Cue-induced reinstatement of alcohol-seeking behavior is associated with increased CaMKII T286 phosphorylation in the reward pathway of mice

Abstract

Cue-induced reinstatement of alcohol-seeking is a hallmark behavioral pathology of addiction. Evidence suggests that reinstatement (e.g., relapse), may be regulated by cell signaling systems that underlie neuroplasticity. A variety of plasticity events require activation of calcium calmodulin-dependent protein kinase II (CaMKII) in components of the reward pathway, such as the nucleus accumbens and amygdala. We sought to determine if cue-induced reinstatement of alcohol-seeking behavior is associated with changes in the activation state (e.g., phosphorylation) of CaMKII-T286. Male C57BL/6J mice (n = 14) were trained to lever press on a fixed-ratio-4 schedule of sweetened alcohol (2% sucrose + 9% EtOH) reinforcement. After 14-d of extinction (no cues or reinforcers), mice underwent a response-contingent reinstatement (n = 7) vs. an additional day of extinction (n = 7). Brains were removed immediately after the test and processed for evaluation of pCaMKII-T286 immunoreactivity (IR). Number of pCaMKII-T286 positive cells/mm2 was quantified from coronal brain sections using Bioquant Image Analysis software. Mice emitted significantly more responses on the alcohol vs. the inactive lever throughout the baseline phase with average alcohol intake of 1.1 ± 0.03 g/kg/1-h. During extinction, responses on the alcohol lever decreased to inactive lever levels by day 7. Significant cue-induced reinstatement of alcohol-seeking was observed during a single test with no effects on the inactive lever. Reinstatement was associated with increased pCaMKII-T286 IR specifically in amygdala (LA and BLA), nucleus accumbens (AcbSh), lateral septum, mediodorsal thalamus, and piriform cortex as compared to extinction control. Brain regions showing no change included the dorsal striatum, medial septum, cingulate cortex, habenula, paraventricular thalamus, and ventral hypothalamus. These results show response contingent cue-induced reinstatement of alcohol-seeking behavior is associated with selective increases in pCaMKII-T286 in specific reward- and memory-related brain regions of male C57BL/6J mice. Primary molecular mechanisms of associative learning and memory may regulate relapse in alcohol addiction.

Alcohol Drinking Exacerbates Neural and Behavioral Pathology in the 3xTg-AD Mouse Model of Alzheimer’s Disease

“…Moreover, cue-specific fear suggests altered AMY function (Phillips and LeDoux, 1992) and is consistent with known effects of alcohol and amygdala-based fear responses (Agoglia and Herman, 2018;McCool et al, 2010). The AMY also regulates positive reinforcing effects of alcohol (Besheer et al, 2003;Besheer et al, 2012;Cannady et al, 2017;Salling et al, 2016;Schroeder et al, 2003) and cue-induced relapse to alcohol-seeking behavior (Cannady et al, 2011;Salling et al, 2017;Schroeder et al, 2008), which suggests common neuroanatomical mechanisms of AD and alcohol addiction.…”

“…As part of ongoing work to discover neural mechanisms of alcohol addiction, we assessed the impact of voluntary alcohol intake on the prefrontal cortex (PFC) and amygdala (AMY) proteome in C57BL/6J mice Salling et al, 2016). These brain regions were chosen for analysis based on known involvement in the rewarding, or reinforcing, effects of alcohol (Agoglia et al, 2015a;Agoglia et al, 2015b;Besheer et al, 2003;Faccidomo et al, 2016;Faccidomo et al, 2015a;Hodge et al, 1996;Olive and Hodge, 2000;Salling et al, 2017;Schroeder et al, 2003;Schroeder et al, 2008;Ueno et al, 2001). In each study, non-dependent mice consumed alcohol (10% v/v) vs water or water only in the homecage for ~1 month (Figure 1A).…”

Individual Differences in Ethanol Drinking and Seeking Behaviors in Rats Exposed to Chronic Intermittent Ethanol Vapor Exposure is Associated with Altered CaMKII Autophosphorylation in the Nucleus Accumbens Shell

“…Further studies are, however, needed to confirm this hypothesis. Taken together, while our results in the nucleus accumbens shell region are coincidental, future mechanistic studies could evaluate a role for CaMKII autophosphorylation in enhanced drinking and seeking behaviors that drive individual differences in outbred rats [22][23][24][25][61][62][63].…”

“…Ca 2+ /calmodulin-dependent protein kinase II (CaMKII) is strongly implicated in the induction and maintenance of synaptic strengthening via autonomous activity and increases in autophosphorylation at T286 [18][19][20]. Mechanistic and correlative studies have demonstrated a role of CaMKII autophosphorylation in alcohol addiction-related behaviors [21][22][23][24][25]. For example, even though CaMKII autophosphorylation-deficient mice self-administer similar amounts of ethanol in an operant paradigm, they do not show psychomotor responses to acute and chronic alcohol injections during an ethanol-free state or during voluntary ethanol-drinking states [23,24].…”

Corticotropin-Releasing Factor Receptor-1 Neurons in the Lateral Amygdala Display Selective Sensitivity to Acute and Chronic Ethanol Exposure

“…The results of the voltage-clamp experiments suggest that this enhanced excitability in the CRF1 1 population is not driven by alterations in GABAergic signaling, which may indicate that these changes are instead regulated by ethanol-induced alterations in intrinsic excitability within the LA. Plasticity in glutamatergic signaling within the LA/BLA has been reported following chronic ethanol exposure (McCool et al, 2010), and the LA specifically exhibits alterations in molecular markers of glutamate signaling following chronic ethanol exposure in nonhuman primates (Alexander et al, 2018) and reinstatement of alcohol seeking in mice (Salling et al, 2017). Future work to characterize glutamatergic signaling in the CRF1 1 and CRF1 À populations of the LA both under basal conditions and following chronic ethanol exposure would help to clarify the mechanisms underlying these ethanol-induced changes in excitability.…”