Ca2+ spike initiation from sensitized inositol 1,4,5-trisphosphate-sensitive Ca2+ stores in megakaryocytes.
Ca(2+)-mediated Ca2+ spikes were analysed in fura-2-loaded megakaryocytes. Direct Ca2+ loading using whole-cell dialysis induced an all-or-none Ca2+ spike on top of a tonic increase in cellular Ca2+ concentration ([Ca2+]i) with a latency of 3-7 s. The latency decreased with increasingly higher concentrations of Ca2+ in the dialysing solution. Spike size and its initiation did not correlate with the tonic level of [Ca2+]i. Thapsigargin completely abolished the Ca(2+)-induced spike initiation, suggesting that Ca2+ spikes originate from thapsigargin-sensitive Ca2+ pools. An inhibitor of phosphatidylinositide-specific phospholipase C (PLC), 2-nitro-4-carboxyphenyl-N,N-diphenyl-carbamate prolonged the latency without changes of spike size in most cases (6/9 cells), but abolished the spike initiation in the other cells (3/9). The results suggest that an increase in [Ca2+]i charges up the inositol-1,4,5-trisphosphate-(InsP3)- and thapsigargin-sensitive Ca2+ pools which progressively sensitize to low or slightly elevated levels of InsP3 by the action of Ca(2+)-dependent PLC until a critical Ca2+ content is reached, and then the Ca2+ spike is triggered. Thus, the limiting step of Ca2+ spike triggering is the initial filling process and the level of InsP3 in megakaryocytes.
Ikeda
M
M
Department of Physiology, Jichi Medical School, Tochigi, Japan.
Kurokawa
K
K
Maruyama
Y
Y
D000818:Animals; D001854:Bone Marrow Cells; D002110:Caffeine / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D002219:Carbamates / Q000494:pharmacology; D015544:Inositol 1,4,5-Trisphosphate / Q000502:physiology*; D008297:Male; D008533:Megakaryocytes / Q000187:drug effects / Q000378:metabolism*; D015591:Microspectrophotometry; D048448:Phenylcarbamates; D011343:Procaine / Q000494:pharmacology; D051381:Rats; D017208:Rats, Wistar; D013050:Spectrometry, Fluorescence; D013729:Terpenes / Q000494:pharmacology; D019284:Thapsigargin; D013917:Thrombin / Q000502:physiology; D010738:Type C Phospholipases / Q000037:antagonists & inhibitors
D002219:Carbamates; D048448:Phenylcarbamates; D013729:Terpenes; C025508:2-nitro-4-carboxyphenyl-N,N-diphenylcarbamate; D002110:Caffeine; D011343:Procaine; D019284:Thapsigargin; D015544:Inositol 1,4,5-Trisphosphate; D010738:Type C Phospholipases; D013917:Thrombin; D002118:Calcium