Pharmacology of a capacitative Ca2+ entry in Xenopus oocytes.
We have characterized pharmacological properties of inositol trisphosphate (InsP3)-mediated calcium entry pathway in Xenopus oocytes via activation of Ca(2+)-dependent Cl- channels (ICl, Ca) as a sensitive indicator for increase in cytosolic [Ca2+]. This type of Ca2+ entry mechanism is known as a capacitative Ca2+ entry (CCE). Voltage-clamped oocytes were maintained in Ca(2+)-free medium and injected with InsP3 which depleted the InsP3-sensitive Ca2+ stores. 10-20 min later, the oocytes were exposed, at 2-3 min intervals, to 5 mM Ca(2+)-containing medium for 5-10 s which evoked repeated inward Cl- current. No effect of external Ca2+ was apparent before InsP3 injection. To determine the pharmacological characteristics of CCE, oocytes were incubated with various chemical agents in Ca(2+)-free solution and exposed to Ca2+ again in presence of the chemical. It was found that organic Ca2+ channel blockers were relatively ineffective in blocking CCE while the inorganic Ca2+ channel blocker La3+ was most efficient in blocking the current. Attempts to measure conductance increase when the Cl- channels were blocked during activation of Ca2+ influx were unsuccessful. Therefore we tested the hypothesis that the Ca2+ influx is mediated via a Ca-H transporter. Lowering the external pH (to pH 6.5) or application of the protonophore carbonylcyanide p-trifluoromethoxyphenyl hydrazone (EC50 = 2 x 10(-8) M) effectively blocked CCE. Since Ca-H countertransport in the plasma membrane is coupled to Ca2+ extrusion by Ca-ATPase in vascular smooth muscle we suggest that the capacitative Ca2+ entry in Xenopus oocytes may possibly arise from slippage of plasma membrane Ca-ATPase coupled to proton countertransport, a mechanism reported in a variety of cells. Ca2+ slippage may arise from the large Ca2+ gradient produced by the Ca2+ depletion protocol.
Gillo
B
B
Department of Physiology, Hebrew University-Hadassah Medical School, Jerusalem, Israel.
Sealfon
S C
SC
Minke
B
B
D000818:Animals; D002118:Calcium / Q000493:pharmacokinetics*; D005260:Female; D009865:Oocytes / Q000378:metabolism*; D014981:Xenopus / Q000378:metabolism*
D002118:Calcium