Descripción: In the mammalian auditory system, the synapse between efferent olivocochlear (OC) neurons and sensory cochlear hair cells is cholinergic, fast, and inhibitory. This efferent synapse is mediated by the nicotinic α9α10 receptor coupled to the activation of SK2 Ca 2+-activated K+ channels that hyperpolarize the cell. So far, the ion channels that support and/or modulate neurotransmitter release from the OC terminals remain unknown. To identify these channels, we used an isolated mouse cochlear preparation and monitored transmitter release from the efferent synaptic terminals in inner hair cells (IHCs) voltage clamped in the whole-cell recording configuration. Acetylcholine (ACh) release was evoked by electrically stimulating the efferent fibers that make axosomatic contacts with IHCs before the onset of hearing. Using the specific antagonists for P/Q- and N-type voltage-gated calcium channels (VGCCs), ω-agatoxin IVA and ω-conotoxin GVIA, respectively, we show that Ca2+ entering through both types of VGCCs support the release process at this synapse. Interestingly, we found that Ca2+ entering through the dihydropiridine-sensitive L-type VGCCs exerts a negative control on transmitter release. Moreover, using immunostaining techniques combined with electrophysiology and pharmacology, we show that BK Ca2+-activated K+ channels are transiently expressed at the OC efferent terminals contacting IHCs and that their activity modulates the release process at this synapse. The effects of dihydropiridines combined with iberiotoxin, a specific BK channel antagonist, strongly suggest that L-type VGCCs negatively regulate the release of ACh by fueling BK channels that are known to curtail the duration of the terminal action potential in several types of neurons. Copyright © 2010 the authors.

Descripción: 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. © 2013 Álvarez et al.