INTRACELLULAR PH-REGULATORY MECHANISMS IN PANCREATIC ACINAR-CELLS .1. CHARACTERIZATION OF H+ AND HCO3- TRANSPORTERS

  • 5 August 1990
    • journal article
    • research article
    • Vol. 265  (22) , 12806-12812
Abstract
Rat pancreatic acini loaded with the pH sensitive fluorescent dye 2'',7-bis(carboxyethyl)-5(6)-carboxyfluorescein were used to characterize intracellular pH (pHi) regulatory mechanisms in these cells. The acini were attached to cover slips and continuously perfused. In 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid (HEPES)-buffered solutions recovery from acid load (H+ efflux) required extracellular Na+ (Naout+) and was blocked by amiloride. Likewise, H+ influx initiated by removal of Naout+ was blocked by amiloride. Hence, in HEPES-buffered medium the major operative pHi regulatory mechanism is a Na+/H+ exchange. In HCO3-buffered medium, amiloride only partially blocked recovery from acid load and acidification due to Naout+ removal. The remaining fraction required Naout,+ was inhibited by H2-4,4''-diisothiocyanostilbene-2,2''-disulfunic acid (H2DIDS) and was independent of C1-. Hence, a transporter with characteristics of a Na+- HCO3- cotransport exists in pancreatic acini. Measurement of pHi changes due to Na+-HCO3- cotransport, suggests that the transporter contributes to HCO3- efflux under physiological conditions. Changing the Cl- gradient across the plasma membrane of acini maintained in HCO3-buffered solutions reveals the presence of an H2DIDS-sensitive, Na+-independent, Cl--dependent, HCO3- transporter with characteristics of a Cl-/HCO3- exchanger. In pancreatic acini the exchanger transports HCO3- but not OH- and under physiological conditions functions to remove HCO3- from the cytosol. In summary, only the Na+/H+ exchanger is functional in HEPES-buffered medium to maintain pHi at 7.28 .+-. 0.03. In the presence of 25 mM HCO3- at pH. of 7.4, all the transporters operate simultaneously to maintain a steady-state pHi of 7.13 .+-. 0.04.