Pharmacology of a capacitative Ca2+ entry in Xenopus oocyte

We have characterized pharmacological properties of inositol trisphosphate (InsP3)-mediated calcium entry pathway in Xenopus oocytes via activation of Ca2+-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 Ca2+-free medium and injected with InsP3which depleted the InsP3-sensitive Ca2+ stores. 10–20 min later, the oocytes were exposed, at 2–3 min intervals, to 5 mM Ca2+-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 Ca2+-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 × 10−8M) 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.

Authors: Gillo, B., Sealfon, S.C. and Minke, B
Year of publication: 1996
Journal: Journal of Photochemistry and Photobiology B: Biology, Volume 35, Issues 1–2, Pages 77-82

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