Intracellular pH in snake renal proximal tubules

Abstract

Intracellular pH (pHi) 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 degrees C] and in response to NH4Cl pulse. pHi was measured with the pH-sensitive fluorescent dye 2',7'-bis(2-carboxyethyl)-5,6-carboxyfluorescein (BCECF). Control resting pHi (7.1) and acidification in response to NH4Cl pulse (minimum pHi, 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 pHi (dpHi/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 pHi and dpHi/dt from acid to resting level were Na+ dependent in the distal portion but not the proximal portion of snake proximal tubules. However, dpHi/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 dpHi/dt and resting pHi may involve membrane potential. This study also indicates that oil-filled lumens do not interfere with measurements of resting pHi and do permit evaluation of pHi regulation at the basolateral membrane without complications from transport at the luminal membrane.