Thapsigargin inhibits Ca2+ uptake, and Ca2+ depletes sarcoplasmic reticulum in intact cardiac myocytes.
We examined the effects of thapsigargin on Ca2+ accumulation by the sarcoplasmic reticulum (SR) and on electrically stimulated beats in single adult rat ventricular myocytes loaded with indo 1 and bathed in N-2-hydroxyethylpiperazine-N'-2-ethanesulfonic acid buffer containing 1 mM Ca2+ at 23 degrees C. The SR Ca2+ content was assessed from the magnitude of intracellular Ca2+ (Ca2+i) transients and contractions elicited by rapid, brief applications of caffeine. After 20-30 min of exposure to 200 nM thapsigargin, the caffeine-dependent Ca2+i transients were abolished or markedly diminished (by 89 +/- 4%). The postrest potentiation of the Ca2+i transient and contraction, typical for rat myocardium, was abolished. Thapsigargin did not significantly change resting Ca2+i but diminished the amplitude of the steady-state Ca2+i transients by 73%, prolonged the time to peak by 24%, and prolonged the half-time (t1/2) of the Ca2+i transient decline by 42%. Progressive SR Ca2+ depletion by thapsigargin was strongly related (r = -0.78) to the prolongation of the t1/2 of relaxation of the steady-state Ca2+i transients, suggesting that the thapsigargin-dependent SR Ca2+ depletion results from an inhibition of the SR Ca2+ uptake. This interpretation was corroborated by comparison of the effects of thapsigargin with those of ryanodine (100 nM), which depletes SR of Ca2+ by accelerating the SR Ca2+ efflux but does not inhibit the SR Ca2+ pump. During rapid pacing (5 Hz), which raises Ca2+i and thus Ca2+ available for SR uptake, the caffeine-dependent SR Ca2+ release was restored in ryanodine-treated cells but not in the presence of thapsigargin.(ABSTRACT TRUNCATED AT 250 WORDS)
Janczewski
A M
AM
Laboratory of Cardiovascular Science, National Institute on Aging, National Institutes of Health, Baltimore, Maryland 21224.
Lakatta
E G
EG
D000818:Animals; D002110:Caffeine / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism / Q000494:pharmacology* / Q000502:physiology*; D002121:Calcium Channel Blockers / Q000494:pharmacology*; D004558:Electric Stimulation; D004594:Electrophysiology; D006321:Heart / Q000187:drug effects; D009200:Myocardial Contraction; D009206:Myocardium / Q000166:cytology*; D012433:Ryanodine / Q000494:pharmacology; D012519:Sarcoplasmic Reticulum / Q000187:drug effects* / Q000378:metabolism; D013729:Terpenes / Q000494:pharmacology*; D019284:Thapsigargin
D002121:Calcium Channel Blockers; D013729:Terpenes; D012433:Ryanodine; D002110:Caffeine; D019284:Thapsigargin; D002118:Calcium