Regulation of phosphate uptake in primary cultured rabbit renal proximal tubule cells by glucocorticoids: Evidence for nongenomic as well as genomic mechanisms

We have investigated the nongenomic as well as the genomic effects of gluco corticoids on phosphate (Pi) uptake in primary rabbit renal proximal tubule cells (PTCs) and have defined the involved signaling pathways. In the pres ent study, cortisol-BSA (cortisol-BSA) (>10(-9) M, 30 min) was found to inh ibit Pi uptake in a time- and concentration-dependent manner. However, prog esterone-BSA (P-4-BSA), 17 beta -estradiol-BSA (E-2-BSA), testosterone-BSA (T-4-BSA), aldosterone, P-4, E-2, and T-4 (10(-9) M, 1 h) had no effect on Pi uptake. In addition, cortisol-BSA (10-9 hr) did not affect either Naf up take or alpha -methylglucopyranoside (cr-MG) uptake. The cortisol-BSA-induc ed inhibition of Pi uptake was associated with a decrease in the V-max for Pi uptake, rather than the K-m. The inhibitory effect of cortisol-BSA was n ot blocked either by actinomycin D (an inhibitor of transcription), cyclohe ximide (an inhibitor of translation), or classical glucocorticoid receptor antagonists (RU 486 or P-4). The cortisol-BSA-induced inhibition of Pi upta ke was blocked by two phospholipase C (PLC) inhibitors (neomycin or U73122) , and two protein kinase C (PKC) inhibitors (staurosporine or bisindolylmal eimide I) but not by two adenylate cyclase/protein kinase A inhibitors [SQ 22536 (an adenylate cyclase inhibitor) or myristoylated protein kinase A in hibitor amide 14-22]. Furthermore, cortisol-BSA promoted the translocation of PKC from the cytosolic fraction to the membrane fraction, while having n o effect on the activity of adenylate cyclase. Our observations may thus be interpreted as indicating that cortisol does indeed inhibit renal Pi uptak e via a nongenomic mechanism, which involves the PLC/PKC pathway.