Abstract
The transport of a fluorescent rapamycin derivative was measured in killifish (Fundulus heteroclitus) renal proximal tubules by means of confocal microscopy and image analysis. Renal cells and tubular lumens rapidly accumulated the rapamycin analog from the medium and attained steady state within 60 min. At steady state, luminal fluorescence intensity was two to four times higher than cellular fluorescence. Cellular fluorescence intensity was a linear function of medium substrate concentration and was not affected by any treatment used. In contrast, luminal fluorescence exhibited a saturable component as the medium concentration of the rapamycin derivative was increased. Secretion into the lumen was blocked by KCN, rapamycin, cyclosporin A and substrates for p-glycoprotein (verapamil, PSC-833 and FK506), but not by substrates for the renal organic anion or organic cation transport systems, such as p-aminohippurate, leukotriene C4 or tetraethylammonium. Finally, rapamycin blockedp-glycoprotein-mediated secretion of a fluorescent cyclosporin A derivative. The data are consistent with the fluorescent rapamycin analog entering proximal tubule cells by simple diffusion and then being pumped into the tubular lumen byp-glycoprotein. They suggest that the parent compound, rapamycin, would be handled similarly.
Footnotes
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Send reprint requests to: Dr. David S. Miller, LPC, NIH/NIEHS, P.O. Box 12233, Research Triangle Park, NC 27709.
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↵1 Supported in part by a grant from the Deutsche Forschungsgemeinschaft to G.F. and a travel grant from Mundipharma Pharmaceuticals to J.D.
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This study was also supported by a NATO CRG grant.
- Abbreviations:
- CSA
- cyclosporin A
- DMSO
- dimethylsulfoxide
- LTC4
- leukotriene C4
- MDR
- multidrug resistance
- NA
- numerical aperature
- NBD-rapamycin
- (4-nitrobenzofurazan-7-yl)-rapamycin
- NBDL-CSA
- [N-ε(4-nitrobenzofurazan-7-yl)-d-Lys8]-cyclosporin
- PAH
- p-aminohippurate
- TEA
- tetraethylammonium
- Received November 22, 1996.
- Accepted March 5, 1997.
- The American Society for Pharmacology and Experimental Therapeutics
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