Modulation of Xenopus laevis Ca-activated Cl currents by protein kinase C and protein phosphatases: Implications for studies of anesthetic mechanisms

Ca-activated Cl currents (ICl(Ca)) are used frequently as reporters in functional studies of anesthetic effects on G protein-coupled receptors using Xenopus laevis oocytes. However, because anesthetics affect protein kinase C (PKC), they could indirectly affect ICl(Ca) if this current is regulated by phosphorylation. We therefore studied the effect of modulation of either PKC or protein phosphatases PP1 and PP2A on ICl(Ca) stimulated either by lysophosphatidate (LPA) signaling or by microinjection of Ca. X. laevis oocytes were studied under voltage clamp. Rat PP1 and PP2A were overexpressed in oocytes. PP, inositoltrisphosphate (IP3), the PP inhibitor okadaic acid (OA), the PKC inhibitor chelerythrine, or CaCl2 were directly injected into the oocyte. Responses to agonists (LPA 10 6 M, IP3 10 4 M, CaCl2 0.5 M) were measured at a holding potential of 70 mV in the presence or absence of the PP inhibitors cantharidin or OA. PP1 and PP2A inhibited ICl(Ca) from 7.6 0.9 Ct o 2.5 0.9 C and 3.2 1.4 C, respectively. PP inhibition enhanced ICl(Ca) in control oocytes and reversed the inhibitory effect in oocytes expressing PP1 or PP2A. PKC inhibition by chelerythrine enhanced both LPAand CaCl2-induced ICl(Ca). Our data indicate that the Xenopus ICl(Ca) is modulated by phosphorylation. This may complicate design and interpretation of studies of G protein-coupled receptors using this model. (Anesth Analg 2004;99:416 –22)