Differential Effects of Single Versus Repeated Alcohol Withdrawal on the Expression of Endocannabinoid System‐Related Genes in the Rat Amygdala

Abstract

Background Endogenous cannabinoids such as anandamide and 2-arachidonoylglycerol (2-AG) exert important regulatory influences on neuronal signaling, participate in short- and long-term forms of neuroplasticity, and modulate stress responses and affective behavior in part through the modulation of neurotransmission in the amygdala. Alcohol consumption alters brain endocannabinoid levels, and alcohol dependence is associated with dysregulated amygdalar function, stress responsivity and affective control. Methods The consequence of long-term alcohol consumption on the expression of genes related to endocannabinoid signaling was investigated using quantitative RT-PCR analyses of amygdala tissue. Two groups of ethanol-exposed rats were generated by maintenance on an ethanol liquid diet (10%): one group received continuous access to ethanol for 15 days, while the second group was given intermittent access to the ethanol diet (5 days/week for 3 weeks). Control subjects were maintained on an isocaloric ethanol-free liquid diet. To provide an initial profile of acute withdrawal amygdala tissue was harvested following either 6 or 24 hours of ethanol withdrawal. Results Acute ethanol withdrawal was associated with significant changes in mRNA expression for various components of the endogenous cannabinoid system in the amygdala. Specifically, reductions in mRNA expression for the primary clearance routes for anandamide and 2-AG (FAAH and MAGL, respectively) were evident, as were reductions in mRNA expression for CB1, CB2 and GPR55 receptors. Although similar alterations in FAAH mRNA were evident following either continuous or intermittent ethanol exposure, alterations in MAGL and cannabinoid receptor-related mRNA (e.g. CB1, CB2, GPR55) were more pronounced following intermittent exposure. In general, greater withdrawal-associated deficits in mRNA expression were evident following 24 versus 6 hours of withdrawal. No significant changes in mRNA expression for enzymes involved in 2-AG biosynthesis (e.g. DAGL-α/β) were found in any condition. Conclusions These findings suggest that ethanol dependence and withdrawal are associated with dysregulated endocannabinoid signaling in the amygdala. These alterations may contribute to withdrawal-related dysregulation of amygdalar neurotransmission.

Blockade of alcohol escalation and “relapse” drinking by pharmacological FAAH inhibition in male and female C57BL/6J mice

“…Together, this indicates that global NAE dysregulation occurs as a response to acute alcohol withdrawal only. This transient increase may be due to decreased FAAH activity, as found in the rats after 1-day alcohol withdrawal [Serrano et al 2012], but not the FAAH expression level as observed in the present study (Figure 8). To our knowledge, this is the first description of global NAE dysregulation, both in terms of multi-localization and number of endogenous substrates (AEA, OEA, and PEA), in response to acute alcohol withdrawal, and indicates that a strong eCB response may act as an important molecular coping mechanism in acute alcohol withdrawal, thereby allowing the mice to better cope with alcohol withdrawal stress.…”

“…eCBs exert prominent modulatory influence on the extended amygdala and corticostriatal circuits, and stress exposure disrupts eCB-enriched regions that participate in emotional control [Serrano et al 2012; Dincheva et al 2015; Morena et al 2016]. Specifically, the relatively deficient eCB function is associated with increased anxiety and depression, and as such, impaired eCB activity may contribute to the negative affective states and increased stress responsivity that underlie negative reinforcement mechanisms driving alcohol drinking by dependent individuals and that contribute to alcohol relapse following periods of abstinence [Parsons & Hurd, 2015].…”

Involvement of Activated Brain Stress Responsive Systems in Excessive and “Relapse” Alcohol Drinking in Rodent Models: Implications for Therapeutics

“…eCBs have considerable modulatory effects on the extended amygdala and corticostriatal circuitries, and stress disrupts these eCB-enriched regions that are involved in emotional control [Serrano et al, 2012;Dincheva et al, 2015;Morena et al, 2016]. Pharmacologic and genetic manipulations (knockout or knock-in) of FAAH are found to alter anxiety-like and depression-like behaviors [Bortolato et al, 2007;Gunduz-Cinar et al, 2013;Kathuria et al, 2003;Moreira et al, 2008;Carnevali et al, 2015].…”

Stress regulates endocannabinoid-CB1 receptor signaling

“…On the other hand, CB1R binding was decreased in the supraoptic hypothalamic nuclei and ventrolateral thalamus and increased in subregions of the caudate putamen in singly-housed compared to group-housed rats [98]. Repeated episodes of alcohol withdrawal, which produces anxiety in rodents [99], resulted in a significant reduction of CB1R mRNA in the amygdala of male rats [100]. …”