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- Title
Protein kinase C and phosphatidylinositol 3-kinase independently contribute to P-glycoprotein-mediated drug secretion in the mouse proximal tubule.
- Authors
Tsuruoka, Shuichi; Sugimoto, Koh-ichi; Fujimura, Akio; Imai, Masashi; Asano, Yasushi; Muto, Shigeaki
- Abstract
We have recently reported that the apical membrane of the proximal tubule S2 segment from wild-type mice (WT mice) has the capacity for P-glycoprotein- (P-gp-) mediated drug efflux, whereas mice in which both mdr1a and mdr1b genes are disrupted (KO mice) do not. To examine the intracellular regulatory mechanisms of drug-transporting P-gp activity, we isolated and perfused proximal tubule S2 segments from WT and KO mice, and measured luminal efflux of the intracellular fluorescence of rhodamine 123, a fluorescent substrate of P-gp. The decay half-time of the intracellular fluorescence (t1/2) was regarded as an index of the drug-transporting P-gp activity. In the WT mice, the t1/2 was 36±5 s (n=35) during the basal period, and was significantly increased to 440±45 s by the luminal addition of verapamil (an inhibitor of P-gp). The addition of phorbol 12-myristate 13-acetate (PMA) [a protein kinase C (PKC) activator] to the bath increased t1/2, but 4α-phorbol (the inactive form of PMA) did not. The PMA-induced increase in t1/2 was further increased by the luminal addition of verapamil, and was partially inhibited by co-treatment with staurosporine or H-7 (inhibitors of PKC). Pretreatment with wortmannin or LY294002 [inhibitors of phosphatidylinositol 3-kinase (PI 3-kinase)] added to the bath also increased t1/2. The wortmannin- and LY294002-induced increase in t1/2 was also further increased by the luminal addition of verapamil. The effects of PMA on t1/2 were additive after co-treatment with wortmannin or LY294002. In the KO mice, t1/2 was 440±25 s (n=18) during the basal period, and was no longer affected by the addition of verapamil, staurosporine, H-7, wortmannin, or LY294002. Based on the use of pharmacological agents, we conclude that in the proximal tubule from WT mice, P-gp-mediated drug secretion occurs independently via PKC- and PI 3-kinase-dependent processes, whereas it is not present in KO mice, independently of PKC- and PI 3-kinase.
- Subjects
PROTEIN kinase C; PHOSPHOINOSITIDES; CELL membranes; GLYCOPROTEINS; DRUGS; GLYCOCONJUGATES
- Publication
Pflügers Archiv: European Journal of Physiology, 2001, Vol 442, Issue 3, p321
- ISSN
0031-6768
- Publication type
Article
- DOI
10.1007/s004240100542