| Abstrakt: |
A slowly activating, outwardly rectifying Cl– channel (ORCC) has been described in rat peritoneal mast cells (RPMCs). This channel is activated by intracellular application of cAMP, an effect that might be mediated by a PKA-type serine/threonine protein kinase. To test this hypothesis, whole-cell patch-clamp experiments (nystatin-perforated patch) were performed and 8-bromoadenosine 3′,5′-cyclic monophosphothioate, Sp-enantiomer (Sp-8Br-cAMPS), a cell membrane-permeable activator of PKA, and three inhibitors of different serine/threonine protein phosphatases (okadaic acid, cantharidin, calyculin A), were tested. In RPMCs application of repetitive series of step hyper- and depolarizations (holding potential 0 mV, test potentials –80 to +80 mV, step size +20 mV) induced a slowly increasing, [half-maximal activation time (t0.5) 11.0±1.1 min, Imax (at +80 mV) 18.7±3.1 pA pF–1], DIDS-sensitive, outwardly rectifying Cl– current ICl,OR. The activation of this current could be accelerated by Sp-8Br-cAMPS, okadaic acid or cantharidin in the extracellular solution. Co-application of Sp-8Br-cAMPS and okadaic acid increased Imax supra-additively. Calyculin A and higher concentrations of cantharidin inhibited the Cl– current via unknown mechanisms. Our findings suggest that ICl,OR in RPMCs is activated by a PKA-type protein kinase, a process which is antagonized functionally by okadaic acid- and cantharidin-sensitive protein phosphatases. [ABSTRACT FROM AUTHOR] |
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