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© The Rockefeller University Press, 0022-1295/1997//703/ $5.00
Journal of General Physiology, Volume 109, Number 6, 1997

Article

Calcium Current Activated by Depletion of Calcium Stores in Xenopus Oocytes

Yong Yao* and Roger Y. Tsien*,{ddagger}

From the * Department of Pharmacology and {ddagger} Howard Hughes Medical Institute, University of California, San Diego, La Jolla, California 92093-0647

Ca2+ currents activated by depletion of Ca2+ stores in Xenopus oocytes were studied with a two-electrode voltage clamp. Buffering of cytosolic Ca2+ with EGTA and MeBAPTA abolished ICl(Ca) and unmasked a current in oocytes that was activated by InsP3 or ionomycin in minutes and by thapsigargin or the chelators themselves over hours. At –60 mV in 10 mM extracellular CaCl2, the current was typically around –90 or –160 nA in oocytes loaded with EGTA or MeBAPTA, respectively. This current was judged to be a Ca2+-selective current for the following reasons: (a) it was inwardly rectifying and reversed at membrane potentials usually more positive than +40 mV; (b) it was dependent on extracellular [CaCl2] with Km = 11.5 mM; (c) it was highly selective for Ca2+ against monovalent cations Na+ and K+, because replacing Na+ and K+ by N-methyl-D-glucammonium did not reduce the amplitude or voltage dependence of the current significantly; and (d) Ca2+, Sr2+, and Ba2+ currents had similar instantaneous conductances, but Sr2+ and Ba2+ currents appeared to inactivate more strongly than Ca2+. This Ca2+ current was blocked by metal ions with the following potency sequence: Mg2+ << Ni2+ {approx} Co2+ {approx} Mn2+ < Cd2+ << Zn2+ << La3+. It was also inhibited by niflumic acid, which is commonly used to block ICl(Ca). PMA partially inhibited the Ca2+ current, and this effect was mostly abolished by calphostin C, indicating that the Ca2+ current is sensitive to protein kinase C. These results are the first detailed electrophysiological characterization of depletion-activated Ca2+ current in nondialyzed cells. Because exogenous molecules and channels are easy to introduce into oocytes and the distortions in measuring ICl(Ca) can now be bypassed, oocytes are now a superior system in which to analyze the activation mechanisms of capacitative Ca2+ influx.

Key Words: calcium influx • ICRAC • ISOC • thapsigargin • MeBAPTA

Address correspondence to Dr. Roger Y. Tsien, Department of Pharmacology/HHMI, CMM-West, Room 310, University of California San Diego, La Jolla, CA 92093-0647. Fax: 619-534-5270. E-mail: rtsien{at}ucsd.edu


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