CHLORIDE DEPENDENCE OF PH MODULATION BY BETA-ADRENERGIC AGONIST IN RAT CARDIOMYOCYTES

The effects of beta-adrenergic agonists on pH(i) were studied on singl e ventricular myocytes isolated from adult rat heart and loaded with t he acetoxymethyl ester (AM) form of the pH indicator SNARF-1. In modif ied Krebs' solution containing 20 mmol/L HEPES and 4.4 mmol/L HCO3-, i soproterenol (1 mu mol/L) caused a significant decrease of steady-stat e pH(i) from 7.20 +/- 0.02 to 7.13 +/- 0.02 (mean +/- SEM) within 2 mi nutes. This acidification, which was also observed in myocytes that we re preloaded with the Ca2+ chelator BAPTA and superfused with nominall y Ca2+-free solution, was blocked by propranolol as well as by the spe cific beta(1)-antagonist CGP 20712 A but not by the beta(2)-antagonist ICI 118,551. Forskolin (10 mu mol/L) induced a similar reversible dec rease of pH(i) (average decrease, 0.11 +/- 0.02 pH unit). Furthermore, adenosine (100 mu mol/L) substantially attenuated the isoproterenol-i nduced decrease of pH(i). The effect of isoproterenol was not prevente d by inhibitors of the Na+-H+ antiport, amiloride (1 mmol/L) and 2-N,N -hexamethylene amiloride (20 mu mol/L). On the other hand, blockers of Cl- transport mechanisms, DIDS (200 mu mol/L) and probenecid (100 mu mol/L), inhibited this acidification. Isoproterenol also failed to ind uce a decrease of steady-state pH(i) in myocytes incubated in Cl--free medium. Rather, the initial rate of rise of pH(i) observed on removal of external Cl- ions was significantly increased in the presence of i soproterenol or dibutyryl cAMP. Because the alkalinization induced by removal of Cl- ions is mainly due to reversal of the Cl--HCO3- exchang er, the augmentation of this initial rate of pH(i) rise directly point s to a beta-adrenergic stimulation of the exchanger. Furthermore, the pH(i) recovery following NH4Cl exposure was accelerated by isoproteren ol in the presence of probenecid, indicating that the Na+-HCO3- cotran sport and/or the Na+-H+ antiport also could be activated. In conclusio n, the present results demonstrate that beta-adrenergic agonist-induce d acidification of rat ventricular myocytes occurs mainly through alte ration of Cl- transport systems, most likely via a cAMP-dependent stim ulation of the Cl--HCO3-exchanger. Since the alkalinizing mechanisms a re also stimulated, an increased apparent cellular buffering capacity is expected.