Microfluorometric monitoring of pHi in cultured heart cells: Na+-H+ exchange

Abstract

Continuous measurement of intracellular pH (pHi) should enhance the likelihood of defining the mechanisms of pHi regulation in actively contracting preparations of cardiac muscle. A filter microfluorometric technique was adapted for use with growth-oriented embryonic chick heart cells to continuously monitor changes in the fluorescence intensity of the pH-sensitive chromophore 6-carboxyfluorescein, generated in situ. Data pertaining to the direction and the rate of pHi changes assisted in thermodynamically characterizing and ascertaining net kinetic parameters of a Na+-H+ exchange mechanism. Imposing an outward Na+ gradient across the cardiac cell membrane rapidly (t 1/2 = 44 s) induced cytosolic acidification, whereas an inward Na+ gradient produced cytosolic alkalinization (t 1/2 = 40 s). Amiloride (10(-3) M) caused the cytoplasm to acidify (t 1/2 = 46 s) and also reversibly blocked the acidification induced by low extracellular Na+. These results are consistent with the presence of a rapid Na+-H+ exchange mechanism in the cardiac cell membrane. Further investigations are required to characterize the involvement of Na+-H+ exchange in pHi regulation and to differentiate the effects of Na+-H+ exchange from other ion gradient-coupled mechanisms, e.g., Na+-Ca2+ exchange.