The effect of rapamycin on single ENaC channel activity and phosphorylation in A6 cells.

Rapamycin and FK-506 are immunosuppressive drugs that bind a ubiquitous immunophilin, FKBP12, but immunosuppressive mechanisms and side effects appear to be different. Rapamycin binds renal FKBP12 to change renal transport. We used cell-attached patch clamp to examine rapamycin's effect on Na(+) channels in A6 cells. Channel NP(o) was 0.5 +/- 0.08 (n = 6) during the first 5 min but fell close to zero after 20 min. Application of 1 microM rapamycin reactivated Na(+) channels (NP(o) = 0.47 +/- 0.1; n=6), but 1 microM FK-506 did not. Also, GF-109203X, a protein kinase C (PKC) inhibitor, mimicked the rapamycin-induced reactivation in a nonadditive manner. However, rapamycin did not reactivate Na(+) channels if cells were exposed to 1 microM FK-506 before rapamycin. In PKC assays, rapamycin was as effective as the PKC inhibitor; however, epithelial Na(+) channel (ENaC) phosphorylation was low under baseline conditions and was not altered by PKC inhibitors or activators. These results suggest that rapamycin activates Na(+) channels by binding FKBP12 and inhibiting PKC, and, in renal cells, despite binding the same immunophilin, rapamycin and FK-506 activate different intracellular signaling pathways.