PHOSPHATASE-MEDIATED ENHANCEMENT OF CARDIAC CAMP-ACTIVATED CL- CONDUCTANCE BY A CL- CHANNEL BLOCKER, ANTHRACENE-9-CARBOXYLATE

An aromatic carboxylate, anthracene-9-carboxylic acid (9-AC), is known as a Cl- channel blocker. However, variable 9-AC effects have hithert o been reported on the cardiac cAMP-activated Cl- conductance, when ap plied extracellularly. We have reexamined the 9-AC effect on the Cl- c onductance activated by isoproterenol or forskolin in guinea pig ventr icular myocytes under whole-cell patch-clamp conditions. The inward cu rrent was blocked by 9-AC at greater than or equal to 0.5 mmol/L, but in contrast, the outward current was enhanced at much lower concentrat ions (ED50, approximate to 13 mu mol/L). 9-AC applied by the intracell ular perfusion technique increased both the inward and outward current s. In the presence of intracellular 9-AC, deactivation of the conducta nce after washout of isoproterenol or forskolin was largely prevented. 9-AC produced an enhancing effect, even after inhibiting the deactiva tion process by okadaic acid (OA), whereas it failed to produce additi onal effects in the presence of orthovanadate. Intracellular applicati on of 9-AC together with OA virtually abolished the current deactivati on The 9-AC effects on the Cl- conductance were not dependent on intra cellular Ca2+ or pH. Putative inhibitors of alkaline (bromotetramisole ) and acid phosphatases (tartrate) were without effect. 9-AC failed to inhibit the activities of purified protein phosphatase (PP)-1, -2A, a nd -2C. In the extract of guinea pig ventricle, 9-AC (greater than or equal to 10 mu mol/L for full action) significantly inhibited a fracti on of endogenous phosphatase activity that was sensitive to orthovanad ate but not to OA, bromotetramisole, and tartrate. It is concluded tha t 9-AC blocks cardiac cAMP-activated (cystic fibrosis transmembrane co nductance regulator) Cl- conductance from the extracellular side but e nhances the conductance from the intracellular side by inhibiting an o rthovanadate-sensitive phosphatase distinct from PP-1, -2A, -2B, or -2 C and alkaline or acid phosphatase.