K-252A AND STAUROSPORINE PROMOTE CHOLINE-ACETYLTRANSFERASE ACTIVITY IN RAT SPINAL-CORD CULTURES

The protein kinase inhibitor K-252a increased choline acetyltransferas e (ChAT) activity in rat embryonic spinal cord cultures in a dose-depe ndent manner (EC50 of approximately 100 nM) with maximal stimulatory a ctivity at 300 nM resulting in as much as a fourfold increase. A singl e application of K-252a completely prevented the marked decline in ChA T activity occurring over a 5-day period following culture initiation. Of 11 kinase inhibitors, only the structurally related inhibitor stau rosporine also increased ChAT activity (EC50 of approximately 0.5 nM). Effective concentrations of K-252a were not cytotoxic or mitogenic an d did not alter the total protein content of treated cultures. Insulin -like growth f actor 1, basic fibroblast growth f actor, ciliary neuro trophic factor, and leukemia inhibitory factor yielded dose-dependent increases in ChAT activity in spinal cord cultures. The combination of K-252a with insulin-like growth factor-I or basic fibroblast growth f actor increased ChAT activity up to eightfold over that of untreated c ontrols, which was greater than that observed with each compound alone . K-252a combined with ciliary neurotrophic factor or leukemia inhibit ory factor demonstrated no additive or synergistic effects on ChAT act ivity. These results suggest that there are multiple mechanisms for th e regulation of ChAT activity in spinal cord cultures. The enhancement of spinal cord ChAT activity by K-252a and staurosporine defines a ne w neurotrophic activity for these small organic molecules and raises t he possibility that they may activate some regulatory elements in comm on with the ciliary neurotrophic factor and leukemia inhibitory factor family of neurotrophic proteins.