Role of Munc13-1 and Munc13-2 Proteins in Alcoholism, and its Potential Clinical Applications

Munc13-1 and Munc13-2 are presynaptic proteins that are involved in the vesicular priming and subsequent release of glutamate neurotransmitter. Previous research has demonstrated some of the effects of ethanol in specific areas of the brain related to addiction, but the effects of ethanol on the expression of Munc13-1 and Munc13-2 and their glutamatergic effects in different brain regions is unknown. First, we performed immunocytochemistry (ICC) of Munc13-1 and Munc13-2 on ethanol-treated differentiated HT22 cells in-vitro and found that there was a significant change in the overall expression of Munc13-1 in a dose and time-dependent manner. Next, we harvested primary hippocampal neurons, treated them with ethanol in a dose and time-dependent manner, and then measured both Munc13-1 and Munc13-2 protein expression using Western-blot analysis. Here, our results showed that ethanol significantly upregulates the expression of both Munc13-1 and Munc13-2. We then tested the effects of ethanol on Munc13 proteins in-vivo on both wild-type (Wt) and heterozygous Munc13-1 knockout mice, using the drinking in the dark (DID) paradigm. In comparison to our previous ex-vivo data, these results showed that with alcohol exposure there was a significant increase in the expression of Munc13-1 in the hippocampus and cerebellum of Wt mice, but a decrease in the expression of Munc13-2 in the hippocampus, cerebellum and cortex. In the heterozygous Munc13-1 knockout mice, ethanol caused significant compensation for the loss of Munc13-1 in the hippocampus and cerebellum, and also an increase in Munc13-2 expression in the cerebellum and cortex. Since both Munc13-1 and Munc13-2 predominantly control glutamatergic synapses, their modulation by ethanol exposure could potentiate an increase in glutamate release from the pre-synaptic terminals of these regions and stimulate excitation of the Central Nervous System (CNS). This is analogous to the CNS excitation we see with chronic AUD patients experiencing alcohol withdrawal symptoms. By identifying these proteins as drug targets, we may be able to unfold a different mechanism of alleviating alcohol withdrawal symptoms in AUD patients, without the GABAergic adverse effects that usually present with the current mainstay treatment options.