Effect of protein kinase C activation and down-regulation on active calcium transport.
Active transcellular Ca2+ transport in primary cultures of the rabbit kidney collecting system has been shown to be responsive to PTH through activation of protein kinase A (PKA). The present study investigates an additional regulatory pathway activated by protein kinase C (PKC). Cells from rabbit kidney connecting tubules and cortical collecting ducts were isolated by immunodissection and subsequently cultured on permeable filters. Incubation of cultured cells with the PKC activator, 12-O-tetradecanoylphorbol 13-acetate (TPA, 10(-8) to 10(-6) M) had a dual effect on active transcellular Ca2+ transport. Short-term incubation increased membrane-associated PKC activity within 10 minutes and decreased active transcellular Ca2+ transport dose-dependently (IC50 = 3.4 +/- 0.4 nM), with a maximal inhibition of 74 +/- 3%. TPA (10(-7) M) concomitantly inhibited the amiloride-sensitive transepithelial potential difference (p.d.) and short-circuit current across the monolayers. After prolonged exposure to TPA, total cellular PKC activity was down-regulated, resulting in a maximal 65 +/- 5% reduction after one hour. Interestingly, this latter event was temporally separated from a gradual return of both Ca2+ absorption rate and transepithelial p.d. to control levels occurring over 96 and 48 hours, respectively, of further incubation with TPA. The inhibitor of protein kinase activity, staurosporine (10(-7) M), when present during incubation with submaximal concentration of TPA (10(-8) M) partly prevented the TPA-induced inhibition of Ca2+ absorption from 54 +/- 4 to 27 +/- 3%. This study demonstrates for the first time that, in addition to PKA, activation of PKC plays a regulatory role in transcellular Ca2+ reabsorption in the renal collecting system.
Bindels
R J
RJ
Department of Physiology, University of Nijmegen, The Netherlands.
Dempster
J A
JA
Ramakers
P L
PL
Willems
P H
PH
van Os
C H
CH
D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002478:Cells, Cultured; D015536:Down-Regulation; D004789:Enzyme Activation / Q000187:drug effects; D017136:Ion Transport / Q000187:drug effects; D007685:Kidney Tubules, Collecting / Q000187:drug effects / Q000378:metabolism*; D011493:Protein Kinase C / Q000378:metabolism*; D011817:Rabbits; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology
D011493:Protein Kinase C; D013755:Tetradecanoylphorbol Acetate; D002118:Calcium