Cholinergic transmission in molluscan neuroendocrine cells

Abstract

Elucidating the process by which an animal can transduce a brief signal into a predictable set of behaviours has important implications for understanding brain function. I explored the transition from quiescence to repetitive activity in the bag cell neurons of Aplysia californica. Using both cultured neurons and intact ganglia, I demonstrated the involvement of cholinergic transmission in this marked change to excitability. The bag cell neurons are a group of 200-400 electrically-coupled neuroendocrine cells that initiate a set of prescribed reproductive behaviours, culminating in deposition of an egg mass. This fixed action pattern, lasting ~1 h, follows a brief (≤10-sec) stimulus from an afferent input to the bag cell neuron cluster, which causes these previously silent neurons to continuously fire for ~30 min. Central to the maintenance of this increased excitability, are the elevation of various second messenger pathways that modulate multiple ion channels. As such, the initiating stimulus for afterdischarge generation was thought to involve metabotropic receptors. However, I report that an acetylcholine-gated ionotropic current triggers the afterdischarge, as well as two, distinct nicotinic responses that participate in excitability: one associated with channel opening and the other through the inhibition of K+ currents. My data suggest that the interplay between inward Ca2+ and cation currents, and outward K+ channels, regulated by intracellular messengers protein kinase C (PKC) and cyclic adenosine monophosphate (cAMP), respectively, set the baseline level of excitability prior to cholinergic activation. I also observed, distinct negative-feedback mechanisms on the acetylcholine ionotropic current. First, an increase in cAMP inhibits the cholinergic current shortly after the start of the afterdischarge, and once the afterdischarge is fully underway, dephosphorylation by a Src family tyrosine kinase further inhibits the channel. In addition, FMRFamide, an afterdischarge suppressor, appears to directly block the cholinergic channel. By exploring both canonical and non-canonical cholinergic roles in the afterdischarge, I have determined that complex signalling pathways can be reduced to a single variable, provided that the necessary precursors are in place. Furthermore, based on post-synaptic receptor composition and regulation, my work indicates the potential for profound diversity in cholinergic pathways.