EFFECTS OF PH ON APICAL CALCIUM-ENTRY AND ACTIVE CALCIUM-TRANSPORT IN RABBIT CORTICAL COLLECTING SYSTEM

Rabbit connecting tubules and cortical collecting ducts were isolated by immunodissection and cultured on permeable supports. The monolayers actively transported Ca2+ with a net transcellular rate of 92 +/- 3 nmol.h(-1).cm(-2). Methoxyverapamil, felodipine, diltiazem, omega-conotoxin GVIA, and omega-agatoxin IVA when added to the apical side had no effect on Ca2+ absorption. Neither hyperpolarization nor depolarization of the apical membrane affected Ca2+ transport rates significantly. Stepwise lowering of the apical pH (pH(a)) from 8.0 to 5.6 gradually inhibited Ca2+ transport from 88 +/- 5 to 7 +/- 2 nmol.h(-1).cm(-2). Measuring the intracellular pH (pH(i)) with 2',7'-bis(carboxyethyl)-5(6)-carboxyfluorescein revealed that lowering the pH(a) from 8.0 to 5.6 decreased pH(i) from 7.8 to 6.7. To determine whether inhibition of Ca2+ absorption results from intracellular acidification, pH(i) was lowered using an NH4Cl pulse while extracellular pH was kept constant. Intracellular acidification from 7.4 +/- 0.2 to 6.9 +/- 0.1 reduced Ca2+ absorption by 26 +/- 6% only. In addition, lowering of the basolateral pH to 6.2 resulted in a pH(i) of 6.8 +/- 0.1, without affecting Ca2+ absorption rates. In conclusion, the basal Ca2+ influx mechanism in the apical membrane is most likely a voltage-independent Ca2+ transporter, insensitive to Ca2+ channel blockers, but strongly inhibited by apical acidification.