Basal forebrain cholinergic systems as circuits through which traumatic stress disrupts emotional memory regulation.

Contextual and spatial systems facilitate changes in emotional memory regulation brought on by traumatic stress. Cholinergic basal forebrain (chBF) neurons provide input to contextual/spatial systems and although chBF neurons are important for emotional memory, it is unknown how they contribute to the traumatic stress effects on emotional memory. Clusters of chBF neurons that project to the prefrontal cortex (PFC) modulate fear conditioned suppression and passive avoidance, while clusters of chBF neurons that project to the hippocampus (Hipp) and PFC (i.e. cholinergic medial septum and diagonal bands of Broca (chMS/DBB neurons) are critical for fear extinction. Interestingly, neither Hipp nor PFC projecting chMS/DBB neurons are critical for fear extinction. The retrosplenial cortex (RSC) is a contextual/spatial memory system that receives input from chMS/DBB neurons, but whether this chMS/DBB-RSC circuit facilitates traumatic stress effects on emotional memory remain unexplored. Traumatic stress leads to neuroinflammation and the buildup of reactive oxygen species. These two molecular processes may converge to disrupt chBF circuits enhancing the impact of traumatic stress on emotional memory.

The role of avoidance in modulating single prolonged stress effects on emotional memory in male and female rats.

The incidence of post traumatic stress disorder (PTSD) is greater in women than men, but mechanisms via which this difference manifests remain under explored. The single prolonged stress (SPS) rodent model of traumatic stress has been used to identify mechanisms through which traumatic stress leads to deficits in retaining extinction (a core PTSD symptom), but has been mostly utilized in male model systems. Recent studies have observed that SPS leads to changes in persistent fear memory in female rats, though these results are variable. This variability could be driven by changes in behavioral strategy in females during extinction, but this possibility has not been sufficiently explored. To address this, we examined the impact of SPS on freezing and avoidance (a core PTSD symptom) during extinction in male and female rats. In male rats, SPS enhanced acquisition of conditioned freezing, but did not enhance freezing during extinction training or testing. SPS also decreased avoidance during extinction training, but not extinction testing. In female rats, SPS had no impact on conditioned freezing. Avoidance was not observed in control rats, but emerged in SPS/female rats during extinction testing. Furthermore, avoidance was negatively correlated with freezing in female rats (high avoidance associated with lower freezing), but this relationship was disrupted with SPS. The results suggest that introducing avoidance during extinction negates SPS effects on extinction retention in male and female rats, control/female rats engage in avoidance to regulate fear expression, and this relationship is disrupted with SPS.

The role of estrogen receptor manipulation during traumatic stress on changes in emotional memory induced by traumatic stress.

RATIONALE: Traumatic stress leads to persistent fear, which is a core feature of post-traumatic stress disorder (PTSD). Women are more likely than men to develop PTSD after trauma exposure, which suggests women are differentially sensitive to traumatic stress. However, it is unclear how this differential sensitivity manifests. Cyclical changes in vascular estrogen release could be a contributing factor where levels of vascular estrogens (and activation of estrogen receptors) at the time of traumatic stress alter the impact of traumatic stress. METHODS: To examine this, we manipulated estrogen receptors at the time of stress and observed the effect this had on fear and extinction memory (within the single prolonged stress (SPS) paradigm) in female rats. In all experiments, freezing and darting were used to measure fear and extinction memory. RESULTS: In Experiment 1, SPS enhanced freezing during extinction testing, and this effect was blocked by nuclear estrogen receptor antagonism prior to SPS. In Experiment 2, SPS decreased conditioned freezing during the acquisition and testing of extinction. Administration of 17ß-estradiol altered freezing in control and SPS animals during the acquisition of extinction, but this treatment had no effect on freezing during the testing of extinction memory. In all experiments, darting was only observed to footshock onset during fear conditioning. CONCLUSION: The results suggest multiple behaviors (or different behavioral paradigms) are needed to characterize the nature of traumatic stress effects on emotional memory in female rats and that nuclear estrogen receptor antagonism prior to SPS blocks SPS effects on emotional memory in female rats.

Fear learning-induced changes in AMPAR and NMDAR expression in the fear circuit.

NMDA receptors (NMDARs) and AMPA receptors (AMPARs) in amygdala nuclei and the dorsal hippocampus (dHipp) are critical for fear conditioning. Enhancements in synaptic AMPAR expression in amygdala nuclei and the dHipp are critical for fear conditioning, with some studies observing changes in AMPAR expression across many neurons in these brain regions. Whether similar changes occur in other nodes of the fear circuit (e.g., ventral hippocampus [vHipp]) or changes in NMDAR expression in the fear circuit occur with fear conditioning have not been sufficiently examined. To address this we used near-infrared immunohistochemistry (IHC) to measure AMPAR and NMDAR subunit expression in several nodes of the fear circuit. Long-term changes in GluR1 and GluR2 expression in the ventral hippocampus (vHipp) and anterior cingulate cortex (ACC), enhanced NR2A expression in amygdala nuclei, and changes in the ratio of GluR1/NR2A and GluR2/NR2A in the dHipp was observed with fear conditioning. Most of these changes were dependent on protein synthesis during fear conditioning and were not observed immediately after fear conditioning. The results of the study suggest that global changes in AMPARs and NMDARs occur in multiple nodes within the fear circuit and raise the possibility that these changes contribute to fear memory. Further research examining how global changes in AMPAR, NMDAR, and AMPAR/NMDAR ratios within nodes of the fear circuit contribute to fear memory is needed.

Maternal Separation Induces Sex-Specific Differences in Sensitivity to Traumatic Stress.

Post-traumatic stress disorder (PTSD) is a debilitating psychiatric disorder with a high economic burden. Two risk factors for increasing the chances of developing PTSD are sex (being female) and early life stress. These risk factors suggest that early life stress-induced changes and sex differences in emotional circuits and neuroendocrinological systems lead to susceptibility to traumatic stress. Exploring mechanisms via which stress leads to specific effects can be accomplished in animal models, but reliable animal models that allow for an examination of how early life stress interacts with sex to increase susceptibility to traumatic stress is lacking. To address this, we examined the effects of early life stress [using the maternal separation (MS) model] and late adolescence/early adult traumatic stress [using the single prolonged stress (SPS) model] on startle reactivity, anxiety-like behavior in the open field (OF), and basal corticosterone levels in male and female rats. Female rats exposed to MS and SPS (MS/SPS) showed enhanced startle reactivity relative to MS/control female rats. Enhanced startle reactivity was not observed in MS/SPS male rats. Instead, non-maternally separated male rats that were exposed to SPS showed enhanced startle reactivity relative to controls. Female rats had enhanced locomotor activity in the OF and higher basal corticosterone levels in comparison to males, but measures in the OF and basal corticosterone were not affected by MS or SPS. Overall the results suggest that the combined MS and SPS models can be used to explore how changes in maternal care during infancy lead to sex differences in sensitivity to the effects of traumatic stress as adolescents and adults.

Systematic Review and Methodological Considerations for the Use of Single Prolonged Stress and Fear Extinction Retention in Rodents.

Posttraumatic stress disorder (PTSD) is a mental health condition triggered by experiencing or witnessing a terrifying event that can lead to lifelong burden that increases mortality and adverse health outcomes. Yet, no new treatments have reached the market in two decades. Thus, screening potential interventions for PTSD is of high priority. Animal models often serve as a critical translational tool to bring new therapeutics from bench to bedside. However, the lack of concordance of some human clinical trial outcomes with preclinical animal efficacy findings has led to a questioning of the methods of how animal studies are conducted and translational validity established. Thus, we conducted a systematic review to determine methodological variability in studies that applied a prominent animal model of trauma-like stress, single prolonged stress (SPS). The SPS model has been utilized to evaluate a myriad of PTSD-relevant outcomes including extinction retention. Rodents exposed to SPS express an extinction retention deficit, a phenotype identified in humans with PTSD, in which fear memory is aberrantly retained after fear memory extinction. The current systematic review examines methodological variation across all phases of the SPS paradigm, as well as strategies for behavioral coding, data processing, statistical approach, and the depiction of data. Solutions for key challenges and sources of variation within these domains are discussed. In response to methodological variation in SPS studies, an expert panel was convened to generate methodological considerations to guide researchers in the application of SPS and the evaluation of extinction retention as a test for a PTSD-like phenotype. Many of these guidelines are applicable to all rodent paradigms developed to model trauma effects or learned fear processes relevant to PTSD, and not limited to SPS. Efforts toward optimizing preclinical model application are essential for enhancing the reproducibility and translational validity of preclinical findings, and should be conducted for all preclinical psychiatric research models.

PI3K-Akt Signaling in the Basolateral Amygdala Facilitates Traumatic Stress Enhancements in Fear Memory.

BACKGROUND: A core symptom of posttraumatic stress disorder is persistent fear memory, which can be defined as fear memory that is resistant to updating, inhibition, or extinction. posttraumatic stress disorder emerges after traumatic stress exposure, but neurobiological mechanisms via which traumatic stress leads to persistent fear memory are not well defined. Akt signaling within the amygdala (Amy) is enhanced with traumatic stress, and phosphatidylinositol kinase 3 (PI3K) activation of Akt within the basolateral Amy (BLA) has been implicated as critical to fear memory formation. These findings raise the possibility that traumatic stress enhances PI3K→Akt signaling in the BLA, which leads to persistent fear memory. METHODS: To test this hypothesis, rats were exposed to traumatic stress using the single prolonged stress model, and changes in Akt phosphorylation were assayed in the Amy at 0 and 30 minutes after fear conditioning (FC). In a separate experiment, we inhibited PI3K→Akt signaling in the BLA prior to FC and observed the effect this had on acquisition, expression, and extinction of FC in stressed and control rats. RESULTS: Enhanced Akt phosphorylation in the Amy at both time points was observed in stressed rats, but not in control rats. PI3K→Akt inhibition in the BLA had no effect on freezing in control rats but decreased freezing during extinction training and testing in stressed rats. CONCLUSION: These findings suggest that PI3K→Akt signaling in the BLA could be a mechanism via which traumatic stress leads to fear memory that is resistant to extinction.

Single prolonged stress alters neural activation in the periacqueductal gray and midline thalamic nuclei during emotional learning and memory.

Clinical and preclinical studies that have examined the neurobiology of persistent fear memory in posttraumatic stress disorder (PTSD) have focused on the medial prefrontal cortex, hippocampus, and amygdala. Sensory systems, the periaqueductal gray (PAG), and midline thalamic nuclei have been implicated in fear and extinction memory, but whether neural activity in these substrates is sensitive to traumatic stress (at baseline or during emotional learning and memory) remains unexplored. To address this, we used the single prolonged stress (SPS) model of traumatic stress. SPS and control rats were either subjected to fear conditioning (CS-fear) or presented with CSs alone (CS-only) during fear conditioning. All rats were then subjected to extinction training and testing. A subset of rats were euthanized after each behavioral stage and c-Fos and c-Jun used to measure neural activation in all substrates. SPS lowered c-Jun levels in the dorsomedial and lateral PAG at baseline, but the elevated c-Jun expression in the PAG during emotional learning and memory. SPS also altered c-Fos expression during fear and extinction learning/memory in midline thalamic nuclei. These findings suggest changes in neural function in the PAG and midline thalamic nuclei could contribute to persistent fear memory induced by traumatic stress. Interestingly, SPS effects were also observed in animals that never learned fear or extinction (i.e., CS-only). This raises the possibility that traumatic stress could have broader effects on the psychological function that are dependent on the PAG and midline thalamic nuclei.

Using Near-infrared Fluorescence and High-resolution Scanning to Measure Protein Expression in the Rodent Brain.

Neuroscience is the study of how cells in the brain mediate various functions. Measuring protein expression in neurons and glia is critical for the study of neuroscience as cellular function is determined by the composition and activity of cellular proteins. In this article, we describe how immunocytochemistry can be combined with near-infrared high-resolution scanning to provide a semi-quantitative measure of protein expression in distinct brain regions. This technique can be used for single or double protein expression in the same brain region. Measuring proteins in this fashion can be used to obtain a relative change in protein expression with an experimental manipulation, molecular signature of learning and memory, activity in molecular pathways, and neural activity in multiple brain regions. Using the correct proteins and statistical analysis, functional connectivity among brain regions can be determined as well. Given the ease of implementing immunocytochemistry in a laboratory, using immunocytochemistry with near-infrared high-resolution scanning can expand the ability of the neuroscientist to examine neurobiological processes at a systems level.

Characterizing changes in glucocorticoid receptor internalization in the fear circuit in an animal model of post traumatic stress disorder.

Glucocorticoid receptors (GRs) shuttle from the cytoplasm (cy) to the nucleus (nu) when bound with glucocorticoids (i.e. GR internalization) and alter transcriptional activity. GR activation within the fear circuit has been implicated in fear memory and post traumatic stress disorder (PTSD). However, no study to date has characterized GR internalization within the fear circuit during fear memory formation or examined how traumatic stress impacts this process. To address this, we assayed cy and nu GR levels at baseline and after auditory fear conditioning (FC) in the single prolonged stress (SPS) model of PTSD. Cy and nu GRs within the medial prefrontal cortex (mPFC), dorsal hippocampus (dHipp), ventral hippocampus (vHipp), and amygdala (AMY) were assayed using western blot. The distribution of GR in the cy and nu (at baseline and after FC) was varied across individual nodes of the fear circuit. At baseline, SPS enhanced cyGRs in the dHipp, but decreased cyGRs in the AMY. FC only enhanced GR internalization in the AMY and this effect was attenuated by SPS exposure. SPS also decreased cyGRs in the dHipp after FC. The results of this study suggests that GR internalization is varied across the fear circuit, which in turn suggests GR activation is selectively regulated within individual nodes of the fear circuit. The findings also suggest that changes in GR dynamics in the dHipp and AMY modulate the enhancing effect SPS has on fear memory persistence.

Disruption of medial septum and diagonal bands of Broca cholinergic projections to the ventral hippocampus disrupt auditory fear memory.

In classical fear conditioning, a neutral conditioned stimulus (CS) is paired with an aversive unconditioned stimulus (US), which leads to a fear memory. If the CS is repeatedly presented without the US after fear conditioning, the formation of an extinction memory occurs, which inhibits fear memory expression. A previous study has demonstrated that selective cholinergic lesions in the medial septum and vertical limb of the diagonal bands of Broca (MS/vDBB) prior to fear and extinction learning disrupt contextual fear memory discrimination and acquisition of extinction memory. MS/vDBB cholinergic neurons project to a number of substrates that are critical for fear and extinction memory. However, it is currently unknown which of these efferent projections are critical for contextual fear memory discrimination and extinction memory. To address this, we induced cholinergic lesions in efferent targets of MS/vDBB cholinergic neurons. These included the dorsal hippocampus (dHipp), ventral hippocampus (vHipp), medial prefrontal cortex (mPFC), and in the mPFC and dHipp combined. None of these lesion groups exhibited deficits in contextual fear memory discrimination or extinction memory. However, vHipp cholinergic lesions disrupted auditory fear memory. Because MS/vDBB cholinergic neurons are the sole source of acetylcholine in the vHipp, these results suggest that MS/vDBB cholinergic input to the vHipp is critical for auditory fear memory. Taken together with previous findings, the results of this study suggest that MS/vDBB cholinergic neurons are critical for fear and extinction memory, though further research is needed to elucidate the role of MS/vDBB cholinergic neurons in these types of emotional memory.

Using c-Jun to identify fear extinction learning-specific patterns of neural activity that are affected by single prolonged stress.

Neural circuits via which stress leads to disruptions in fear extinction is often explored in animal stress models. Using the single prolonged stress (SPS) model of post traumatic stress disorder and the immediate early gene (IEG) c-Fos as a measure of neural activity, we previously identified patterns of neural activity through which SPS disrupts extinction retention. However, none of these stress effects were specific to fear or extinction learning and memory. C-Jun is another IEG that is sometimes regulated in a different manner to c-Fos and could be used to identify emotional learning/memory specific patterns of neural activity that are sensitive to SPS. Animals were either fear conditioned (CS-fear) or presented with CSs only (CS-only) then subjected to extinction training and testing. C-Jun was then assayed within neural substrates critical for extinction memory. Inhibited c-Jun levels in the hippocampus (Hipp) and enhanced functional connectivity between the ventromedial prefrontal cortex (vmPFC) and basolateral amygdala (BLA) during extinction training was disrupted by SPS in the CS-fear group only. As a result, these effects were specific to emotional learning/memory. SPS also disrupted inhibited Hipp c-Jun levels, enhanced BLA c-Jun levels, and altered functional connectivity among the vmPFC, BLA, and Hipp during extinction testing in SPS rats in the CS-fear and CS-only groups. As a result, these effects were not specific to emotional learning/memory. Our findings suggest that SPS disrupts neural activity specific to extinction memory, but may also disrupt the retention of fear extinction by mechanisms that do not involve emotional learning/memory.

Neural circuits via which single prolonged stress exposure leads to fear extinction retention deficits.

Single prolonged stress (SPS) has been used to examine mechanisms via which stress exposure leads to post-traumatic stress disorder symptoms. SPS induces fear extinction retention deficits, but neural circuits critical for mediating these deficits are unknown. To address this gap, we examined the effect of SPS on neural activity in brain regions critical for extinction retention (i.e., fear extinction circuit). These were the ventral hippocampus (vHipp), dorsal hippocampus (dHipp), basolateral amygdala (BLA), prelimbic cortex (PL), and infralimbic cortex (IL). SPS or control rats were fear conditioned then subjected to extinction training and testing. Subsets of rats were euthanized after extinction training, extinction testing, or immediate removal from the housing colony (baseline condition) to assay c-Fos levels (measure of neural activity) in respective brain region. SPS induced extinction retention deficits. During extinction training SPS disrupted enhanced IL neural activity and inhibited BLA neural activity. SPS also disrupted inhibited BLA and vHipp neural activity during extinction testing. Statistical analyses suggested that SPS disrupted functional connectivity within the dHipp during extinction training and increased functional connectivity between the BLA and vHipp during extinction testing. Our findings suggest that SPS induces extinction retention deficits by disrupting both excitatory and inhibitory changes in neural activity within the fear extinction circuit and inducing changes in functional connectivity within the Hipp and BLA.

The role of basal forebrain cholinergic neurons in fear and extinction memory.

Cholinergic input to the neocortex, dorsal hippocampus (dHipp), and basolateral amygdala (BLA) is critical for neural function and synaptic plasticity in these brain regions. Synaptic plasticity in the neocortex, dHipp, ventral Hipp (vHipp), and BLA has also been implicated in fear and extinction memory. This finding raises the possibility that basal forebrain (BF) cholinergic neurons, the predominant source of acetylcholine in these brain regions, have an important role in mediating fear and extinction memory. While empirical studies support this hypothesis, there are interesting inconsistencies among these studies that raise questions about how best to define the role of BF cholinergic neurons in fear and extinction memory. Nucleus basalis magnocellularis (NBM) cholinergic neurons that project to the BLA are critical for fear memory and contextual fear extinction memory. NBM cholinergic neurons that project to the neocortex are critical for cued and contextual fear conditioned suppression, but are not critical for fear memory in other behavioral paradigms and in the inhibitory avoidance paradigm may even inhibit contextual fear memory formation. Medial septum and diagonal band of Broca cholinergic neurons are critical for contextual fear memory and acquisition of cued fear extinction. Thus, even though the results of previous studies suggest BF cholinergic neurons modulate fear and extinction memory, inconsistent findings among these studies necessitates more research to better define the neural circuits and molecular processes through which BF cholinergic neurons modulate fear and extinction memory. Furthermore, studies determining if BF cholinergic neurons can be manipulated in such a manner so as to treat excessive fear in anxiety disorders are needed.

Cholinergic neuronal lesions in the medial septum and vertical limb of the diagonal bands of Broca induce contextual fear memory generalization and impair acquisition of fear extinction.

Previous research has shown that the ventral medial prefrontal cortex (vmPFC) and hippocampus (Hipp) are critical for extinction memory. Basal forebrain (BF) cholinergic input to the vmPFC and Hipp is critical for neural function in these substrates, which suggests BF cholinergic neurons may be critical for extinction memory. In order to test this hypothesis, we applied cholinergic lesions to different regions of the BF and observed the effects these lesions had on extinction memory. Complete BF cholinergic lesions induced contextual fear memory generalization, and this generalized fear was resistant to extinction. Animals with complete BF cholinergic lesions could not acquire cued fear extinction. Restricted cholinergic lesions in the medial septum and vertical diagonal bands of Broca (MS/vDBB) mimicked the effects that BF cholinergic lesions had on contextual fear memory generalization and acquisition of fear extinction. Cholinergic lesions in the horizontal diagonal band of Broca and nucleus basalis (hDBB/NBM) induced a small deficit in extinction of generalized contextual fear memory with no accompanying deficits in cued fear extinction. The results of this study reveal that MS/vDBB cholinergic neurons are critical for inhibition and extinction of generalized contextual fear memory, and via this process, may be critical for acquisition of cued fear extinction. Further studies delineating neural circuits and mechanisms through which MS/vDBB cholinergic neurons facilitate these emotional memory processes are needed. © 2015 Wiley Periodicals, Inc.

Inhibiting corticosterone synthesis during fear memory formation exacerbates cued fear extinction memory deficits within the single prolonged stress model.

Using the single prolonged stress (SPS) animal model of post-traumatic stress disorder (PTSD), previous studies suggest that enhanced glucocorticoid receptor (GR) expression leads to cued fear extinction retention deficits. However, it is unknown how the endogenous ligand of GRs, corticosterone (CORT), may contribute to extinction retention deficits in the SPS model. Given that CORT synthesis during fear learning is critical for fear memory consolidation and SPS enhances GR expression, CORT synthesis during fear memory formation could strengthen fear memory in SPS rats by enhancing GR activation during fear learning. In turn, this could lead to cued fear extinction retention deficits. We tested the hypothesis that CORT synthesis during fear learning leads to cued fear extinction retention deficits in SPS rats by administering the CORT synthesis inhibitor metyrapone to SPS and control rats prior to fear conditioning, and observed the effect this had on extinction memory. Inhibiting CORT synthesis during fear memory formation in control rats tended to decrease cued freezing, though this effect never reached statistical significance. Contrary to our hypothesis, inhibiting CORT synthesis during fear memory formation disrupted extinction retention in SPS rats. This finding suggests that even though SPS exposure leads to cued fear extinction memory deficits, CORT synthesis during fear memory formation enhances extinction retention in SPS rats. This suggests that stress-induced CORT synthesis in previously stressed rats can be beneficial.

Sex differences in the single prolonged stress model.

Post traumatic stress disorder (PTSD) is a debilitating anxiety disorder resulting from traumatic stress exposure. Females are more likely to develop PTSD than males, but neurobiological mechanisms underlying female susceptibility are lacking. This can be addressed by using nonhuman animal models. Single prolonged stress (SPS), a nonhuman animal model of PTSD, results in cued fear extinction retention deficits and hippocampal glucocorticoid receptor (GR) upregulation in male rats. These effects appear linked in the SPS model, as well as in PTSD. However, the effects of SPS on cued fear extinction retention and hippocampal GRs in female rats remain unknown. Thus, we examined sex differences in SPS-induced cued fear extinction retention deficits and hippocampal GR upregulation. SPS induced cued fear extinction retention deficits in male rats but not female rats. SPS enhanced GR levels in the dorsal hippocampus of female rats, but not male rats. SPS had no effects on ventral hippocampal GR levels, but ventral hippocampal GR levels were attenuated in female rats relative to males. These results suggest that female rats are more resilient to the effects of SPS. The results also suggest that GR upregulation and cued fear extinction retention deficits can be dissociated in the SPS model.

Early handling attenuates enhancement of glucocorticoid receptors in the prefrontal cortex in an animal model of post-traumatic stress disorder.

BACKGROUND: Changes in glucocorticoid receptors (GRs) have been implicated in the pathogenesis of stress related psychiatric disorders such as depression and post-traumatic stress disorder (PTSD). Abnormal adaptation of the stress-response system following traumatic stress can lead to an altered hypothalamic-pituitary-adrenal axis that may contribute to PTSD development. Indeed, elevated GR expression in the hippocampus and prefrontal cortex linked to PTSD-like characteristics have been reported in the validated animal model of PTSD, single-prolonged stress. These findings implicate increased levels of GRs in the development of post-traumatic psychopathology and suggest that exploration of GR-targeted interventions may have potential for PTSD prevention. Early handling during the neonatal phase alters GR expression and is proposed to confer resilience to stress. We therefore examined the effects of combined early handling and single prolonged stress treatments on GR expression. METHODS: Timed pregnant dams gave birth to pups that were subjected to early handling (n = 11) or control (n = 13) procedures during the neonatal phase. At postnatal day 45 animals underwent single prolonged stress or a control procedure. Rats were euthanized one day later and GR levels were assayed using western blot electrophoresis. RESULTS: Single prolonged stress exposure enhanced GR expression in the hippocampus and prefrontal cortex. Early handling treatment protected against single prolonged stress-induced enhancement of GR expression in the prefrontal cortex, but not in the hippocampus. CONCLUSIONS: These data are a first step in highlighting the importance of targeting GR systems in prevention/resilience and may suggest that preventive strategies targeting GR upregulation might be particularly effective when prefrontal rather than hippocampal GRs are the target.

Altered locus coeruleus-norepinephrine function following single prolonged stress.

Data from preclinical and clinical studies have implicated the norepinephrine system in the development and maintenance of post-traumatic stress disorder. The primary source of norepinephrine in the forebrain is the locus coeruleus (LC); however, LC activity cannot be directly measured in humans, and previous research has often relied upon peripheral measures of norepinephrine to infer changes in central LC-norepinephrine function. To directly assess LC-norepinephrine function, we measured single-unit activity of LC neurons in a validated rat model of post-traumatic stress disorder - single prolonged stress (SPS). We also examined tyrosine hydroxylase mRNA levels in the LC of SPS and control rats as an index of norepinephrine utilisation. For electrophysiological recordings, 92 LC neurons were identified from 19 rats (SPS, 12; control, 7), and spontaneous and evoked responses to a noxious event (paw compression) were recorded. Baseline and restraint stress-evoked tyrosine hydroxylase mRNA expression levels were measured in SPS and control rats (n = 16 per group) in a separate experiment. SPS rats showed lower spontaneous activity but higher evoked responses, leading to an enhanced signal-to-noise ratio of LC neurons, accompanied by impaired recovery from post-stimulus inhibition. In concert, tyrosine hydroxylase mRNA expression in the LC of SPS rats tended to be lower at baseline, but was exaggerated following restraint stress. These data demonstrate persistent changes in LC function following stress/trauma in a rat model of post-traumatic stress, as measured by differences in both the electrophysiological properties of LC neurons and tyrosine hydroxylase mRNA transcription.