Activation of Na/H exchanger in mesangial cells is associated with translocation of PKC isoforms

We investigated the potential coupling of the Na/H exchanger to protein kinase C (PKC) activation. Mesangial cells (MC), passage 3-8, were exposed to either serotonin (5-HT), arginine vasopressin (AVP), or fibroblast growth factor (FGF). We assessed the effect of these agonists on recovery from an acid load (NH4+/NH3) in the nominal absence of CO2/HCO3. 5-HT, AVP, and FGF significantly enhanced the rate of recovery from 24.2 +/- 4.5 x 10(-4) intracellular pH units/s to 67.7 +/- 5.7, 70.5 +/- 5.1, and 55.7 +/- 6.8, respectively (n > 6, P < 0.0001). The increase in activity was abolished by ethylisopropylamiloride and removal of extracellular Na+, thereby suggesting that activation of Na/H exchanger activity was responsible for enhanced recovery by 5-HT, AVP, and FGF. MC were then pretreated with phorbol ester [phorbol 12-myristate 13-acetate (PMA); 10 microM for 40 h] and acid loaded. 5-HT and AVP did not enhance recovery of PMA-pretreated cells (23.3 +/- 6.8 and 24.9 +/- 4.7, n > 4, not significant), whereas FGF stimulated activity (48.2 +/- 5.5, n > 4, P < 0.001). Similar results were found when MC were pretreated with the PKC antagonist, sphingosine. Specific antibodies were raised against alpha-, beta I-, beta II-, and gamma-isoforms of PKC. We observed that MC express the alpha-, gamma-, and beta I-isoforms, but not the beta II-isoform of PKC.(ABSTRACT TRUNCATED AT 250 WORDS)