Characterization of the TRH-induced activation of Na+/H(+)-exchange in pituitary GH4C1 cells.
In the present study in GH4C1 cells, the dependence of TRH-induced activation of Na+/H(+)-exchange on extracellular Na+ and Ca2+ was examined. Furthermore, the effects of both extracellular and intracellular H+ on Na+/H(+)-exchange were investigated. The buffering capacity was 63 +/- 11.8 mM (pH unit)-1 at basal intracellular pH (pHi) of 7.02 +/- 0.02. The initial rate of alkalinization in cells acidified with nigericin increased with increasing concentrations of extracellular Na+ according to simple Michaelis-Menten kinetics. The apparent Km-value for Na+ was 53 +/- 17.5 mM and the Vmax value was 28 +/- 4.5 mM H+/min. Addition of Na+ together with TRH increased Vmax to 56 +/- 6.4 mM H+/min (P < 0.05), while no difference was observed in Km. Decreasing extracellular pH (pHo) decreased the rate of alkalinization of acid-loaded cells, despite a large inward Na+ gradient. Furthermore, a decrease in pHi was necessary to obtain activation of Na+/H+ exchange. At pHi-values close to basal pHi no activation of Na+/H(+)-exchange was obtained. In addition, the results showed that extracellular Ca2+ was necessary for TRH-induced activation of Na+/H+ exchange. Blocking influx of extracellular Ca2+ with Ni2+ abolished the effect of TRH, suggesting that the TRH-induced activation of Na+/H(+)-exchange in GH4C1 cells is dependent on influx of extracellular Ca2+.
Törnquist
K
K
Endocrine Research Laboratory, University of Helsinki, Minerva Foundation, Institute for Medical Research, Finland.
D000584:Amiloride / Q000494:pharmacology; D000818:Animals; D001692:Biological Transport / Q000187:drug effects; D002021:Buffers; D002118:Calcium / Q000494:pharmacology; D002478:Cells, Cultured; D006859:Hydrogen / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D009532:Nickel / Q000494:pharmacology; D010902:Pituitary Gland / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D051381:Rats; D012964:Sodium / Q000378:metabolism*; D013973:Thyrotropin-Releasing Hormone / Q000494:pharmacology*
D002021:Buffers; D013973:Thyrotropin-Releasing Hormone; D000584:Amiloride; D009532:Nickel; D006859:Hydrogen; D012964:Sodium; D002118:Calcium