Induction of alkalinization in cultured renal cells (MDCK line) by prostaglandin E2.

The effect of prostaglandin E2 (PGE2), on the intracellular pH (pHi) in BCECF-loaded Madin Darby Canine Kidney (MDCK) cells was investigated. PGE2 elevated the pHi. Under resting conditions, pHi of MDCK cells suspended in PBS at pH 7.4 was 7.11 +/- 0.08; PGE2 increased pHi with an EC50 of 0.16 microM. PGF2 alpha elicited a similar response to PGE2, with an EC50 of 0.24 microM. Amiloride (0.4 mM) reversed the response to PGE2 (control 7.18 +/- 0.05; PGE2 7.26 +/- 0.05; after amiloride 7.18 +/- 0.05). In MDCK cells exposed to a Na(+)-free solution, alkalinization induced by this eicosanoid was blocked (Ringer-choline 7.16 +/- 0.03; PGE2 7.16 +/- 0.02). PGE2 increased by 100% the rate of recovery after an acidification pulse with ammonium chloride. In the presence of Ringer-HCO3- (pH 7.4), there was a delay in the maximal response to this prostaglandin (PBS 2.2 +/- 0.27, Ringer-bicarbonate 3.4 +/- 0.55 min) and the pHi increment was less marked than in PBS (0.09 pH units in HCO3- versus 0.16 pH units in PBS; P < 0.001). This effect of PGE2 was not blocked by 4,4'-diisothiocyanatostilbene-2,2'-disulfonic acid (1.0 mM). PMA (100 nM), activator of protein kinase C, mimicked the response to PGE2, suggesting the participation of this kinase on the effect of the prostanoid. As expected, two inhibitors of protein kinase C, staurosporine and sphingosine, abolished the response to PGE2. Staurosporine (0.10 microM), an inhibitor of protein kinase C, blocked the response to PGE2 (control 7.02 +/- 0.04; PGE2 and staurosporine 7.03 +/- 0.04, n = 9, not significant). Sphingosine, another inhibitor of protein kinase C, also blocked the response to PGE2. Two analogues of cAMP did not modify the pHi. In summary, PGE2 induced an intracellular alkalinization via stimulation of a Na+/H+ exchanger, with the participation of protein kinase C, in MDCK cells.