Intracellular pH and growth hormone-releasing factor-stimulated adenosine 3'5'-monophosphate, intracellular calcium and growth hormone release from rat anterior pituitary cells.
In this study, we examined the effect of changes in intracellular pH (pHi) on basal and GH-releasing factor (GRF)-stimulated cyclic AMP (cAMP), intracellular Ca2+ and GH release using a static monolayer culture prepared from dispersed rat anterior pituitary cells. To modulate pHi, two approaches were used: variation of extracellular pH (pHo) and addition of sodium propionate and ammonium chloride which alter pHi directly. Direct pHi measurement with 2'7'-bis(carboxyethyl)-5(6)-carboxyfluorescein showed that for pHo values between 6.9 and 7.6, a change in pHo of 0.1 units resulted in a change in pHi of 0.045 units. Sodium propionate (30 mmol/l) reduced pHi by 0.06 units whereas ammonium chloride (30 mmol/l) increased pHi by 0.1 units. Increasing pHo from 6.6 to 7.8 enhanced the maximal GRF-stimulated cAMP and GH responses by 80% and 300% respectively, indicating that the GRF-stimulated cAMP and GH release were both pH-dependent. Acute elevation of pHo from 6.6 to 7.8 also increased basal GH release by sixfold. Reduction of pHi by sodium propionate, however, had no significant effect on GRF-stimulated cAMP levels while the corresponding GRF-stimulated GH release was reduced by up to 40%. In comparison, elevation of pHi by ammonium chloride enhanced the GRF-stimulated cAMP release by up to 75% and the corresponding increase in GH was less than 20%. When the relationship between pHi and intracellular Ca2+ was determined with the fluorescent Ca2+ indicator, Fura-2, it was found that increasing pHo and treatment with ammonium chloride increased intracellular Ca2+, while sodium propionate and reducing pHi had no effect on intracellular Ca2+. These results indicate that activation of adenylate cyclase and mobilization of intracellular Ca2+, two intracellular signalling pathways of importance to GH secretion, are both sensitive to changes in pHi.
Puttagunta
A L
AL
Department of Physiology, Faculty of Medicine, University of Alberta, Edmonton, Canada.
Chik
C L
CL
Girard
M
M
O'Brien
L
L
Ho
A K
AK
D000643:Ammonium Chloride / Q000494:pharmacology; D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002478:Cells, Cultured; D000242:Cyclic AMP / Q000378:metabolism*; D013006:Growth Hormone / Q000096:biosynthesis*; D013007:Growth Hormone-Releasing Hormone / Q000494:pharmacology*; D006863:Hydrogen-Ion Concentration; D010903:Pituitary Gland, Anterior / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D051381:Rats; D013268:Stimulation, Chemical
D000643:Ammonium Chloride; D013006:Growth Hormone; D013007:Growth Hormone-Releasing Hormone; D000242:Cyclic AMP; D002118:Calcium