Abstract
Defolliculated oocytes of Xenopus laevis responded to removal of external divalent cations with large depolarizations and, when voltage clamped, with huge currents. Single channel analysis revealed a Cl- channel with a slope conductance of about 90 pS at positive membrane potentials with at least four substates. Single channel amplitudes and mean channel currents had a reversal potential of approximately -15 mV as predicted by the Nernst equation for a channel perfectly selective for Cl-. Readdition of Ca2+ immediately inactivated the channel and restored the former membrane potential or clamp current. The inward currents were mediated by a Ca2+ inactivated Cl- channel (CaIC). The inhibitory potency of Ca2+ was a function of the external Ca2+ concentration with a half maximal blocker concentration of about 20 microM. These channels were inhibited by the Cl- channel blockers flufenamic acid, niflumic acid and diphenylamine-2-carboxylate (DPC). In contrast, 4,4'-acetamido-4'-isothiocyanatostilbene-2, 2'-disulfonicacid (SITS), another Cl- channel blocker, led to activation of this Cl- channel. Like other Cl- channels, the CaIC was activated by cytosolic cAMP. Extracellular ATP inhibited the channel while ADP was without any effect. Injection of phorbol 12-myristate 13-acetate (PMA), a protein kinase C activating phorbol ester, stimulated the Cl- current. Cytochalasin D, an actin filament disrupting compound, reversibly decreased the clamp current demonstrating an influence of the cytoskeleton. The results indicate that removal of divalent cations activates Cl- channels in Xenopus oocytes which share several features with Cl- channels of the CLC family. The former so-called leak current of oocytes under divalent cation-free conditions is nothing else than an activation of Cl- channels.
MeSH Terms
1-Methyl-3-isobutylxanthine/pharmacology
4-Acetamido-4'-isothiocyanatostilbene-2,2'-disulfonic Acid/pharmacology
Adenosine Triphosphate/pharmacology
Animals
Calcium/pharmacology
Cell Membrane Permeability/drug effects,physiology
Chloride Channels/drug effects,physiology
Cholera Toxin/pharmacology
Cyclic AMP/physiology
Cytochalasin D/pharmacology
Cytoskeleton/drug effects,physiology
Flufenamic Acid/pharmacology
Membrane Potentials/drug effects
Oocytes/drug effects,physiology
Oogenesis
Patch-Clamp Techniques
Pertussis Toxin
Tetradecanoylphorbol Acetate/pharmacology
Virulence Factors, Bordetella/pharmacology
Xenopus laevis
ortho-Aminobenzoates/pharmacology
Chemicals
Chloride Channels
Virulence Factors, Bordetella
ortho-Aminobenzoates
Cytochalasin D
4-Acetamido-4'-isothiocyanatostilbene-2,2'-disulfonic Acid
Flufenamic Acid
Adenosine Triphosphate
Cholera Toxin
fenamic acid
Cyclic AMP
Pertussis Toxin
Tetradecanoylphorbol Acetate
Calcium
1-Methyl-3-isobutylxanthine
Authors & Affiliations
4 authors, click to expand affiliations / ORCID
Weber W M
Institut für Tierphysiologie, Jusois-Liebig-Universităt, Giessen, FRG.
Liebold K M
Reifarth F W
Clauss W
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