Intracellular in snake renal proximal tubules

Intracellular pH (pH(i)) was studied in isolated proximal renal tubules of garter snakes (Thamnophis spp.) with oil-filled lumens under control conditions [N-2-hydroxyethylpiperazine-N'-2-ethanesulfonic acid (HEPES)- buffered medium with pH 7.4 at 25°C] and in response to NH⁴Cl pulse. pH(i) was measured with the pH-sensitive fluorescent dye 2',7'-bis(2-carboxyethyl)- 5,6-carboxyfluorescein (BCECF). Control resting pH(i) (7.1) and acidification in response to NH₄Cl pulse (minimum ph(i), 6.6) were essentially the same in snake tubules with oil-filled lumens or perfused lumens and in rabbit S2 proximal tubules with oil filled lumens. Rate of recovery of pH(i) (dpH(i)/dt) from acid to resting level in snake tubules (2.5 x 10^-3 pH U/s was about one-third of that in rabbit tubules. Resting pH(i) and dpH(i)/dt from acid to resting level were Na⁺ dependent in the distal portion but not the proximal portion of snake proximal tubules. However, dpH(i)/dt was not influenced by amiloride or 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid in snake proximal tubules, suggesting that the effect of Na⁺ on dpH(i)/dt and resting pH(i) may involve membrane potential. This study also indicates that oil-filled lumens do not interfere with measurements of resting pH(i) and do permit evaluation of pH(i) regulation at the basolateral membrane without complications from transport at the luminal membrane.