. Data analysis

3.2. Rapid effects of corticosterone on BLA synaptic plasticity

All statistical analyses were carried out with SPSS version 16.0 (SPSS, Gorinchem, The Netherlands). A two-tailed paired Student’s

Next, we wondered if and how corticosterone would rapidly affect BLA-LTP. For this purpose we perfused brain slices from control mice with 100 nM corticosterone for 20 min (Fig. 3A). Within-group comparison to the pre-corticosterone baseline (t = 30 to 20 min) showed that corticosterone application itself did not alter baseline properties (Fig. 3A; mean baseline fEPSP ± SEM before vs during perfusion: 98.4 ± 1.6% vs 101.2 ± 2.4%, p > 0.05). Corticosterone application co-terminated with HFS. Tetanization resulted in moderate, though stable, levels of potentiation in control slices (Fig. 3A). The average synaptic response over the 60 min period amounted to 125.8 ± 3.4% (mean fEPSP amplitude ± SEM, p < 0.01 compared to pre-tetanus baseline), and was significantly lower than seen in control slices (p < 0.05; Fig. 3A and B). These data suggests that corticosterone rapidly decreases the level of synaptic plasticity in the BLA. In view of the previously described metaplasticity of rapid corticosterone effects on mEPSC frequency in the BLA (Karst et al., 2010), we wondered whether this metaplasticity is also seen at the level of synaptic plasticity in BLA circuits. Hereto, 100 nM corticosterone was administered on brain slices of mice that had been subjected to restraint stress prior to decapitation (Fig. 3C). Withingroup comparisons did not indicate a statistically significant effect of corticosterone infusion on baseline transmission (mean baseline fEPSP ± SEM before vs during perfusion: 100.7 ± 0.6% vs 102.7 ± 2.2%, p > 0.05). Interestingly, compared to slices from stressed mice not exposed to corticosterone in vitro, corticosterone strongly reduced the level of potentiation, especially towards to

Fig. 1. Schematic overview of the experimental method and outcome. (A) Positioning of the stimulation (SLA) and the recording electrode (RBLA) at their sites in the lateral amygdala (LA) and basolateral amygdala (BLA), respectively in mouse coronal brain slices. (B) Representative local fEPSP evoked by stimulation of the LA– BLA pathway. The amplitude of the signal was calculated according to the formula (a + b)/2 as illustrated in the figure. * indicates position of the stimulus artifact.

3. Results 3.1. Effect of restraint stress on synaptic plasticity in the BLA vs CA1 We have previously demonstrated that acute stress effectively enhances synaptic plasticity in the BLA (Sarabdjitsingh et al., 2012). In short, no significant difference in baseline transmission parameters were found between brain slices from control and stressed mice (using 20 min restraint stress or control treatment in vivo; LTP recording in subsequently prepared brain slices, causing a delay of >1 h between stress and recording). High frequency stimulation (HFS) evoked substantial synaptic potentiation, which was significantly more pronounced in slices from mice exposed to stress compared to control mice (Fig. 2A and B; p < 0.001). These data suggest that acute stress itself does not affect synaptic responses in the BLA but may specifically target activity-dependent plasticity in the LA-BLA pathway. To illustrate the opposing effects stress can have on different brain regions, we also studied the effects of stress on LTP in CA1. Similar to the BLA, brief exposure to acute stress did not affect baseline transmission in subsequently prepared slices compared to control (mean slope ± SEM: control 632 ± 57 mV/s vs 651 ± 60 mV/s; p > 0.05). Different from the BLA and control CA1 slices, HFS-induced synaptic plasticity was strongly attenuated in CA1 slices from stressed mice (Fig. 2C and D; p < 0.001).

Please cite this article in press as: Sarabdjitsingh, R. A., & Joëls, M. Rapid corticosteroid actions on synaptic plasticity in the mouse basolateral amygdala: Relevance of recent stress history and b-adrenergic signaling. Neurobiology of Learning and Memory (2013), http://dx.doi.org/10.1016/j.nlm.2013.10.011

4

R.A. Sarabdjitsingh, M. Joëls / Neurobiology of Learning and Memory xxx (2013) xxx–xxx

end of the 60 min period (Fig. 3C and D; p < 0.001). The average level of attenuation over 60 min was similar to control slices treated with corticosterone (F(3,36) = 21.50; p < 0.001; 100 nM corticosterone 125.8 ± 3.4% vs stress + 100 nM corticosterone 129.1 ± 5.2; p > 0.05). Taken together, these data suggest that, without altering baseline properties, exogenous corticosterone administration very rapidly and potently suppresses HFS-induced synaptic potentiation in the BLA. Additionally, the persistence of LTP after corticosterone administration seems to depend on the recent stress history of the mouse. 3.3. The effect of corticosterone on propranolol-mediated actions on BLA synaptic plasticity We have previously described that the delayed effects of corticosterone or stress on enhancement of BLA LTP depend on the badrenoceptor (Pu et al., 2009; Sarabdjitsingh et al., 2012). It is however not known whether the rapid suppressive effects of corticosterone also depend on the b-adrenoceptor. In the third experiment we studied the possible interactive effects of stress, noradrenergic transmission and corticosterone on LTP. First, we investigated the effect of 100 nM corticosterone application in brain slices of non-stressed mice pre-treated with propranolol (Fig. 4A and B). Within-group comparison to the

pre-corticosterone baseline (10 min) showed that corticosterone infusion on top of propranolol pre-treatment did not alter baseline properties in undisturbed controls (mean baseline fEPSP ± SEM before vs during perfusion: 98.9 ± 0.5% vs 101.0 ± 0.5%, p > 0.05). Tetanization resulted in significant potentiation with an average synaptic response of 128.2 ± 2.1% (average fEPSP amplitude ± SEM over the 60 min period, p < 0.001 vs pre-tetanus baseline), comparable to the group without propranolol. Compared to slices that only received corticosterone, pre-treatment with propranolol, did not result in additional suppression of synaptic plasticity (Fig. 4B; F(3,28) = 5.48, p < 0.01; 100 nM corticosterone vs propranolol + 100 nM corticosterone, p > 0.05). Next we studied whether the suppressive effect of 100 nM corticosterone after stress depends on the b-adrenoceptor. For this, we studied the effect of 100 nM corticosterone in sections of mice that were pre-treated with propranolol before stress exposure. Baseline fEPSPs amplitude was not affected by corticosterone infusion (before vs during: 98.6 ± 0.6% vs 101.4 ± 1.8%, p > 0.05). Yet, HFS resulted in only a transient form of LTP. While tetanization initially seemed to evoke potent synaptic plasticity, the signals rapidly declined to pre-tetanus levels (Fig. 4C), and were significantly lower compared to signals in stressed controls (Fig. 4D; F(3,28) = 33.43, p < 0.001; stress vs propranolol + stress + 100 nM corticosterone, p < 0.001), suggesting rapid interactive effects between stress, propranolol and corticosterone.

Fig. 2. The effect of stress on LTP in the BLA and hippocampus. (A) High frequency stimulation (HFS) of the LA afferents resulted in sustained and stable LTP at BLA synapses in slices of control mice (open diamonds, n = 15), which was more pronounced in slices of stressed mice (black diamonds, n = 14). (B) Bar chart illustrating the averages (+SEM) per treatment group for the entire 60 min post-tetanic period, showing significantly increased LTP in the BLA after stress. (C) By contrast, in the CA1 area, significant reduction of LTP is seen in slices from stressed (black circles, n = 8) compared to control animals (open circles, n = 8). (D) Bart chart illustrating the averages per treatment group for the entire 60 min post-tetanic period. Averaged mean fEPSP amplitude normalized against baseline with error bars indicating SEM. Dashed lines indicate pre-tetanus baseline levels. Two-tailed unpaired Student’s t-test, *** p < 0.001. Figure partly based on results from Sarabdjitsingh et al., 2012.

Please cite this article in press as: Sarabdjitsingh, R. A., & Joëls, M. Rapid corticosteroid actions on synaptic plasticity in the mouse basolateral amygdala: Relevance of recent stress history and b-adrenergic signaling. Neurobiology of Learning and Memory (2013), http://dx.doi.org/10.1016/j.nlm.2013.10.011

R.A. Sarabdjitsingh, M. Joëls / Neurobiology of Learning and Memory xxx (2013) xxx–xxx

5

Fig. 3. The effect of stress and corticosterone on LTP. (A) 100 nM corticosterone perfusion onto slices from control mice (black circles, n = 5) attenuates BLA LTP compared to control treatment (open diamonds, n = 15). (B) This is also illustrated by the averages per treatment group for the entire 60 min post-tetanic period. (C) Corticosterone perfusion onto slices from stressed mice (black circles, n = 6) also attenuates HFS-induced LTP compared to control slices from stressed mice. (D) Bar chart illustrating the averages per treatment group for the entire 60 min post-tetanic period. Averaged mean fEPSP amplitude normalized against baseline with error bars indicating SEM. Dashed lines indicate pre-tetanus baseline levels. Two-tailed unpaired Student’s t-test, * p < 0.05; *** p < 0.001.

3.4. Persistence of synaptic potentiation To more accurately examine the changes in LTP over time and overall persistence of the post-tetanization curve, we calculated the ratio between the late (50–60 min) and early (0–10 min) phase of the LTP curve of each experimental group (Fig. 5). The results were statistically analyzed using a 2  2  2 design with stress, propranolol treatment and 100 nM corticosterone perfusion as between-group factors. Between-group comparisons indicated a main effect of stress (F(7,1) = 20.30; p < 0.001), propranolol treatment (F(7,1) = 8.05, p < 0.01) and 100 nM corticosterone perfusion (F(7,1) = 16.83, p < 0.001). Additionally, we found an interaction between all three factors (F(7,1) = 3.93, p = 0.05), suggesting that stress, propranolol and corticosterone interact to affect the persistence of synaptic potentiation after tetanization. To simplify the interpretation of the results and considering the overall effect of stress on LTP (F(7,1) = 20.30; p < 0.001), we grouped the data according to their stress background and performed within-group analyses. One-way ANOVA showed a comparable ratio of persistence between all animal groups that were not exposed to stress (F(3,28) = 0.89; p > 0.05; Fig. 5A), suggesting that neither corticosterone nor propranolol significantly affects the maintenance of LTP in the absence of stress. However, this changed significantly when animals were exposed to stress (F(3,27) = 22.18; p < 0.001; Fig. 5B). Post-hoc analysis showed that relative to the stress group, the maintenance of LTP

diminished when animals received either propranolol, in vitro corticosterone or a combination of these treatments. Interestingly, the combination of propranolol, stress and in vitro application of 100 nM corticosterone resulted in the most transient form of LTP (vs. propranolol + stress or vs propranolol + 100 nM corticosterone, p < 0.05). These data suggest that both the level of potentiation and particularly the long-term maintenance during the post-tetanic period are affected by additive interactions among stress, propranolol and corticosterone.

4. Discussion In the current study we demonstrate that corticosterone can rapidly modulate and supress synaptic plasticity in the BLA. This suppression and loss of persistence particularly occurs in animals that have recently experienced restraint stress. Additionally, we probed the role of b-adrenergic interactions with corticosterone. We found that blocking b-adrenergic transmission with propranolol before stress exposure also results in transient LTP, an effect that is even more pronounced when tissue is subsequently exposed to corticosterone. In line with previous work from our lab (Pu et al., 2009), we first showed that acute administration of corticosterone can rapidly decrease synaptic potentiation in the BLA. We refer to these effects as ‘rapid’ since they occur in the same time-domain as effects earlier

Please cite this article in press as: Sarabdjitsingh, R. A., & Joëls, M. Rapid corticosteroid actions on synaptic plasticity in the mouse basolateral amygdala: Relevance of recent stress history and b-adrenergic signaling. Neurobiology of Learning and Memory (2013), http://dx.doi.org/10.1016/j.nlm.2013.10.011

6

R.A. Sarabdjitsingh, M. Joëls / Neurobiology of Learning and Memory xxx (2013) xxx–xxx

Fig. 4. The effect of stress and corticosterone on LTP in propranolol-pretreated mice. (A) 100 nM corticosterone administration resulted in attenuated and transient BLA-LTP in control (grey squares, n = 6) and (C) stressed mice (grey squares, n = 6) injected with propranolol before stress exposure. (B) and (D) Bar chart illustrating the averages per treatment group for the 60 min post-tetanus period. Averaged mean fEPSP amplitude normalized against baseline with error bars indicating SEM. Dashed lines indicate pretetanus baseline levels. Tukey’s post hoc test, * p < 0.05, *** p < 0.001.

described in patch clamp studies, which were shown to occur independent from protein synthesis (Karst et al., 2010). The rapid decrease in LTP is opposite to the delayed actions of stress (and thus presumably also corticosterone) in the BLA, because we and others have previously shown a slow enhancement in BLA excitability and synaptic plasticity in vivo and in brain slices hours after exposure to stress (Kavushansky & Richter-Levin, 2006; Maroun, 2006; Rodriguez Manzanares et al., 2005; Sarabdjitsingh et al., 2012; Vouimba et al., 2006). These bidirectional effects of corticosterone on LTP are reminiscent of the difference in rapid and delayed effects of corticosterone observed in the hippocampus (Joels & Krugers, 2007; Wiegert et al., 2006), though the direction of the BLA effects (rapid as well as delayed) is exactly opposite to what was found in the hippocampus. The rapid suppression of BLA LTP by corticosterone is a somewhat unexpected finding because rapid effects of corticosterone in the BLA and hippocampus at the single cell level are highly comparable, i.e. in both areas the hormone enhances mEPSC frequency (Joels et al., 2012). Apparently that is not translated to synaptic plasticity at the circuit level. Possible explanations may be found in mechanisms downstream of glutamatergic transmission and important for synaptic plasticity; these may differ between the hippocampus and BLA. There is some evidence that LTP in the BLA involves voltage-dependent calcium signaling, more so than NMDA-receptor pathways (Bauer, Schafe, & LeDoux, 2002). LTP involving the former, as opposed to the

latter, was earlier shown to be facilitated by corticosterone in the hippocampus (Krugers et al., 2005). Also, the late phase of LTP depends on protein kinase A and mitogen-activated protein kinase (Huang, Martin, & Kandel, 2000; Wu, Rowan, & Anwyl, 2006). These pathways may be differently affected by corticosterone in the hippocampus vs BLA, but this is speculative at this stage. We were also interested to see if metaplasticity in response to corticosterone, as observed at the single cell level, can be discerned with regard to LTP at the circuit level. Thus, within the BLA, several studies have provided evidence for metaplastic effects of corticosterone on BLA neuronal excitability and function: the nature of these corticosteroid actions depends on the recent stress history of an animal (Karst et al., 2010; Rao, Anilkumar, McEwen, & Chattarji, 2012). It is important to note that we did not see any changes in baseline fEPSP amplitude after stress, which contrasts with earlier studies (e.g. Karst et al., 2010; Kavushansky & Richter-Levin, 2006; Vouimba, Yaniv, Diamond, & Richter-Levin, 2004). It should be noted however that these other studies were performed either in vivo or with patch clamp recordings. These methodological differences could potentially contribute to the discrepancy. For instance, patch clamp recording of single neurons was performed in the presence of bicuccilline (to block GABAergic transmission) as opposed to the current design, so that putative effects of stress on GABAergic transmission (which contribute to the overall outcome) may have remained unnoticed. In vivo stimulation may

Please cite this article in press as: Sarabdjitsingh, R. A., & Joëls, M. Rapid corticosteroid actions on synaptic plasticity in the mouse basolateral amygdala: Relevance of recent stress history and b-adrenergic signaling. Neurobiology of Learning and Memory (2013), http://dx.doi.org/10.1016/j.nlm.2013.10.011

R.A. Sarabdjitsingh, M. Joëls / Neurobiology of Learning and Memory xxx (2013) xxx–xxx

Fig. 5. Ratio indicating persistence of BLA-LTP. The overall persistence in BLA-LTP was calculated as ratio between the late (50–60 min) and early (0–10 min) phase after tetanization. (A) The different treatment groups did not differ in ratio under non-stressed conditions. (B) Under stress conditions, the persistence in LTP was decreased in animals treated with 100 nM corticosterone, propranolol or both compared to stress control group. Averaged mean ratio with error bars indicating SEM. One-way ANOVA and Tukey’s post hoc test, *** p < 0.001; LSD post hoc test, # p < 0.05.

have activated other pathways than the ones activated in the current (reduced) slice preparations. To some extent we did observe different responses to corticosterone regarding LTP when the organism had earlier been exposed to restraint stress. In particular the stability of the LTP evoked in the presence of corticosterone turned out to be sensitive to earlier stress exposure. While a single pulse of corticosterone (i.e. in tissue from non-stressed mice) rapidly attenuated LTP but still yielded considerably elevated synaptic transmission 60 min later, recent exposure to stress gradually led to a much stronger suppression of LTP by corticosterone. Interestingly, this corticosterone-induced instability in maintenance of synaptic plasticity is highly reminiscent of what is seen when b-adrenoceptors are blocked during stress by propranolol administration (Sarabdjitsingh et al., 2012). Possibly the two hormones share elements in their downstream effector pathways, such as protein kinase A. The fact, though, that propranolol and corticosterone appear to have an additive effect on the maintenance of LTP, specifically in stressed mice, seems to argue against this assumption. Our current set of experiments does not allow further conclusions about the mechanisms underlying corticosteroid metaplasticity. For instance, it is unknown which corticosteroid receptor is involved in the transient character of LTP upon a second exposure to corticosterone or whether the response to corticosterone involves gene transcription causing a decline in signals. Regardless of the mechanism, however, the present results do indicate that BLA cells and circuits respond differently to corticosterone when the organism has recently experienced a stressor, compared to undisturbed control conditions. At this moment we can only speculate about the significance of these findings. Extensive evidence in humans and rodents suggests

7

that the amygdala, especially its basolateral part, has an orchestrating role in the circuit controlling emotional memory formation (de Quervain, Aerni, Schelling, & Roozendaal, 2009; McGaugh, 2004; Roozendaal et al., 2009). Behavioral as well as electrophysiological studies have shown that optimal memory formation and behavioral performance can only be established through interactive effects of corticosteroids and noradrenaline. The impact of such interactive actions is not restricted to the BLA only, but also involves brain regions to which the BLA is connected. For instance, activation of the BLA has modulatory effects on LTP in other brain regions such as the hippocampus (Akirav & Richter-Levin, 1999; Frey, Bergado-Rosado, Seidenbecher, Pape, & Frey, 2001; Nakao, Matsuyama, Matsuki, & Ikegaya, 2004), and prefrontal cortex (Richter-Levin & Maroun, 2010; Tan, Lauzon, Bishop, Bechard, & Laviolette, 2010). Very few studies so far have examined the consequences of renewed exposure to corticosterone some hours after stress. We here show that such a situation affects LTP in the BLA differently than when corticosterone is given against an undisturbed background, i.e. by reducing the BLA ability to express sustained synaptic plasticity. Our results suggest a protective role for renewed corticosteroid exposure, by resetting BLA excitability and synaptic plasticity and hence preventing the stress response to cause inappropriate or excessive memory consolidation. In that respect, insufficient levels of corticosterone—as have been hypothesized to occur in individuals vulnerable to posttraumatic stress disorder (Yehuda, 2009)—might fail to reset the BLA excitability and synaptic plasticity. It should be noted, though, that in real life re-exposure of BLA circuits to corticosterone will not occur without also involving noradrenaline (among other stress modulators, such as CRH or endocannabinoids). More in general, extrapolation of the current in vitro findings to the in vivo situation is preliminary as are speculations about their relevance for human psychopathology. In conclusion, we have demonstrated that corticosterone and badrenergic transmission are major players in the long-term maintenance and persistence of stress-induced increases in synaptic plasticity in the mouse BLA. These results will be important not only for our understanding of how emotional memory is formed under stress conditions but possibly could provide clues for the elucidation of the pathophysiology and etiology of stress-related disorders.

References Adamec, R., Muir, C., Grimes, M., & Pearcey, K. (2007). Involvement of noradrenergic and corticoid receptors in the consolidation of the lasting anxiogenic effects of predator stress. Behavioural Brain Research, 179, 192–207. Akirav, I., & Richter-Levin, G. (1999). Biphasic modulation of hippocampal plasticity by behavioral stress and basolateral amygdala stimulation in the rat. Journal of Neuroscience, 19, 10530–10535. Bauer, E. P., Schafe, G. E., & LeDoux, J. E. (2002). NMDA receptors and L-type voltagegated calcium channels contribute to long-term potentiation and different components of fear memory formation in the lateral amygdala. Journal of Neuroscience, 22, 5239–5249. de Kloet, E. R., Joels, M., & Holsboer, F. (2005). Stress and the brain: from adaptation to disease. Nature Reviews Neuroscience, 6, 463–475. de Quervain, D. J., Aerni, A., Schelling, G., & Roozendaal, B. (2009). Glucocorticoids and the regulation of memory in health and disease. Frontiers in Neuroendocrinology, 30, 358–370. Frey, S., Bergado-Rosado, J., Seidenbecher, T., Pape, H. C., & Frey, J. U. (2001). Reinforcement of early long-term potentiation (early-LTP) in dentate gyrus by stimulation of the basolateral amygdala: Heterosynaptic induction mechanisms of late-LTP. Journal of Neuroscience, 21, 3697–3703. Gibbs, M. E., & Summers, R. J. (2002). Role of adrenoceptor subtypes in memory consolidation. Progress in Neurobiology, 67, 345–391. Groc, L., Choquet, D., & Chaouloff, F. (2008). The stress hormone corticosterone conditions AMPAR surface trafficking and synaptic potentiation. Nature Neuroscience, 11, 868–870. Herbert, J., Goodyer, I. M., Grossman, A. B., Hastings, M. H., de Kloet, E. R., Lightman, S. L., et al. (2006). Do corticosteroids damage the brain? Journal of Neuroendocrinology, 18, 393–411. Huang, Y. Y., Martin, K. C., & Kandel, E. R. (2000). Both protein kinase A and mitogenactivated protein kinase are required in the amygdala for the macromolecular

Please cite this article in press as: Sarabdjitsingh, R. A., & Joëls, M. Rapid corticosteroid actions on synaptic plasticity in the mouse basolateral amygdala: Relevance of recent stress history and b-adrenergic signaling. Neurobiology of Learning and Memory (2013), http://dx.doi.org/10.1016/j.nlm.2013.10.011

8

R.A. Sarabdjitsingh, M. Joëls / Neurobiology of Learning and Memory xxx (2013) xxx–xxx

synthesis-dependent late phase of long-term potentiation. Journal of Neuroscience, 20, 6317–6325. Huge, V., Rammes, G., Beyer, A., Zieglgansberger, W., & Azad, S. C. (2009). Activation of kappa opioid receptors decreases synaptic transmission and inhibits longterm potentiation in the basolateral amygdala of the mouse. European Journal of Pain, 13, 124–129. Joels, M., & Baram, T. Z. (2009). The neuro-symphony of stress. Nature Reviews Neuroscience, 10, 459–466. Joels, M., & Krugers, H. J. (2007). LTP after stress: Up or down? Neural Plasticity, 2007, 93202. Joels, M., Sarabdjitsingh, R. A., & Karst, H. (2012). Unraveling the time domains of corticosteroid hormone influences on brain activity: Rapid, slow, and chronic modes. Pharmacological Reviews, 64, 901–938. Karst, H., Berger, S., Erdmann, G., Schutz, G., & Joels, M. (2010). Metaplasticity of amygdalar responses to the stress hormone corticosterone. Proceeding of the National Academic Science USA, 107, 14449–14454. Karst, H., Berger, S., Turiault, M., Tronche, F., Schutz, G., & Joels, M. (2005). Mineralocorticoid receptors are indispensable for nongenomic modulation of hippocampal glutamate transmission by corticosterone. Proceedings of the National Academic Science USA, 102, 19204–19207. Kavushansky, A., & Richter-Levin, G. (2006). Effects of stress and corticosterone on activity and plasticity in the amygdala. Journal of Neuroscience Research, 84, 1580–1587. Krugers, H. J., Alfarez, D. N., Karst, H., Parashkouhi, K., van, G. N., & Joels, M. (2005). Corticosterone shifts different forms of synaptic potentiation in opposite directions. Hippocampus, 15, 697–703. Krugers, H. J., Karst, H., & Joels, M. (2012). Interactions between noradrenaline and corticosteroids in the brain: From electrical activity to cognitive performance. Front Cell Neuroscience, 6, 15. Maroun, M. (2006). Stress reverses plasticity in the pathway projecting from the ventromedial prefrontal cortex to the basolateral amygdala. European Journal of Neuroscience, 24, 2917–2922. Martin, S., Henley, J. M., Holman, D., Zhou, M., Wiegert, O., van, S. M., et al. (2009). Corticosterone alters AMPAR mobility and facilitates bidirectional synaptic plasticity. PLoS One, 4, e4714. McEwen, B. S. (2007). Physiology and neurobiology of stress and adaptation: Central role of the brain. Physiological Reviews, 87, 873–904. McGaugh, J. L. (2004). The amygdala modulates the consolidation of memories of emotionally arousing experiences. Annual Review of Neuroscience, 27, 1–28. McGaugh, J. L. (2013). Making lasting memories: Remembering the significant. Proceeding of the National Academic Sciences USA, 110(Suppl 2), 10402–10407. Nakao, K., Matsuyama, K., Matsuki, N., & Ikegaya, Y. (2004). Amygdala stimulation modulates hippocampal synaptic plasticity. Proceedings of the National Academic Science USA, 101, 14270–14275. Pitkanen, A., Stefanacci, L., Farb, C. R., Go, G. G., LeDoux, J. E., & Amaral, D. G. (1995). Intrinsic connections of the rat amygdaloid complex: Projections originating in the lateral nucleus. Journal of Comparative Neurology, 356, 288–310. Popoli, M., Yan, Z., McEwen, B. S., & Sanacora, G. (2012). The stressed synapse: The impact of stress and glucocorticoids on glutamate transmission. Nature Reviews Neuroscience, 13, 22–37.

Pu, Z., Krugers, H. J., & Joels, M. (2009). Beta-adrenergic facilitation of synaptic plasticity in the rat basolateral amygdala in vitro is gradually reversed by corticosterone. Learning and Memory, 16, 155–160. Rammes, G., Steckler, T., Kresse, A., Schutz, G., Zieglgansberger, W., & Lutz, B. (2000). Synaptic plasticity in the basolateral amygdala in transgenic mice expressing dominant-negative cAMP response element-binding protein (CREB) in forebrain. European Journal of Neuroscience, 12, 2534–2546. Rao, R. P., Anilkumar, S., McEwen, B. S., & Chattarji, S. (2012). Glucocorticoids protect against the delayed behavioral and cellular effects of acute stress on the amygdala. Biological Psychiatry, 72, 466–475. Reul, J. M., & de Kloet, E. R. (1985). Two receptor systems for corticosterone in rat brain: Microdistribution and differential occupation. Endocrinology, 117, 2505–2511. Richter-Levin, G., & Maroun, M. (2010). Stress and amygdala suppression of metaplasticity in the medial prefrontal cortex. Cerebral Cortex, 20, 2433–2441. Rodriguez Manzanares, P. A., Isoardi, N. A., Carrer, H. F., & Molina, V. A. (2005). Previous stress facilitates fear memory, attenuates GABAergic inhibition, and increases synaptic plasticity in the rat basolateral amygdala. Journal of Neuroscience, 25, 8725–8734. Roozendaal, B., McEwen, B. S., & Chattarji, S. (2009). Stress, memory and the amygdala. Nature Reviews Neuroscience, 10, 423–433. Sapolsky, R. M., McEwen, B. S., & Rainbow, T. C. (1983). Quantitative autoradiography of [3H]corticosterone receptors in rat brain. Brain Research, 271, 331–334. Sapolsky, R. M., Romero, L. M., & Munck, A. U. (2000). How do glucocorticoids influence stress responses? Integrating permissive, suppressive, stimulatory, and preparative actions. Endocrine Reviews, 21, 55–89. Sarabdjitsingh, R. A., Kofink, D., Karst, H., de Kloet, E. R., & Joels, M. (2012). Stressinduced enhancement of mouse amygdalar synaptic plasticity depends on glucocorticoid and ss-adrenergic activity. PLoS One, 7, e42143. Tan, H., Lauzon, N. M., Bishop, S. F., Bechard, M. A., & Laviolette, S. R. (2010). Integrated cannabinoid CB1 receptor transmission within the amygdala– prefrontal cortical pathway modulates neuronal plasticity and emotional memory encoding. Cerebral Cortex, 20, 1486–1496. Ulrich-Lai, Y. M., & Herman, J. P. (2009). Neural regulation of endocrine and autonomic stress responses. Nature Reviews Neuroscience, 10, 397–409. Vouimba, R. M., Munoz, C., & Diamond, D. M. (2006). Differential effects of predator stress and the antidepressant tianeptine on physiological plasticity in the hippocampus and basolateral amygdala. Stress, 9, 29–40. Vouimba, R. M., Yaniv, D., Diamond, D., & Richter-Levin, G. (2004). Effects of inescapable stress on LTP in the amygdala versus the dentate gyrus of freely behaving rats. European Journal of Neuroscience, 19, 1887–1894. Yehuda, R. (2009). Status of glucocorticoid alterations in post-traumatic stress disorder. Ann N Y Acad Sci., 1179, 56–69. Wiegert, O., Joels, M., & Krugers, H. (2006). Timing is essential for rapid effects of corticosterone on synaptic potentiation in the mouse hippocampus. Learning and Memory, 13, 110–113. Wu, J., Rowan, M. J., & Anwyl, R. (2006). Long-term potentiation is mediated by multiple kinase cascades involving CaMKII or either PKA or p42/44 MAPK in the adult rat dentate gyrus in vitro. Journal of Neurophysiology, 95, 3519–3527.

Please cite this article in press as: Sarabdjitsingh, R. A., & Joëls, M. Rapid corticosteroid actions on synaptic plasticity in the mouse basolateral amygdala: Relevance of recent stress history and b-adrenergic signaling. Neurobiology of Learning and Memory (2013), http://dx.doi.org/10.1016/j.nlm.2013.10.011

Rapid corticosteroid actions on synaptic plasticity in the mouse basolateral amygdala: relevance of recent stress history and β-adrenergic signaling.

The rodent stress hormone corticosterone rapidly enhances long-term potentiation in the CA1 hippocampal area, but leads to a suppression when acting i...
1MB Sizes 0 Downloads 0 Views