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Am J Physiol Cell Physiol 270: C1051-C1060, 1996;
0363-6143/96 $5.00
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AJP - Cell Physiology, Vol 270, Issue 4 C1051-C1060, Copyright © 1996 by American Physiological Society

ARTICLES

ANP-stimulated cGMP egression in renal principal cells: abrogation of polarity by SV40 large T

V. Millul, D. Prie, M. Geniteau-Legendre, M. C. Verpont, B. Baudouin and P. M. Ronco
Institut National de la Sante et de la Recherche Medicale Unite 64, Hopital Tenon, Paris, France.

Egression of atrial natriuretic peptide (ANP)-stimulated guanosine 3', 5'-cyclic monophosphate (cGMP) was compared with that of isoproterenol-stimulated adenosine 3', 5'-cyclic monophosphate (cAMP) in a rabbit collecting duct cell line transformed with a temperature-sensitive strain of simian virus 40 (SV40). At 39.5 degrees C (inactivated large T), cells exhibit major features of principal cells, whereas at 33 degrees C (functional large T) they lose most of their specific properties. When cells were grown on plastic at 39.5 degrees C, both cyclic nucleotides were predominantly released extracellularly via probenecid-sensitive carriers. Probenecid (3mM) reduced the ratios of extracellular cGMP and cAMP by 84 and 70%, respectively. The amount of extracellular cGMP or cAMP ws linearly correlated with the time integral of the intracellular cyclic nucleotide, suggesting first-order kinetics. The apparent first-order rate constant (k) was sixfold greater for cGMP (0.139 +/- 0.037 min-1, n = 3 experiments) than for cAMP (0.022 +/- 0.003(-1), n = 3 experiments). 3-Isobutyl-1-methylxanthine markedly inhibited extrusion of cGMP (k = 0.022 +/- 0.003 min-1), whereas that of cAMP was unchanged. When cells were grown on filters at 39.5 degrees C, both nucleotides were predominantly released in the apical medium but with a greater polarity for cGMP (83 +/- 4%, n = 6 experiments) than for cAMP (60 +/- 6%, n = 3 experiments) and a prevailing apical localization of the probenecid-sensitive carrier. Activation of SV40 large T at 33 degrees C did not alter cyclic nucleotide transport characteristics but abolished the polarity of probenecid-sensitive cyclic nucleotide extrusion. These results suggest a physiological role for luminal cGMP in the rabbit collecting duct and a specific effect of large T on the probenecid-sensitive carrier polarity.


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