The Immediately Releasable Pool of Mouse Chromaffin Cell Vesicles Is Coupled to P/Q-Type Calcium Channels via the Synaptic Protein Interaction Site

Abstract

It is generally accepted that the immediately releasable pool is a group of readily releasable vesicles that are closely associated with voltage dependent Ca2+ channels. We have previously shown that exocytosis of this pool is specifically coupled to P/Q Ca2+ current. Accordingly, in the present work we found that the Ca2+ current flowing through P/Q-type Ca2+ channels is 8 times more effective at inducing exocytosis in response to short stimuli than the current carried by L-type channels. To investigate the mechanism that underlies the coupling between the immediately releasable pool and P/Q-type channels we transiently expressed in mouse chromaffin cells peptides corresponding to the synaptic protein interaction site of Cav2.2 to competitively uncouple P/Q-type channels from the secretory vesicle release complex. This treatment reduced the efficiency of Ca2+ current to induce exocytosis to similar values as direct inhibition of P/Q-type channels via ω-agatoxin-IVA. In addition, the same treatment markedly reduced immediately releasable pool exocytosis, but did not affect the exocytosis provoked by sustained electric or high K+ stimulation. Together, our results indicate that the synaptic protein interaction site is a crucial factor for the establishment of the functional coupling between immediately releasable pool vesicles and P/Q-type Ca2+ channels.

The Physiology, Pathology, and Pharmacology of Voltage-Gated Calcium Channels and Their Future Therapeutic Potential

“…The ensuing entry of calcium ions then triggers the fusion of synaptic vesicles, culminating in the release of neurotransmitters into the synaptic cleft. The three major Ca V 2 channel isoforms support not only rapid neurotransmitter release but also hormone release from secretory cells such as chromaffin cells (Santana et al, 1999;Albillos et al, 2000;Wykes et al, 2007;Álvarez et al, 2013).…”

Ca2+‐independent and voltage‐dependent exocytosis in mouse chromaffin cells

“…The most recognized and well characterized voltage sensors in excitable cells are voltage‐dependent ion channels. It is known that some VDCC are functionally and spatially associated with the exocytotic machinery and with immediately releasable vesicles in neurons and neuroendocrine cells . Therefore, VDCC are major candidates to be the voltage sensors responsible of CIVDE.…”

“…Consequently, the addition of an exogenous free synprint peptide that competes with the endogenous synprint site of the Ca 2+ channel molecule disrupts vesicle‐channel interaction and inhibits neurotransmission . Particularly in chromaffin cells we showed previously that the exocytosis of IRP is markedly reduced in presence of the free synprint, and concluded that synprint is important in the coupling between IRP vesicles and P/Q‐type Ca 2+ channels . In order to evaluate if synprint mediates CIVDE, we transfected our cells with an IRES plasmid encoding the synprint peptide and EGFP.…”

“…In chromaffin cells, fast synchronous secretion induced by short depolarizations is generally attributed to the exocytosis of the immediately releasable pool (IRP) of vesicles. It is accepted that IRP in chromaffin and other neuroendocrine cells is a group of ready releasable secretory vesicles highly coupled to Ca 2+ entry through VDCC . However, the short depolarizations used experimentally to activate IRP exocytosis might also potentially trigger a Ca 2+ ‐independent but voltage‐dependent exocytosis (CIVDE) mechanism.…”

Expression of CaV2.2 and Splice Variants of CaV2.1 in Oxytocin‐ and Vasopressin‐Releasing Supraoptic Neurones

“…An alternative mechanism for the preferential role of Ca V 2.2 in somatodendritic release of OT is an interaction between the synprint site and exocytotic proteins. The immediately releasable pool of granules in mouse chromaffin cells is coupled to Ca V 2.1 channels through an interaction mediated by the synprint site and whole cell capacitance measurements have suggested that a rapidly releasable pool of granules exists in MNC somata . It is therefore possible that a pool of granules in OT MNC somata may be coupled to Ca V 2.2 channels via an interaction between the synprint site and the complexes of exocytotic proteins attached to those granules.…”