Intracellular pH (pH_i) and its basolateral regulation were studied in isolated proximal-proximal and distal-proximal segments of garter snake (Thamnophis spp.) renal tubules with oil-filled lumens in HEPES-buffered and in HEPES-HCO₃-buffered media (pH 7.4 at 25°C). pH_i was measured with the pH-sensitive fluorescent dye 2',7'-bis(2-carboxyethyl)-5,6-carboxyfluorescein (BCECF) under resting conditions and in response to NH₄Cl pulse. Resting pH_i (~7.1-7.2) and its response to and rate of recovery (dpH_i/dt) from an NH₄Cl pulse were not affected by the presence or absence of HCO₃ in either segment. Rate of recovery was depressed by Na⁺ removal in distal-proximal segments only and only in HEPES buffer. It was not affected by removal of Cl or of both Na⁺ and Cl or by reduction in membrane potential through addition of Ba²⁺ (5 mM) or high K⁺ (75 mM) in either segment in either HEPES or HEPES-HCO₃ buffer. The Na⁺/H⁺ exchange inhibitor ethylisopropylamiloride (EIPA) (100 µM) and the anion exchange inhibitor DIDS (100 µM) reduced dpH_i/dt in the distal-proximal segments only and only in HEPES-HCO₃ buffer. The H⁺-ATPase inhibitor bafilomycin (1 µM), H⁺-K⁺-ATPase and K⁺/NH⁺₄ exchange inhibitor Schering 28080 (10-100 µM), organic cation efflux inhibitor tetrapentylammonium (25 µM-20 mM), and K⁺ channel blocker tetraethylammonium (20 mM) had no effect on dpH_i/dt in either segment. These data do not clearly support basolateral regulation of pH_i in snake proximal renal tubules by commonly recognized Na⁺-dependent or Na⁺-independent acid or base transporters.

Thamnophis spp.; garter snakes; buffers; ammonium chloride pulse; bafilomycin; Schering 28080