pH homeostasis in pituitary GH4C1 cells: basal intracellular pH is regulated by cytosolic free Ca2+ concentration.
In GH4C1 cells, membrane depolarization induces a rapid and sustained increase in the cytosolic free calcium concentration ([Ca2+]i). In the present study we have investigated the role of [Ca2+]i in the regulation of basal intracellular pH (pHi). Depolarizing GH4C1 cells in buffer containing 0.4 mM extracellular Ca2+ decreased basal pHi from 7.02 +/- 0.04 to 6.85 +/- 0.03 (P less than 0.05). If the depolarization-induced influx of Ca2+ was inhibited by chelating extracellular Ca2+ or blocking influx through voltage-operated Ca2+ channels with nimodipine, no acidification was observed. Addition of TRH induced a rapid activation of Na+/H+ exchange in acidified cells, increasing pHi by 0.14 +/- 0.03 U. The action of TRH was blunted if extracellular Ca2+ was chelated; however, if influx of Ca2+ via voltage-operated channels was blocked by nimodipine, TRH still increased pHi. To deplete ATP, we incubated cells with 2-deoxy-D-glucose for 15-20 min and observed a decrease in basal pHi to 6.75 +/- 0.03 (P less than 0.05). No additional acidification was obtained when 2-deoxy-D-glucose-treated cells were depolarized, and no TRH-induced activation of Na+/H+ exchange was observed. Addition of ionomycin or 12-O-tetradecanoyl-phorbol-13-acetate separately to acidified cells had only modest effects on pHi; however, addition of 12-O-tetradecanoyl-phorbol-13-acetate and ionomycin together increased pHi markedly. We conclude that in GH4C1 cells, increasing [Ca2+]i reduces basal pHi through a mechanism dependent on influx of extracellular Ca2+ and independent of Na+/H+ exchange. In addition, elevation of [Ca2+]i and activation of protein kinase C act synergistically to enhance Na+/H+ exchange and increase pHi in acidified cells. Finally, normal cellular ATP is necessary for the activation of Na+/H+ exchange.
Törnquist
K
K
Endocrine Research Laboratory, University of Helsinki, Finland.
Tashjian
A H
AH
D000255:Adenosine Triphosphate / Q000378:metabolism; D000584:Amiloride / Q000494:pharmacology; D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002999:Clone Cells; D003600:Cytosol / Q000378:metabolism; D003847:Deoxyglucose / Q000494:pharmacology; D004347:Drug Interactions; D004734:Energy Metabolism / Q000187:drug effects; D004789:Enzyme Activation; D006706:Homeostasis; D006863:Hydrogen-Ion Concentration; D006982:Hypertonic Solutions; D015759:Ionomycin / Q000494:pharmacology; D007700:Kinetics; D010911:Pituitary Neoplasms; D011188:Potassium / Q000494:pharmacology; D011493:Protein Kinase C / Q000378:metabolism; D051381:Rats; D013050:Spectrometry, Fluorescence; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D013973:Thyrotropin-Releasing Hormone / Q000494:pharmacology
D006982:Hypertonic Solutions; D015759:Ionomycin; D013973:Thyrotropin-Releasing Hormone; D000584:Amiloride; D000255:Adenosine Triphosphate; D003847:Deoxyglucose; D011493:Protein Kinase C; D013755:Tetradecanoylphorbol Acetate; D011188:Potassium; D002118:Calcium