The mechanisms of sarcoplasmic reticulum Ca2+ release in toad pacemaker cells.
The mechanisms of sarcoplasmic reticulum (SR) Ca2+ release in pacemaker cells from the sinus venosus of the cane toad (Bufo marinus) were studied. Single, isolated cells were voltage clamped using a nystatin-perforated patch. Ionic currents and intracellular Ca2+ concentration ([Ca2+]i) were recorded simultaneously. Depolarizations of 300 ms duration from a holding potential of -55 mV produced an inward current which had a bell-shaped relationship with voltage. Inward current first appeared at about -45 mV, reached a maximum of -343 +/- 46 pA at -15 mV and reversed at +45 mV. In contrast the amplitude of the increase in [Ca2+]i caused by depolarization (Ca2+ transient) increased monotonically with the increasing depolarization. At -15 mV the amplitude of the Ca2+ transient was 243 +/- 33 nM and at +45 mV it was 411 +/- 43 nM. The inward current produced by depolarizations to -5 mV was largely eliminated by the L-type Ca2+ channel blocker nifedipine (10 microM) while 37 +/- 7 % of the Ca2+ transient persisted. A significantly larger proportion of the Ca2+ transient (56 +/- 5 %) remained at +85 mV in the presence of nifedipine. The SR Ca2+ pump inhibitor 2, 5-di(tert-butyl)-1,4-hydroquinone (10 microM), which causes depletion of the SR Ca2+, reduced the amplitude of the Ca2+ transient to 34 +/- 1 % of control, irrespective of the voltage. Brief exposure to extracellular Ca2+-free solution abolished the Ca2+ transients caused by depolarization while the caffeine-induced Ca2+ release persisted. Tetrodotoxin (1 microM) had no effect on the amplitude of the depolarization-induced Ca2+ transient, although it reduced the fast component of the inward current. In contrast, Ni2+ (5 mM) abolished the Ca2+ transients at any given voltage. Ni2+ also abolished spontaneous Ca2+ transients. In conclusion, in toad pacemaker cells Ca2+ release from SR contributes approximately 66 % of the Ca2+ involved in the Ca2+ transient and requires extracellular Ca2+ influx to trigger its release. The L-type Ca2+ channels and Na+-Ca2+ exchange are major sources of Ca2+ influx under physiological conditions.
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Department of Physiology and The Institute for Biomedical Research, University of Sydney (F13), Sydney, NSW 2006, Australia.
Allen
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D000200:Action Potentials / Q000502:physiology; D000818:Animals; D016227:Benzoquinones / Q000494:pharmacology; D001683:Biological Clocks / Q000502:physiology*; D002024:Bufo marinus; D002118:Calcium / Q000493:pharmacokinetics*; D002121:Calcium Channel Blockers / Q000494:pharmacology; D020746:Calcium Channels, L-Type / Q000378:metabolism; D000252:Calcium-Transporting ATPases / Q000037:antagonists & inhibitors / Q000378:metabolism; D066298:In Vitro Techniques; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology; D009206:Myocardium / Q000737:chemistry / Q000166:cytology / Q000378:metabolism*; D009532:Nickel / Q000494:pharmacology; D009543:Nifedipine / Q000494:pharmacology; D018408:Patch-Clamp Techniques; D012519:Sarcoplasmic Reticulum / Q000201:enzymology*; D012964:Sodium / Q000378:metabolism; D019831:Sodium-Calcium Exchanger / Q000378:metabolism; D013779:Tetrodotoxin / Q000494:pharmacology
D016227:Benzoquinones; D002121:Calcium Channel Blockers; D020746:Calcium Channels, L-Type; D019831:Sodium-Calcium Exchanger; C067344:2,5-di-tert-butylbenzoquinone; D013779:Tetrodotoxin; D009532:Nickel; D012964:Sodium; D000252:Calcium-Transporting ATPases; D009543:Nifedipine; D002118:Calcium