Voltage-activated Ca2+ channels (VACCs) mediate Ca2+ influx to trigger action potential-evoked neurotransmitter release but the mechanism by which Ca2+ regulates spontaneous transmission is unclear. Here we show VACCs are the major physiological triggers for spontaneous release at murine neocortical inhibitory synapses. Moreover, despite the absence of a synchronizing action potential, we find that spontaneous fusion of a GABA-containing vesicle requires the activation of multiple tightly-coupled VACCs of variable type.

Silent voltage-gated potassium channel subunits (KS) interact selectively with members of the K2 channel family to modify their functional properties. The localization and functional roles of these silent subunits remain poorly understood. Mutations in the KS subunit, K8.2 (), lead to severe visual impairment in humans, but the basis of these deficits remains unclear. Here, we examined the localization, native interactions, and functional properties of K8.2-containing channels in mouse, macaque, and human photoreceptors of either sex. In human retina, K8.2 colocalized with K2.1 and K2.2 in cone inner segments and with K2.1 in rod inner segments. K2.1 and K2.2 could be coimmunoprecipitated with K8.2 in retinal lysates indicating that these subunits likely interact directly. Retinal K2.1 was less phosphorylated than cortical K2.1, a difference expected to alter the biophysical properties of these channels. Using voltage-clamp recordings and pharmacology, we provide functional evidence for Kv2-containing channels in primate rods and cones. We propose that the presence of K8.2, and low levels of K2.1 phosphorylation shift the activation range of K2 channels to align with the operating range of rod and cone photoreceptors. Our data indicate a role for K2/K8.2 channels in human photoreceptor function and suggest that the visual deficits in patients with mutations arise from inadequate resting activation of K channels in rod and cone inner segments. Mutations in a voltage-gated potassium channel subunit, K8.2, underlie a blinding inherited photoreceptor dystrophy, indicating an important role for these channels in human vision. Here, we have defined the localization and subunit interactions of K8.2 channels in primate photoreceptors. We show that the K8.2 subunit interacts with different Kv2 channels in rods and cones, giving rise to potassium currents with distinct functional properties. Our results provide a molecular basis for retinal dysfunction in patients with mutations in the gene encoding K8.2.

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Coupling of an intracellular Ca(2+) clock to surface membrane ion channels, i.e., a "membrane clock, " via coupling of electrochemical Na(+) and Ca(2+) gradients (ENa and ECa, respectively) has been theorized to regulate sinoatrial nodal cell (SANC) normal automaticity. To test this hypothesis, we measured responses of [Na(+)]i, [Ca(2+)]i, membrane potential, action potential cycle length (APCL), and rhythm in rabbit SANCs to Na(+)/K(+) pump inhibition by the digitalis glycoside, digoxigenin (DG, 10-20 μmol/l). Initial small but significant increases in [Na(+)]i and [Ca(2+)]i and reductions in ENa and ECa in response to DG led to a small reduction in maximum diastolic potential (MDP), significantly enhanced local diastolic Ca(2+) releases (LCRs), and reduced the average APCL. As [Na(+)]i and [Ca(2+)]i continued to increase at longer times following DG exposure, further significant reductions in MDP, ENa, and ECa occurred; LCRs became significantly reduced, and APCL became progressively and significantly prolonged. This was accompanied by increased APCL variability. We also employed a coupled-clock numerical model to simulate changes in ENa and ECa simultaneously with ion currents not measured experimentally. Numerical modeling predicted that, as the ENa and ECa monotonically reduced over time in response to DG, ion currents (ICaL, ICaT, If, IKr, and IbNa) monotonically decreased. In parallel with the biphasic APCL, diastolic INCX manifested biphasic changes; initial INCX increase attributable to enhanced LCR ensemble Ca(2+) signal was followed by INCX reduction as ENCX (ENCX = 3ENa - 2ECa) decreased. Thus SANC automaticity is tightly regulated by ENa, ECa, and ENCX via a complex interplay of numerous key clock components that regulate SANC clock coupling.