Involvement Of L-type Channels Research Articles

The Contribution of L-Type Calcium Channels to Acetylcholine Secretion in Frog and Mouse Neuromuscular Junctions with Active and Inactivated Voltage-Gated Potassium Channels

We report here experiments addressing the contribution of L-type Ca2+ channels to evoked acetylcholine secretion from frog and mouse motor nerve endings with active and inactivated voltage-gated K+ channels. These studies evaluated the effects of the specific L-type Ca2+ channel blocker nitrendipine on the quantum composition of endplate currents and the time course of the secretion of acetylcholine quanta in intact preparations and after preliminary blockade of voltage-gated K+ channels with 4-aminopyridine (4-AP) in medium with depressed and physiological Ca2+ levels. A fluorescence method was used to measure calcium transients reflecting the integral influx of Ca2+ into nerve endings; computer modeling was applied to the processes underlying exocytosis in the presence of the two types of Ca2+ channel (N and L) and with different durations of nerve ending action potentials. In frog synapses, L-type Ca2+ channels were found to contribute to evoked acetylcholine secretion in the presence of active K+ channels, but only in the presence of a depressed Ca2+ level in the medium; on inactivation of voltage-gated K+ channels, the contribution of L-type channels to the secretory process became less significant. At a physiological Ca2+ level, the involvement of L-type channels in evoked acetylcholine secretion was apparent, as in mouse synapses, only in conditions of blockade of voltage-gated K+ channels.

Вклад кальциевых каналов L-типа в секрецию ацетилхолина в нервно-мышечных соединениях лягушки и мыши при активных и инактивированных потенциал-зависимых калиевых каналах

In this work, the contribution of L-type Ca2+ channels to the evoked secretion of acetylcholine from the frog and mouse motor nerve endings with active and inactivated potential-dependent K+ channels was studied. The effects of the specific Ca2+ blocker of the L-type channels nitrendipine on the quantal content of endplate currents and the timing of acetylcholine quanta secretion in intact preparations and after the preliminary blockade of potential-activated K+ channels with 4-aminopyridine (4-AP) were evaluated under conditions of a decreased and physiological level of ambient Ca2+. Fluorescent measurement of calcium transient reflecting the integral input of Ca2+ into the nerve ending was performed as well as the computer simulation of the processes underlying exocytosis assuming the presence of two types of Ca2+ channels (N- and L-types) and varying duration of the nerve action potential. It was shown that in the frog synapses, L-type calcium channels contribute to the evoked secretion of acetylcholine at active K+ channels only under conditions of decreased Ca2+ level; after inactivation of potential-dependent K+ channels, the contribution of L-type channels to the secretory process becomes less significant. At the physiological level of ambient Ca2+, the involvement of L-type channels in the evoked acetylcholine secretion, as in mice synapses, is manifested only under conditions of inactivated potential-dependent K+ channels.

Role of presynaptic L-type Ca2+ channels in GABAergic synaptic transmission in cultured hippocampal neurons.

Using dual whole cell patch-clamp recordings of monosynaptic GABAergic inhibitory postsynaptic currents (IPSCs) in cultured rat hippocampal neurons, we have previously demonstrated posttetanic potentiation (PTP) of IPSCs. Tetanic stimulation of the GABAergic neuron leads to accumulation of Ca2+ in the presynaptic terminals. This enhances the probability of GABA-vesicle release for up to 1 min, which underlies PTP. In the present study, we have examined the effect of altering the probability of release on PTP of IPSCs. Baclofen (10 microM), which depresses presynaptic Ca2+ entry through N- and P/Q-type voltage-dependent Ca2+ channels (VDCCs), caused a threefold greater enhancement of PTP than did reducing [Ca2+]o to 1.2 mM, which causes a nonspecific reduction in Ca2+ entry. This finding prompted us to investigate whether presynaptic L-type VDCCs contribute to the Ca2+ accumulation in the boutons during spike activity. The L-type VDCC antagonist, nifedipine (10 microM), had no effect on single IPSCs evoked at 0.2 Hz but reduced the PTP evoked by a train of 40 Hz for 2 s by 60%. Another L-type VDCC antagonist, isradipine (5 microM), similarly inhibited PTP by 65%. Both L-type VDCC blockers also depressed IPSCs during the stimulation (i.e., they increased tetanic depression). The L-type VDCC "agonist" (-)BayK 8644 (4 microM) had no effect on PTP evoked by a train of 40 Hz for 2 s, which probably saturated the PTP process, but enhanced PTP evoked by a train of 1 s by 91%. In conclusion, the results indicate that L-type VDCCs do not participate in low-frequency synchronous transmitter release, but contribute to presynaptic Ca2+ accumulation during high-frequency activity. This helps maintain vesicle release during tetanic stimulation and also enhances the probability of transmitter release during the posttetanic period, which is manifest as PTP. Involvement of L-type channels in these processes represents a novel presynaptic regulatory mechanism at fast CNS synapses.

Modulation of nicotine-evoked [3H]dopamine release from rat striatal synaptosomes by voltage-sensitive calcium channel ligands

The calcium channel subtypes mediating nicotine-evoked [3H]dopamine release from rat striatal synaptosomes were probed with L-, N-, and P-type calcium channel ligands. Responses to nicotine were blocked by the peptides omega-conotoxin GVIA and omega-agatoxin IVA. The affinity constants for these compounds were consistent with their actions at N- and P-type channels, respectively. Together, these channels mediate at least 90% of the calcium-dependent response to nicotine. The L-type antagonists nifedipine, verapamil, and nicardipine were also effective blockers of nicotine-evoked release with maximal effects of 80-100% inhibition. However, these effects occurred at concentrations 2-3 orders of magnitude higher than those necessary to block L-type channels. Moreover, Bay K8644, an L-type agonist, also blocked nicotine-evoked release. Together, these findings argue strongly against the involvement of L-type channels.

Kappa-opioid receptors on astrocytes stimulate l-type Ca 2+ channels

Cultured astrocytes from the cerebral cortex of the rat respond to opioid κ-receptor stimulation with a substantial elevation of the cytoplasmic free calcium, visualized through the use of the fluorescent calcium indicator Fura-2. The stimulation of κ-receptors with U-50488H increases the level of calcium through a dose-related stimulatory effect on the transmembrane calcium influx. The κ-receptor stimulation was completely blocked by the selective κ-receptor blocker nor-binaltorphimine. Furthermore, the transmembrane calcium influx was completely blocked by nifedipine, indicating the involvement of l-type channels. The presence of l-type channels was verified by stimulation of l-type channels with Bay K8644. The effects of Bay K8644 were completely blocked by nifedipine. l-type channel-coupled κ-receptors on astrocytes might represent a novel mechanism contributing to the depressant action of opioids on synaptic transmission via decreasing the availability of extracellular calcium necessary for presynaptic transmitter release.