sirolimus and cypermethrin

The interaction of FK-506 with K(V)1.3, stably expressed in Chinese hamster ovary cells, was investigated with the whole cell patch-clamp technique. FK-506 inhibited K(V)1.3 in a reversible, concentration-dependent manner with an IC(50) of 5.6 microM. Rapamycin, another immunosuppressant, produced effects that were similar to those of FK-506 (IC(50) = 6.7 microM). Other calcineurin inhibitors (cypermethrin or calcineurin autoinhibitory peptide) alone had no effect on the amplitude or kinetics of K(V)1.3. In addition, the inhibitory action of FK-506 continued, even after the inhibition of calcineurin activity. The inhibition produced by FK-506 was voltage dependent, increasing in the voltage range for channel activation. At potentials positive to 0 mV (where maximal conductance is reached), however, no voltage-dependent inhibition was found. FK-506 exhibited a strong use-dependent inhibition of K(V)1.3. FK-506 shifted the steady-state inactivation curves of K(V)1.3 in the hyperpolarizing direction in a concentration-dependent manner. The apparent dissociation constant for FK-506 to inhibit K(V)1.3 in the inactivated state was estimated from the concentration-dependent shift in the steady-state inactivation curve and was calculated to be 0.37 microM. Moreover, the rate of recovery from inactivation of K(V)1.3 was decreased. In inside-out patches, FK-506 not only reduced the current amplitude but also accelerated the rate of inactivation during depolarization. FK-506 also inhibited K(V)1.5 and K(V)4.3 in a concentration-dependent manner with IC(50) of 4.6 and 53.9 microM, respectively. The present results indicate that FK-506 inhibits K(V)1.3 directly and that this effect is not mediated via the inhibition of the phosphatase activity of calcineurin.

Topics: Animals; Calcineurin; Calcineurin Inhibitors; CHO Cells; Cricetinae; Cricetulus; Dose-Response Relationship, Drug; Immunosuppressive Agents; Kinetics; Kv1.3 Potassium Channel; Kv1.5 Potassium Channel; Membrane Potentials; Patch-Clamp Techniques; Potassium Channel Blockers; Pyrethrins; Shal Potassium Channels; Sirolimus; Tacrolimus; Transfection

S100betabeta is a calcium binding, neurotrophic protein produced by nonneuronal cells in the nervous system. The pathway by which it enhances neuronal survival is unknown. Here we show that S100betabeta enhances survival of embryonic chick forebrain neurons in a dose-dependent manner. In the presence of suboptimal amounts of S100betabeta, neuronal survival is enhanced by the immunosuppressants FK506 and cyclosporin A at concentrations that inhibit calcineurin, which is present in these cells. Rapamycin, an immunosuppressant that does not inhibit calcineurin, did not enhance cell survival. Cypermethrin, a direct and highly specific calcineurin inhibitor, mimicked the immunophilin ligands in its neurotrophic effect. None of the drugs stimulated neuronal survival in the absence of S100betabeta. In the presence of suboptimal amounts of S100betabeta, FK506, cyclosporin A, and cypermethrin (but not rapamycin) also increased NF-kappaB activity, as measured by immunofluorescence of cells stained with antibody to the active subunit (p65) and by immunoblotting of nuclear extracts. Antioxidant and glucocorticoid inhibitors of NF-kappaB decreased both the amount of active NF-kappaB and the survival of neurons caused by S100betabeta alone or in the presence of augmenting drugs. We conclude that S100betabeta enhances the survival of chick embryo forebrain neurons through the activation of NF-kappaB.

Topics: Animals; Antioxidants; Brain Chemistry; Calcineurin Inhibitors; Calcium-Binding Proteins; Cattle; Cell Nucleus; Cell Survival; Cells, Cultured; Chick Embryo; Coumarins; Cyclosporine; Dexamethasone; Dimerization; Immunophilins; Immunosuppressive Agents; Isocoumarins; Macromolecular Substances; Nerve Growth Factors; Neurons; NF-kappa B; Prosencephalon; Pyrethrins; Pyrrolidines; S100 Calcium Binding Protein beta Subunit; S100 Proteins; Serine Proteinase Inhibitors; Sirolimus; Tacrolimus; Thiocarbamates