title: Voltage-dependent Ca2+ release in rat megakaryocytes requires functional IP3 receptors.
abstract: Using simultaneous whole-cell patch-clamp and fluorescence measurements of [Ca2+]i in rat megakaryocytes we have investigated the requirement for functional inositol 1,4,5-trisphosphate (IP3) receptors in Ca2+ release induced by membrane depolarization during agonist stimulation. Voltage-dependent Ca2+ release was observed during application of the IP3-generating agonists U46619 (a thromboxane A2 analogue) and ADP. Furthermore, voltage-dependent Ca2+ release was observed in the absence of exogenous agonist following sensitization of IP3 receptors with thimerosal. Depolarization-induced Ca2+ release was not detected during depletion of intracellular Ca2+ stores by thapsigargin. Thus, depletion of stores alone is not sufficient to confer voltage dependence upon the Ca2+ release mechanism. Block of IP3 receptors by carbacyclin-stimulated elevations in cAMP, uncaging of cAMP or exposure to a high concentration of caffeine reversibly abolished Ca2+ increases stimulated by both ADP and depolarization. The cAMP-dependent block was prevented by a peptide inhibitor of protein kinase A, indicating that an alteration of adenylate cyclase activity leading to modulation of protein kinase A activity does not underlie the control of Ca2+ release by voltage. These results are consistent with the requirement for functional IP3 receptors for voltage control of Ca2+ release from intracellular stores during inositol lipid signalling. The data also indicate the involvement of a voltage sensor downstream of surface membrane receptors in the depolarization-evoked Ca2+ response.
authors.affiliation: Department of Physiology, University of Cambridge, Cambridge CB2 3EG, UK. mjm39@cam.ac.uk
authors.forename: M J
authors.initials: MJ
authors.lastname: Mason
authors.forename: M P
authors.initials: MP
authors.lastname: Mahaut-Smith
mesh_terms: D019796:15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5,13-dienoic Acid / Q000494:pharmacology; D000244:Adenosine Diphosphate / Q000494:pharmacology; D000818:Animals; D000891:Anti-Infective Agents, Local / Q000494:pharmacology; D002110:Caffeine / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D015220:Calcium Channels / Q000378:metabolism*; D020013:Calcium Signaling / Q000187:drug effects / Q000502:physiology*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D011464:Epoprostenol / Q000031:analogs & derivatives / Q000494:pharmacology; D053496:Inositol 1,4,5-Trisphosphate Receptors; D008297:Male; D008533:Megakaryocytes / Q000378:metabolism*; D018408:Patch-Clamp Techniques; D010726:Phosphodiesterase Inhibitors / Q000494:pharmacology; D010975:Platelet Aggregation Inhibitors / Q000494:pharmacology; D051381:Rats; D017208:Rats, Wistar; D018160:Receptors, Cytoplasmic and Nuclear / Q000378:metabolism*; D019284:Thapsigargin / Q000494:pharmacology; D013849:Thimerosal / Q000494:pharmacology; D014662:Vasoconstrictor Agents / Q000494:pharmacology
chemical_list: D000891:Anti-Infective Agents, Local; D015220:Calcium Channels; D004791:Enzyme Inhibitors; D053496:Inositol 1,4,5-Trisphosphate Receptors; D010726:Phosphodiesterase Inhibitors; D010975:Platelet Aggregation Inhibitors; D018160:Receptors, Cytoplasmic and Nuclear; D014662:Vasoconstrictor Agents; D013849:Thimerosal; D002110:Caffeine; D000244:Adenosine Diphosphate; D019284:Thapsigargin; C026678:carboprostacyclin; D019796:15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5,13-dienoic Acid; D011464:Epoprostenol; D002118:Calcium