Neurexin mediates neuropeptide release from cholinergic motor neurons through dense-core vesicle localization

Neurexins are critical synaptic cell adhesion molecules that play many roles in modulating neurotransmitter release and synaptic function, and are high-confidence risk genes for neurodevelopmental conditions such as autism. Understanding the function of the neurexin superfamily has been challenging in mammalian systems that have 3 genes (NRXN1-3) that encode 2-3 major isoforms, which undergo extensive alternative splicing and generate thousands of transcripts. In contrast to mammals, C. elegans has a single gene, nrx-1, encoding only long alpha and short gamma isoforms. Neurexins canonically regulate synapse morphology and function in a neuron- and context-specific manner, through mechanisms related to release of chemical neurotransmitters and receptors. Whether neurexins (nrx-1) impact other secretory molecules such as neuropeptides (NPs) and NP containing dense-core vesicles (DCVs) is not well understood. Here, we report that nrx-1 regulates the release of multiple NPs from cholinergic motor neurons in C. elegans. Using tissue specific expression and degradation of endogenous NRX-1, we find that nrx-1 functions in NP release in a cell-autonomous manner and that the short gamma-isoform is required to regulate NP secretion from the cholinergic neurons. We confirm that loss of nrx-1 gamma-isoform impacts cholinergic active-zone number, but also find it regulates the clustering, distribution, and expression of the DCV protein, IDA-1 (PTPRN), and the DCV secretion regulator, UNC-31 (CADPS). We find that nrx-1 functions to maintain separation and juxtaposition of neurotransmitter and NP release sites and DCV localization. Loss of cholinergic excitation (unc-17) or GABAergic inhibition (unc-25) did not impact cholinergic NP release, but that the increased NP release upon loss of nrx-1 is dependent on the calcium channel unc-2. We find that neurexins can regulate NP signaling, a novel mechanism to modify circuits and behaviors, and of potential importance for NRXN1 associated human conditions.