Ca2+-dependent capacitance increases in rat basophilic leukemia cells following activation of store-operated Ca2+ entry and dialysis with high-Ca2+-containing intracellular solution.
Ca2+-dependent vesicular fusion was studied in single whole-cell patch-clamped rat basophilic leukemia (RBL) cells using the capacitance technique. Dialysis of the cells with 10 microM free Ca2+ and 300 microM guanosine 5'-O-(3-thiotriphosphate) (GTP[gamma-S]) resulted in prominent capacitance increases. However, dialysis with either Ca2+ (225 nM to 10 microM) or GTP[gamma-S] alone failed to induce a capacitance change. Under conditions of weak Ca2+ buffering (0.1 mM EGTA), activation of Ca2+-release-activated Ca2+ (CRAC) channels by dialysis with inositol 1,4,5-trisphosphate (InsP3) failed to induce a capacitance increase even in the presence of GTP[gamma-S]. However, when Ca2+ATPases were inhibited by thapsigargin, InsP3 and GTP[gamma-S] led to a pronounced elevation in membrane capacitance. This increase was dependent on a rise in intracellular Ca2+ because it was abolished when cells were dialysed with a high level of EGTA (10 mM) in the recording pipette. The increase was also dependent on Ca2+ influx because it was effectively suppressed when external Ca2+ was removed. Our results demonstrate that ICRAC represents an important source of Ca2+ for triggering a secretory response.
Artalejo
A R
AR
Department of Physiology, University of Oxford, Parks Road, Oxford, UK.
Ellory
J C
JC
Parekh
A B
AB
D000818:Animals; D002118:Calcium / Q000378:metabolism* / Q000494:pharmacology*; D015220:Calcium Channels / Q000187:drug effects / Q000502:physiology*; D000252:Calcium-Transporting ATPases / Q000037:antagonists & inhibitors; D002614:Chelating Agents / Q000494:pharmacology; D003956:Dialysis; D004533:Egtazic Acid / Q000494:pharmacology; D004553:Electric Conductivity; D004791:Enzyme Inhibitors / Q000494:pharmacology; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000494:pharmacology; D015544:Inositol 1,4,5-Trisphosphate / Q000494:pharmacology; D015471:Leukemia, Basophilic, Acute / Q000503:physiopathology*; D051381:Rats; D019284:Thapsigargin / Q000494:pharmacology; D014407:Tumor Cells, Cultured
D015220:Calcium Channels; D002614:Chelating Agents; D004791:Enzyme Inhibitors; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D004533:Egtazic Acid; D019284:Thapsigargin; D015544:Inositol 1,4,5-Trisphosphate; D000252:Calcium-Transporting ATPases; D002118:Calcium