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Effects of Ca2+ channel blockers on transmitter release and presynaptic currents at the frog neuromuscular junction
E Katz1, P A Ferro, B D Cherksey
1Instituto de Biología Celular, Facultad de Medicina, Universidad de Buenos Aires, Argentina.
Abstract:
1. The effects of the calcium channel blockers, funnel-web spider toxin (FTX), omega-agatoxin IVA (omega-Aga IVA) and omega-conotoxin GVIA (omega-CgTX), were tested on transmitter release and presynaptic currents in frog motor nerve endings. 2. Evoked transmitter release was blocked by FTX (IC50 = 0.02 microliter ml-1) and omega-CgTX (1 microM) but was not affected by omega-Aga IVA (0.5 microM). When FTX (0.1 microliter ml-1) was assayed on spontaneous release either in normal Ringer solution or in low Ca(2+)-high Mg2+ solution, it was found not to affect miniature endplate potential (MEPP) amplitude but to increase MEPP frequency by approximately 2-fold in both conditions. 3. Presynaptic calcium currents (ICa), measured by the perineurial technique in the presence of 10 mM tetraethylammonium chloride (TEA) and 200 microM BaCl2 to block K+ currents, were blocked by omega-CgTX (5 microM), partially blocked by FTX (1 microliter ml-1) and not affected by omega-Aga IVA (0.5 microM). 4. The presynaptic calcium-activated potassium current (IK(Ca)) measured by the perineurial technique in the presence of 0.5 microM 3,4-aminopyridine (DAP) to block voltage-dependent K+ currents, was strongly affected by charybdotoxin (ChTX) (300 nM) and completely abolished by BaCl2 (200 microM). This current was also blocked by omega-CgTX (5 microM) and by CdCl2 (200 microM) but was not affected by FTX (1 microliter ml-1). The blockade by omega-CgTX could not be reversed by elevating [Ca]o to 10 mM. 5. The results suggest that in frog synaptic terminals two omega-CgTX-sensitive populations might coexist. The transmitter release process seems to be mediated by calcium influx through a omega-CgTX- and FTX-sensitive population.
Insights
This study investigated calcium channel blockers, funnel-web spider toxin (FTX) and omega-conotoxin GVIA (omega-CgTX), on frog motor nerve endings. Results show these toxins affect transmitter release and presynaptic currents, suggesting specific calcium channels mediate neurotransmission.
Area of Science:
- Neuroscience
- Pharmacology
- Molecular Biology
Background:
- Calcium channels play a crucial role in neurotransmitter release at nerve terminals.
- Specific toxins can selectively block different types of calcium channels, providing tools to study their function.
Purpose of the Study:
- To investigate the effects of funnel-web spider toxin (FTX) and omega-conotoxin GVIA (omega-CgTX) on transmitter release and presynaptic currents in frog motor nerve endings.
- To identify the specific calcium channel populations involved in neurotransmission.
Main Methods:
- Electrophysiological recordings of evoked transmitter release and spontaneous miniature endplate potentials (MEPPs).
- Perineurial voltage clamp technique to measure presynaptic calcium currents (ICa) and calcium-activated potassium currents (IK(Ca)).
- Application of specific calcium channel blockers: FTX, omega-CgTX, and omega-agatoxin IVA (omega-Aga IVA).
Main Results:
- FTX and omega-CgTX blocked evoked transmitter release, while omega-Aga IVA did not.
- FTX increased MEPP frequency without affecting MEPP amplitude.
- Omega-CgTX and partially FTX blocked presynaptic ICa, whereas omega-Aga IVA had no effect.
- Omega-CgTX and cadmium chloride (CdCl2) blocked IK(Ca), but FTX did not.
Conclusions:
- Frog motor nerve endings possess at least two omega-CgTX-sensitive calcium channel populations.
- Transmitter release is mediated by calcium influx through omega-CgTX- and FTX-sensitive channels.