Intracellular pH in frog skin: effects of Na+, volume, and cAMP

Single skins were analyzed by {sup 31}P-nuclear magnetic resonance (NMR) spectroscopy during alternative perfusion with control and experimental solutions. Intracellular (pH{sub c}) and extracellular (pH{sub o}) pH were monitored by measuring the spectral frequencies of intracellular P{sub i} and external methylphosphonate, respectively. Base-line pH{sub c} was 7.20 {plus minus} 0.02 (SE) when pH{sub o} was 6.99 {plus minus} 0.02. A 4-acetamido-4{prime}-isothiocyanostilbene-2,2{prime}-disulfonic acid (SITS)-inhibitable, HCO{sup {minus}}{sub 3}-dependent alkaline shift in pH{sub c} can be elicited by replacing external Cl{sup {minus}} by gluconate or sulfate. The authors now report that this effect is observed even in sodium-free media. The substitution of gluconate for external Cl{sup {minus}} has also been reported to shrink cell volume. This shrinkage can be minimized by replacing Cl{sup {minus}} with gluconate during perfusion with hypotonic, rather than isotonic, media. Conducted in this manner, the anionic substitution produced a much smaller alkaline shift in pH{sub c}. In the presence of the Na-H antiport blocker 5-(N-methyl-N-isobutyl)amiloride (MIA), restoration of external Na{sup +} did not increase pH{sub c}. Adenosine 3{prime},5{prime}-cyclic monophosphate (cAMP) also alters pH{sub c}. The data suggest that frog skin regulates pH{sub c} by the parallel operation of Na-H and Na{sup +}-independent Cl-HCO{sub 3} antiports. Cell volume and cAMPmore » may play regulating roles in this epithelium.« less