The roles of the CaVbeta2 subunit and the synaptic protein interaction site in determining Ca2+ channel function in neuroendocrine secretion

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The flow of Ca2+ into neurons through voltage-gated Ca2+ (CaV) channels triggers the release of neurotransmitters and hormones and the intracellular targeting of these channels is therefore of fundamental importance for intercellular communication. CaV channels are composed of a CaValpha1 subunit and one of four CaVbeta subunits, which may help transport channels to axon terminals. The two types of Ca2+ channels most important in synaptic release (CaV2.1 and CaV2.2) contain a peptide sequence that allows them to interact with synaptic proteins and is thus called the "synprint" site.****We propose to study the roles of the CaVbeta2 subunit and the synprint site in central neurons called the magnocellular neurosecretory cells (MNCs), which secrete the hormones oxytocin and vasopressin. Release occurs from both MNC cell bodies and terminals and increases as plasma osmolality increases. We found that the CaVbeta2 is enriched in MNC terminals, suggesting that it might play a role in CaV transport. We have also shown that MNCs express a variant of CaV2.1 that lacks the synprint site, but do not express an equivalent variant of CaV2.2. These data may explain the observation that CaV2.2, but not CaV2.1, triggers release of oxytocin from the MNC cell bodies.****We propose to test 1) whether the CaVbeta2 subunit plays a role in axonal targeting of CaV channels and 2) whether the interaction between the synprint site of CaV2.2 and synaptic proteins plays a role in triggering release of oxytocin from MNC cell bodies.****We will compare CaV expression in the MNC terminals of transgenic mice that lack the CaVbeta2 subunit in the brain to that of control littermates and predict that the expression will be lower. We further predict that these mice will fail to release vasopressin properly. We will test this by measuring vasopressin levels and plasma osmolality of control and knockout mice exposed to osmotic stress. We will test the role of the synprint site in facilitating neuropeptide secretion in MNC somata and terminals by interfering with the interaction between Ca2+ channels and synaptic proteins and measuring oxytocin release from MNC cell bodies. We predict that the disruption of this interaction will inhibit the ability of influx through CaV2.2 to trigger release, which would support a role for this interaction in hormone release. These studies will therefore help elucidate the mechanisms that allow neurons to regulate the release of hormones and neurotransmitters.***