A Novel Cholinergic Action of Alcohol and the Development of Tolerance to That Effect inCaenorhabditis elegans

Abstract

Understanding the genes and mechanisms involved in acute alcohol responses has the potential to allow us to predict an individual's predisposition to developing an alcohol use disorder. To better understand the molecular pathways involved in the activating effects of alcohol and the acute functional tolerance that can develop to such effects, we characterized a novel ethanolinduced hypercontraction response displayed by Caenorhabditis elegans. We compared body size of animals prior to and during ethanol treatment and showed that acute exposure to ethanol produced a concentration-dependent decrease in size followed by recovery to their untreated size by 40 min despite continuous treatment. An increase in cholinergic signaling, leading to muscle hypercontraction, is implicated in this effect because pretreatment with mecamylamine, a nicotinic acetylcholine receptor (nAChR) antagonist, blocked ethanol-induced hypercontraction, as did mutations causing defects in cholinergic signaling (cha-1 and unc-17). Analysis of mutations affecting specific subunits of nAChRs excluded a role for the ACR-2R, the ACR-16R, and the levamisole-sensitive AChR and indicated that this excitation effect is dependent on an uncharacterized nAChR that contains the UNC-63 a-subunit. We performed a forward genetic screen and identified eg200, a mutation that affects a conserved glycine in EAT-6, the a-subunit of the Na + /K + ATPase. The eat-6(eg200) mutant fails to develop tolerance to ethanol-induced hypercontraction and remains contracted for at least 3 hr of continuous ethanol exposure. These data suggest that cholinergic signaling through a specific a-subunit-containing nAChR is involved in ethanol-induced excitation and that tolerance to this ethanol effect is modulated by Na + /K + ATPase function.T HE abuse of alcohol is a cause of significant societal and health-related problems. Despite the common usage of this drug, the molecular underpinnings of alcohol's acute actions on the brain are poorly understood. Alcohol (ethanol) has biphasic behavioral effects in humans and other animals, acting as a stimulant at lower concentrations and as a depressant at higher concentrations. Understanding the molecular nature of these biphasic effects is made difficult by the fact that ethanol interacts with, and alters the function of, many proteins, including neurotransmitter receptors and ion channels (Harris et al. 2008;Spanagel 2009). A commonly suggested class of ethanol targets is ligand-gated ion channels (LGICs), which include NMDA glutamate receptors, and members of a subclass of LGICs, the Cys-loop superfamily, including GABA A , glycine, and nicotinic acetylcholine receptors (nAChRs) (Olsen et al. 2014). Effects on the GABA A and glutamate receptors are hypothesized to play a significant role in the depressant effects of ethanol (Harris et al. 2008). Locomotor activation by low to moderate concentrations of ethanol in rodents is thought to model the euphoric effects of ethanol in humans (Phillips and Shen 1996). Locomotor activat...

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Na+/K+-pump and neurotransmitter membrane receptors

“…Ethanol can cause hypercontraction of C. elegans through activation of a novel α subunit associated with the cholinergic body wall muscle receptor (Hawkins et al 2015 ). This hypercontraction can reverse after 40 min despite the continual presence of ethanol, indicating ethanol tolerance.…”

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