Chronic intermittent ethanol (CIE) exposure in rats induces neuroplastic adaptations in the prelimbic cortex (PL) and nucleus accumbens core (NAcore) during withdrawal, contributing to deficits in decision-making and escalation of alcohol consumption. Increased evidence has linked neuroplastic changes associated with the development of alcohol dependence and withdrawal to neuroimmune signaling and activation of microglia. Microglia exhibit a well-defined structure/function relationship, with alterations in cytokine release occurring concomitantly with reconfiguration of cellular dimensions and redistribution of fine processes. Accordingly, gaining a better understanding of ethanol-induced microglia activation will provide insight into how neuroimmune signaling may impact neural structure and function following ethanol exposure. The present study characterized CIE-induced alterations in microglia morphology in the PL and NAcore using confocal microscopy and 3D image reconstruction.Rats were sacrificed at peak withdrawal, 10 hours after discontinuation of 15 days of CIE or Air exposure. CIE-exposed rats exhibited an increase in microglia soma size in the PL and NAcore compared to Air-exposed control rats. However, the CIE-mediated increase in soma volume in the PL cortex was accompanied by a reduction in microglial complexity. While systemic LPS elicited similar increases in soma volume in the PL cortex and NAcore, the LPS-induced increase in soma size was associated with increased complexity of microglial processes in the PL cortex, an effect opposite to that of CIE. These data are consistent with the hypothesis that the microglial activation state following CIE is structurally and chemically distinct from the activation state associated with induction of the innate neuroimmune system following systemic LPS exposure.

Abbreviations: SA (self-administration), NO (nitric oxide), MSNs (medium spiny neurons), nNOS (neuronal nitric oxide synthase), NAcore (nucleus accumbens core), PL (prelimbic), vSub (ventral subiculum), VTA (ventral tegmental area), GLT-1 (glutamate transporter-1), BLA (basolateral amygdala), DETA (DETA-NONOate), CNO (Clozapine-n-oxide), FR (Fixed ratio), standard error of the mean (SEM), eGFP (enhanced green fluorescent protein), cGMP (cyclic GMP), sGC (soluble guanylyl cyclase), CPP (conditioned place preference), MMPs (matric metallo-proteinases), i.v. (intravenous), LOD (limit of detection), GluOx (glutamate oxidase) AbstractCue-induced reinstatement of cocaine seeking after self-administration (SA) and extinction relies on glutamate release in the nucleus accumbens core (NAcore), which in turn activates neuronal nitric oxide synthase (nNOS) interneurons. Nitric oxide (NO) is required for structural plasticity in NAcore medium spiny neurons (MSNs), as well as cued cocaine seeking. However, NO release in the NAcore during reinstatement has yet to be directly measured. Further, the temporal relationship between glutamate release, and the induction of a NO response also remains unknown. Using wireless amperometric recordings in awake behaving rats, we quantified the magnitude and temporal dynamics of novel context-and cue-induced reinstatement-evoked glutamate and NO release in the NAcore. We found that re-exposure to cocaine-conditioned stimuli following SA and extinction increased extracellular glutamate, leading to release of NO in the NAcore. In contrast, exposing drug-naïve rats to a novel context led to a lower magnitude rise in glutamate in the NAcore relative to cue-induced reinstatement. Interestingly, novel context exposure evoked a higher magnitude NO response relative to cue-induced reinstatement. Despite differences in magnitude, novel context evoked-NO release in the NAcore was also temporally delayed when compared to glutamate. These results demonstrate a dissociation between the magnitude of cocaine cue-and novel context-evoked glutamate and NO release in the NAcore, yet similarity in the temporal dynamics of their release. Together, these data contribute to a greater understanding of the relationship between glutamate and NO, two neurotransmitters implicated in encoding the valence of distinct contextual stimuli.