The diversity of Ca2+ channel types in rat cerebellar granule neurons was investigated with whole-cell recordings (5 mM external Ba2+). Contributions of five different high-voltage-activated Ca2+ channel current components were distinguished pharmacologically. Nimodipine-sensitive L-type current and omega-CTx-GVIA-sensitive N-type current contributed 15 and 20% of the total current, respectively. The bulk of the remaining current (46%) was inhibited by omega-Aga-IVA. The current blocked by this toxin was further subdivided into two components, P-type and Q-type, on the basis of differences in their inactivation kinetics and sensitivity to omega-Aga-IVA. P-Type current was noninactivating during 0.1 sec depolarizations, half-blocked at about 1-3 nM omega-Aga-IVA, and contributed approximately 11% of the total current; Q-type current was prominently inactivating, half-blocked at approximately 90 nM omega-Aga-IVA, and comprised 35% of the total current. Both P- and Q-type currents were potently inhibited by the Conus magus toxin omega-CTx-MVIIC. A current component resistant to all of the aforementioned blockers (R-type) displayed more rapid inactivation than the other components and constituted 19% of the total current. The Q-type current, the largest of the current components in the granule neurons, resembles currents that can be generated in Xenopus oocytes by expression of cloned alpha 1A subunits.

This study describes a Ca2+ store in fura-2-loaded bullfrog sympathetic neurons that modulates [Ca2+]i responses elicited by either depolarization or Ca2+ release from a caffeine- and ryanodine-sensitive store. This store is insensitive to caffeine and ryanodine, but is sensitive to the protonophore carbonyl cyanide p-trifluoromethoxyphenylhydrazone (FCCP). The FCCP-sensitive store slows both the rise in [Ca2+]i during stimulation (apparently by accumulating Ca2+ from the cytosol) and the recovery following stimulation (by releasing the accumulated Ca2+ into the cytosol). For a fixed level of depolarization, recovery is slowed to an extent that depends on stimulus duration. [Ca2+]i imaging shows that these effects are prominent in the soma but not in growth cones. Ca2+ uptake by the FCCP-sensitive store appears to be strongly [Ca2+]i dependent, since it becomes influential only when [Ca2+]i approaches approximately 500 nM. Therefore, this store may specifically influence [Ca2+]i during moderate and strong stimulation. The effect of the store on responses to depolarization can be accounted for by a simple three-compartment scheme consisting of the extracellular medium, the cytosol, and a single internal store with a [Ca2+]i-dependent uptake mechanism resembling the mitochondrial Ca2+ uniporter. The store's effect on responses to caffeine-induced Ca2+ release can be accounted for by including a second internal compartment to represent the caffeine-sensitive store. While the identity of the FCCP-sensitive store is unknown, its sensitivity to FCCP is consistent with a mitochondrial pool. It is suggested that by modulating the temporal properties of [Ca2+]i following stimulation, the FCCP-sensitive store may influence the degree of activation of intracellular [Ca2+]i-dependent processes.

It has been established that multiple types of Ca2+ channels participate in triggering neurotransmitter release at central synapses, but there is uncertainty about the nature of their combined actions. We investigated synaptic transmission at CA3-CA1 synapses of rat hippocampal slices and asked whether the dependence on omega-CTx-GVIA-sensitive N-type channels and omega-Aga-IVA-sensitive P/Q-type Ca2+ channels can be altered by physiological mechanisms. The reliance on multiple types of Ca2+ channels was not absolute but depended strongly on the amount of Ca2+ influx through individual channels, which was manipulated by prolonging the presynaptic action potential with the K+ channel blocker 4-aminopyridine (4-AP) and by varying the extracellular Ca2+ concentration ([Ca2+]o). We quantified the influence of spike broadening on Ca2+ influx through various Ca2+ channels by imposing mock action potentials on voltage-clamped cerebellar granule neurons. In field recordings of the EPSP in hippocampal slices, action potential prolongation increased the EPSP slope by 2-fold and decreased its reliance on either N-type or P/Q-type Ca2+ channels. The inhibition of synaptic transmission by N-type channel blockade was virtually eliminated in the presence of 4-AP, but it could be restored by lowering [Ca2+]o. These results rule out a scenario in which a significant fraction of presynaptic terminals rely solely on N-type channels to trigger transmission. The change in sensitivity to the neurotoxins with 4-AP could be explained in terms of a nonlinear relationship between Ca2+ entry and synaptic strength, which rises steeply at low [Ca2+]o, but approaches saturation at high [Ca2+]o. This relationship was evaluated experimentally by varying [CA2+]o in the absence and presence of 4-AP. One consequence of this relationship is that down-modulation of presynaptic Ca2+ channels by various modulators would increase the relative impact of spike broadening greatly.

No abstract