Forced abstinence (FA) from alcohol has been shown to produce a variety of anxietyand depression-like symptoms in animal models. Somatostatin (SST) neurons, a subtype of GABAergic neurons found throughout the brain, are a novel neural target with potential treatment implications in affective disorders, yet their role in alcohol use disorders (AUD) remains to be explored. Here, we examined the neuroadaptations of SST neurons during forced abstinence from voluntary alcohol consumption. Following 6 weeks of two-bottle choice alcohol consumption and protracted forced abstinence, male and female C57BL/6J mice exhibited a heightened, but sex-specific, depressivelike behavioral profile in the sucrose preference test (SPT) and forced swim test (FST), without changes in anxiety-like behaviors in the elevated plus maze (EPM) and open field test (OFT). FST-induced cFos expressions in the prefrontal cortex (PFC) and ventral bed nucleus of the stria terminalis (vBNST) were altered in FA-exposed female mice only, suggesting a sex-specific effect of forced abstinence on the neural response to acute stress. SST immunoreactivity in these regions was unaffected by forced abstinence, while differences were seen in SST/cFos co-expression in the vBNST. No differences in cFos or SST immunoreactivity were seen in the lateral central nucleus of the amygdala (CEA) and the basolateral amygdala (BLA). Additionally, SST neurons in female mice displayed opposing alterations in the PFC and vBNST, with heightened intrinsic excitability in the PFC and diminished intrinsic excitability in the vBNST. These findings provide an overall framework of forced abstinence-induced neuroadaptations in these key brain regions involved in emotional regulation and processing.

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251 words Main Text: 6367 words Figures: 6 Dao et al. Forced Abstinence in Cortico-Amygdalar Regions 2 ABSTRACT Forced abstinence (FA) from alcohol has been shown to produce a variety of anxiety-and depression-like symptoms in animal models. Somatostatin (SST) neurons, a subtype of GABAergic neurons found throughout the brain, are a novel neural target with potential treatment implications in affective disorders, yet their role in alcohol use disorders (AUD) remains to be explored. Here, we examined the neuroadaptations of SST neurons during forced abstinence from voluntary alcohol consumption. Following six weeks of two-bottle choice alcohol consumption and protracted forced abstinence, male and female C57BL/6J mice exhibited a heightened, but sexspecific, depressive-like behavioral profile in the sucrose preference test (SPT) and forced swim test (FST), without changes in anxiety-like behaviors in the elevated plus maze (EPM) and open field test (OFT). FSTinduced cFos expressions in the prefrontal cortex (PFC) and ventral bed nucleus of the stria terminalis (vBNST)were altered in FA-exposed female mice only, suggesting a sex-specific effect of forced abstinence on the neural response to acute stress. SST immunoreactivity in these regions was unaffected by forced abstinence, while differences were seen in SST/cFos co-expression in the vBNST. No differences in cFos or SST immunoreactivity were seen in the lateral central nucleus of the amygdala (CEA) and the basolateral amygdala (BLA). Additionally, SST neurons displayed opposing alterations in the PFC and vBNST, with heightened intrinsic excitability in the PFC and diminished intrinsic excitability in the vBNST. These findings provide an overall framework of forced abstinence-induced neuroadaptations in these key brain regions involved in emotional regulation and processing.

BackgroundSomatostatin (SST) neurons in the prelimbic (PL) cortex mediate a variety of behavioral states, ranging from alcohol consumption to fear learning and avoidance-related behaviors. However, little is known about the role of somatostatin peptide signaling itself to cortical functioning and behavior. Here, we sought to characterize the unique physiological and behavioral roles of the SST peptide in the PL cortex.MethodsWe employed a combination of ex vivo electrophysiology, in vivo calcium monitoring, and in vivo peptide pharmacology to explore the role of SST neuron and peptide signaling in the mouse PL cortex. Whole-cell slice electrophysiology was conducted in C57BL/6J male and female mice in pyramidal and GABAergic neurons of the PL cortex to characterize the pharmacological mechanism of SST signaling. Fiber photometry recordings of GCaMP6f fluorescent calcium signals from SST neurons were conducted to characterize the activity profile of SST neurons during exploration of an elevated plus maze (EPM) and open field (OF). We further used local delivery of a broad SST receptor (SSTR) agonist into bilateral PL cortex to test causal effects of SST signaling on these same exploratory behaviors.ResultsSSTR activation broadly hyperpolarized layer 2/3 pyramidal neurons in the PL cortex in both male and female mice ex vivo, through both monosynaptic and polysynaptic GABA neuron-mediated mechanisms of action. This hyperpolarization was blocked by pre-application of the SSTR antagonist cyclo-somatostatin (cyclo-SST) and was non-reversible. SST neurons in PL were activated during EPM and OF exploration, indicating task-related recruitment of these neurons. Lastly, in line with this exploration-related activity profile, SSTR agonist administration directly into the PL enhanced open arm exploration in the EPM.ConclusionsHere we reveal a novel role for the SST peptide system within the PL cortex, by demonstrating a peptide-induced hypoexcitability of PL circuits and modulation of PL-dependent exploratory behaviors.