ABSTRACT
Clozapine binds and has significant effects on multiple neurotransmitter receptors, notably including some dopamine receptors. Downstream of these receptors, clozapine affects the balance of Gi− and Gq-dependent second-messenger signaling. We used Caenorhabiditis elegans as a genetic model to study further how clozapine affects both dopamine receptors and downstream Gq mediated signaling. Four of six worm dopamine receptor orthologs, dop-1, dop-2, dop-4, and dop-5 produced resistance to clozapine induced developmental delay when mutated, suggesting that both type I and type II dopamine receptors mediate the behavioral effects of clozapine in C. elegans. Beyond these receptors, reduction of function of one of the G proteins, egl-30 (Gαq), produced greatly increased susceptibility to clozapine. Gαq has multiple known downstream effects. Among these is the control of acetylcholine release, which is in balance with monoamines in the human brain and is another target of clozapine and other antipsychotic drugs. We tested for downstream effects on acetylcholine at the neuromuscular junction upon clozapine treatment but found no evidence for effects of clozapine. In contrast, modulation of Gαq upstream leads to worms that are either more resistant or more susceptible to clozapine, emphasizing the importance of Gαq proteins in mediating effects of clozapine. A genetic screen for suppressors of egl-30 recovered eight mutants. By characterizing the behavioral effects of these mutants, we found that clozapine exerts its function on development by affecting Gαq signaling through control of the pharyngeal pumping rate. A whole-genome sequencing technique was utilized and identified a list of candidate genes for these suppressor mutations. Further characterization of these mutants promises the discovery of novel components participating in Gαq signaling and a better understanding of the mechanisms of action of clozapine.
