Serotonin and neuropeptides are both released by the HSN command neuron to initiate C. elegans egg laying

Abstract

1Neurons typically release both a small molecule neurotransmitter and one or more neuropeptides, but how these two types of signal from the same neuron might act together remains largely obscure. For example, serotonergic neurons in mammalian brain express the neuropeptide Substance P, but it is unclear how serotonin signaling might be modulated by a coreleased neuropeptide. We studied this issue in C. elegans, in which all serotonergic neurons express the neuropeptide NLP-3. The serotonergic Hermaphrodite Specific Neurons (HSNs) are command motor neurons within the egg-laying circuit that have previously been shown to release serotonin to initiate egg-laying behavior. We found that egg-laying defects in animals lacking serotonin were far milder than in animals lacking HSNs, suggesting that HSNs must release other signal(s) in addition to serotonin to stimulate egg laying. While null mutants for nlp-3 had only mild egg-laying defects, animals lacking both serotonin and NLP-3 had severe defects, like those of animals lacking HSNs. Optogenetic activation of HSNs induced egg laying in wild-type animals, or in mutant animals lacking either serotonin or NLP-3, but failed to induce egg laying in animals lacking both. We recorded calcium activity in the egg-laying muscles of animals lacking either serotonin, NLP-3, or both. The single mutants, and to a greater extent the double mutant, showed muscle activity that was uncoordinated and unable to expel eggs, such that the vm2 muscles cells that are direct postsynaptic targets of the HSN failed to contract simultaneously with other egg-laying muscle cells. Our results show that the HSN neurons use serotonin and the neuropeptide NLP-3 as partially redundant cotransmitters that together stimulate and coordinate activity of the target cells onto which they are released.