STUDY ON THE POSSIBLE INVOLVEMENT OF PROTEIN-KINASES IN THE MODULATION OF BRAIN PRESYNAPTIC SODIUM-CHANNELS - COMPARISON WITH CALCIUM CHANNELS

A possible modulatory role of kinases on voltage sensitive Na+ channel s of presynaptic brain nerve endings was investigated by testing the e ffect of several kinase activators and inhibitors on the elevation of [Na-i] induced by veratridine in mouse brain synaptosomes loaded with a selective Na+ indicator dye. Veratridine (20 mu M) increases the bas al [Na-i] level (20 mM) more than twofold. This increase is independen t of external Ca2+, but abolished by tetrodotoxin (1 mu M). Activation of cAMP dependent protein kinase with forskolin or cAMP analogs, or o f protein kinase C with diacylglycerol did not affect the veratridine- induced elevation in [Na-i]. Drugs reported to inhibit calmodulin-depe ndent events, as well as the regulatory domain of protein kinase C, we re potent and effective inhibitors of the increase in [Na-i] induced b y veratridine, as well as other veratridine induced responses, namely elevation of [Ca-i] (monitored with the Ca2+ indicator dye fura-2) and neurotransmitter (GABA) release. Drugs that inhibit kinases by bindin g to the catalytic site were ineffective, however, as was the phosphat ase inhibitor, okadaic acid. A selective inhibitor of Ca2+ and calmodu lin dependent protein kinase II also did not affect the elevation of [ Na-i] induced by veratridine, but markedly diminished the elevation of [Ca-i] induced by depolarization either with veratridine or with high K+ (15 and 30 mM). On the basis of these results it is concluded that , the dramatic inhibition exerted by some of the drugs tested on the e levation of [Na-i] induced by veratridine is not due to their effects on kinases, but to a possible interaction of these compounds with an i ntracellular site of the Naf channel. On the other hand, while Ca2+ an d calmodulin dependent protein kinase II is unable to modulate brain p resynaptic voltage sensitive Na+ channels, it facilitates the activati on of brain presynaptic voltage sensitive Ca2+ channels. (C) 1998 Else vier Science Ltd. All rights reserved.