The modulation of action potential generation by calcium-induced calcium release is enhanced by mitochondrial inhibitors in mudpuppy parasympathetic neurons.
Previously, we demonstrated that outward currents activated by calcium-induced calcium release (CICR) opposed depolarization-induced action potential (AP) generation in dissociated mudpuppy parasympathetic neurons [J Neurophysiol 88 (2002) 1119]. In the present study, we tested whether AP generation by depolarizing current ramps could be altered by dissipating the mitochondrial membrane potential and thus interrupting mitochondrial Ca2+ buffering. Exposure to the protonophore carbonyl cyanide m-chlorophenylhydrazone (CCCP; 2 microM) alone or in combination with the mitochondrial ATP synthase inhibitor oligomycin (8 microg/ml), increased the latency to AP generation. Exposure to the electron transport chain inhibitor rotenone (10 microM) alone or in combination with oligomycin (8 microg/ml) similarly increased the latency to AP generation. CCCP and oligomycin or rotenone and oligomycin treatment caused rhodamine 123 loss from mitochondria within a few minutes, confirming that the mitochondrial membrane potential was dissipated during drug exposure. Oligomycin alone had no effect on the latency to AP generation and did not cause loss of rhodamine 123 from mitochondria. The increase in latency induced by CCCP and oligomycin was similar when recordings were made with either the perforated patch or standard whole cell patch recording configuration. Exposure to the endoplasmic reticulum Ca-ATPase inhibitor thapsigargin (1 microM), decreased the latency to AP generation. In cells pretreated with thapsigargin to eliminate CICR, CCCP and oligomycin had no effect on AP latency. Pretreatment with iberiotoxin (IBX; 100 nM), an inhibitor of large conductance, calcium- and voltage-activated potassium channels, reduced the extent of the CCCP- and oligomycin-induced increase in latency to AP generation. These results indicate that treatment with CCCP or rotenone to dissipate the mitochondrial membrane potential, a condition which should minimize sequestration of Ca2+ by mitochondria, facilitated the Ca(2+)-induced Ca2+ release activation of IBX-sensitive and IBX-insensitive conductances that regulate AP generation.
Department of Anatomy and Neurobiology, University of Vermont College of Medicine, 89 Beaumont Avenue, Burlington, VT 05405, USA.
K L
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Barstow
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Locknar
L A
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Merriam
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Parsons
D000200:Action Potentials / Q000187:drug effects* / Q000502:physiology; D000814:Aniline Compounds / Q000378:metabolism; D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002258:Carbonyl Cyanide m-Chlorophenyl Hydrazone / Q000494:pharmacology; D002614:Chelating Agents / Q000494:pharmacology; D004347:Drug Interactions; D004533:Egtazic Acid / Q000031:analogs & derivatives* / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005726:Ganglia, Parasympathetic / Q000166:cytology* / Q000187:drug effects; D007462:Iodobenzenes / Q000494:pharmacology; D007476:Ionophores / Q000494:pharmacology; D008564:Membrane Potentials; D008928:Mitochondria / Q000187:drug effects; D009337:Necturus; D009474:Neurons / Q000166:cytology / Q000187:drug effects*; D009840:Oligomycins / Q000494:pharmacology*; D009924:Organ Culture Techniques; D018408:Patch-Clamp Techniques / Q000379:methods; D011930:Reaction Time / Q000187:drug effects; D020112:Rhodamine 123 / Q000378:metabolism; D012402:Rotenone / Q000494:pharmacology; D014475:Uncoupling Agents / Q000494:pharmacology; D014966:Xanthenes / Q000378:metabolism
D000814:Aniline Compounds; D002614:Chelating Agents; D004791:Enzyme Inhibitors; D007462:Iodobenzenes; D007476:Ionophores; D009840:Oligomycins; D014475:Uncoupling Agents; D014966:Xanthenes; D012402:Rotenone; D020112:Rhodamine 123; C059715:Fluo-3; C476101:o-iodoxybenzoic acid; D004533:Egtazic Acid; D002258:Carbonyl Cyanide m-Chlorophenyl Hydrazone; C025603:1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid; D002118:Calcium