THE ROLE OF PROTEIN-KINASE-C IN CELLULAR TOLERANCE TO ETHANOL

We have shown that ethanol inhibits uptake of adenosine by a specific nucleoside transporter in NG108-15 neuroblastoma x glioma cells and th at cAMP-dependent protein kinase (PKA) activity is required for this i nhibition. After chronic exposure to ethanol, adenosine uptake is no l onger inhibited on rechallenge with ethanol, i.e. transport has become tolerant to ethanol. Here we show that protein kinase C (PKC) contrib utes to ethanol-induced tolerance of adenosine transport. Activation o f PKC by phorbol esters in control cells results in an ethanol-toleran t phenotype, similar to that produced by chronic ethanol exposure. In addition, chronic exposure to ethanol increases the amounts of alpha, delta, and epsilon PKC. However, reducing PKC activity by inhibition w ith chelerythrine during chronic exposure to ethanol or down-regulatio n by phorbol esters prevents the development of ethanol-induced tolera nce of adenosine transport. By contrast, the inhibition of PKA activit y produces tolerance to ethanol inhibition of adenosine uptake. When p rotein phosphatase inhibitors are present, inhibiting PKA activity has no effect on ethanol sensitivity of adenosine uptake, suggesting a ro le for protein phosphatases in the regulation of ethanol sensitivity o f uptake. Taken together, our results suggest that PKA and PKC have op posing effects on the ethanol sensitivity of adenosine transport; PKA activity is required for ethanol sensitivity, and PKC activation produ ces tolerance. Based on these data, we propose that chronic ethanol ex posure increases PKC activity, leading to the activation of a protein phosphatase (1 or 2A). This phosphatase then dephosphorylates a PKA-ph osphorylated site, which is required for ethanol to inhibit adenosine uptake. Therefore, the sensitivity of adenosine transport to ethanol a ppears to be maintained by a balance of PKA and protein phosphatase ac tivities, and PKC may regulate phosphatase activity.