Thapsigargin activates a calcium influx pathway in the unfertilized mouse egg and suppresses repetitive calcium transients in the fertilized egg.
At fertilization, the sperm initiates development of the mouse egg by inducing a large transient increase in the intracellular Ca2+ concentration ([Ca2+]i), which is followed by repetitive transient increases in [Ca2+]i. To determine how the repetitive Ca2+ transients are produced, thapsigargin, an inhibitor of the endoplasmic reticulum Ca-ATPase, was used to deplete intracellular Ca2+ stores within the egg. In the unfertilized egg, thapsigargin (1-50 microM) caused a slowly rising and falling transient increase in [Ca2+]i with or without extracellular Ca2+. An influx pathway for Ca2+ is activated by thapsigargin, since an immediate increase in [Ca2+]i occurred when Ca2+ was added to eggs after thapsigargin treatment in a Ca2+, Mg(2+)-free medium. This suggests that Ca2+ entry in the mouse egg may be coupled to the emptying of an intracellular store. The magnitude of the first Ca2+ transient at fertilization was reduced by as much as 84% in eggs pretreated with thapsigargin. Reduction of extracellular Ca2+, by addition of a Ca2+ chelator, suppressed the repetitive Ca2+ transients following fertilization. The Ca2+ transients also require filling of an intracellular store; they were suppressed when thapsigargin was added before or after fertilization. These results support the hypothesis that the first sperm-induced Ca2+ transient at fertilization depletes an intracellular Ca2+ store, triggering an increase in plasma membrane Ca2+ permeability, and that the enhanced Ca2+ influx causes repetitive Ca2+ transients due to the periodic filling and emptying of an intracellular Ca2+ store.
Kline
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Department of Biological Sciences, Kent State University, Ohio 44242.
Kline
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JT
D000251:Adenosine Triphosphatases / Q000037:antagonists & inhibitors; D000814:Aniline Compounds; D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002413:Cations, Divalent; D005306:Fertilization; D005307:Fertilization in Vitro; D005456:Fluorescent Dyes; D016257:Fura-2; D015544:Inositol 1,4,5-Trisphosphate / Q000494:pharmacology*; D007700:Kinetics; D008297:Male; D051379:Mice; D008815:Mice, Inbred Strains; D010063:Ovum / Q000187:drug effects / Q000378:metabolism* / Q000502:physiology; D013050:Spectrometry, Fluorescence; D013094:Spermatozoa / Q000502:physiology; D013729:Terpenes / Q000494:pharmacology*; D019284:Thapsigargin; D014966:Xanthenes
D000814:Aniline Compounds; D002413:Cations, Divalent; D005456:Fluorescent Dyes; D013729:Terpenes; D014966:Xanthenes; C059715:Fluo-3; D019284:Thapsigargin; D015544:Inositol 1,4,5-Trisphosphate; D000251:Adenosine Triphosphatases; D002118:Calcium; D016257:Fura-2