Presynaptic neurons control postsynaptic GABAa receptor trafficking in Caenorhabditis elegans

Inhibitory neurotransmission is critical for regulating neuronal excitability. The main inhibitory neurotransmitter is gamma-aminobutyric acid (GABA). It acts predominantly at postsynaptic GABA(A) receptors. GABAA receptors are heteropentameric chloride ion channels. Receptor activation results in chloride influx, which rapidly reduces membrane excitability. Balance between inhibition and excitation is tightly regulated to ensure efficient neurotransmission. Significant reduction in GABA(A) receptor efficacy causes anxiety and epilepsy. GABA(A) receptor-agonists are the primary treatment for these disorders. Although regulation of these receptors plays a major role in controlling excitability, we understand very little about the physiological mechanisms underlying the development and maintenance of inhibitory synapses. Mammalian inhibitory synapses are difficult to study, in vivo, due to limited genetic accessibility as well as receptor heterogeneity. In contrast, Caenorhabditis elegans is well suited to study these synapses. Its nervous system is genetically accessible and its GABA (A) receptor is simple and uniform. It functions in bodywall muscles to promote muscle relaxation during locomotion. Normally, GABA (A) receptors are localized at synapses. We reasoned that if genes required for proper receptor trafficking were mutated, synaptic localization of these receptors would be lost. We performed a simple genetic screen to identify mutants with mislocalized receptors, and isolated mutants with intracellular accumulation of GABA (A) receptors in large organelles. One of these mutants carried a mutation in the unc-3 gene. UNC-3 acts in neurons, yet our phenotype is in muscles. To explain this conundrum, we hypothesized that presynaptic neurons control postsynaptic GABA(A) receptors. To test this hypothesis, we examined GABA(A) receptor localization in an axonal pathfinding mutant that removes innervation from postsynaptic cells. Normally these cells receive GABA and ACh innervation. In cells lacking both types of inputs, we observed UNC-49 accumulation in similar organelles. We identified these organelles as autophagosomes. Autophagosomes deliver cellular components to the lysosome for degradation. Selective restoration of either cholinergic or GABAergic innervation suppresses GABA(A) receptor autophagy; however, only GABAergic motor neurons induce receptor clustering. We conclude first that presynaptic terminals induce GABA(A) receptor cluster formation by independently regulating receptor localization and receptor stability and second, that autophagy is a novel degradative trafficking pathway for GABA(A) receptors.