Depolarization-Secretion Coupling in Nerve Terminals

点击翻译按钮获取中文摘要

DESCRIPTION (provided by applicant): Although there is considerable evidence that the electrical activity of neuronal somata leads to the entry of Ca2+ and to the subsequent secretion of transmitters (i.e., Depolarization-secretion coupling), the molecular details of how ionic currents control the release of specific neuroactive substances from nerve terminals remain undetermined. Vasopressin (AVP) and oxytocin (OT) are synthesized by magnocellular neurons (MCN) of the hypothalamus and secreted from neurohypophysial (NH) terminals. OT neurons are characterized by a high frequency discharge during suckling which leads to the pulsatile release of OT. AVP neurons are characterized by their asynchronous phasic activity (bursting) during maintained AVP release. In both cases, it is the clustering, albeit with different time courses for each peptide, of spikes, which facilitates hormone release. We have discovered that there are different Calcium-channel subtypes in AVP vs. OT terminals, but that their biophysical properties cannot explain this differential facilitation of release. Therefore, we hypothesize that autocrine/paracrine feedback effects determine efficacy of bursting patterns of electrical activity to facilitate release of AVP vs. OT. ATP is thought to be co-released with the HNS peptides. Purines, such as ATP and adenosine, interact with specific receptors on neurons and glia, leading to a variety of effects. It is not known, however, whether these effects are at somata and/or synapses in the central nervous system (CNS). We have characterized the electrical and secretory effects on the HNS by exogenous purines, including effects on membrane ionic conductances in these CNS neurons vs. their nerve terminals. The HNS affords the unique opportunity of unraveling the complicated effects of endogenous purines in the CNS by comparing such effects on different neuronal compartments. Our goal is to determine membrane mechanisms that mediate endogenous purinergic- induced modifications of neurohormone secretion during physiological patterns of electrical stimulation. To achieve these objectives, perforated-patch recordings of Ca2+ and K+ currents will be made from identified, isolated nerve terminals and somata of the HNS. Effects on release will be compared between the intact HNS and NH terminals by the use of ELISAs and capacitance measurements. Loose patch-clamp recordings from nerve terminals and somata in the intact HNS will allow analysis of how bursting activity regulates peptide release in the complete system. These studies will provide a unique opportunity to determine if endogenous purinergic feedback regulation occurs at the terminals of CNS neurons.