PHOSPHORYLATION AT A SINGLE-SITE IN THE RAT-BRAIN SODIUM-CHANNEL IS NECESSARY AND SUFFICIENT FOR CURRENT REDUCTION BY PROTEIN-KINASE-A

Voltage-gated sodium channels respond to excitatory inputs in nerve ce lls, generating spikes of depolarization at axon hillock regions and p ropagating the initial rising phase of action potentials through axons . It previously has been shown that protein kinase A (PKA) attenuates sodium current amplitude 20-50% by phosphorylating serines located in the I-II linker of the sodium channel. We have tested the individual c ontributions of five PKA consensus sites in the I-II linker by measuri ng sodium currents expressed in Xenopus oocytes during conditions of P KA induction, PKA was induced by perfusing oocytes with a cocktail tha t contained forskolin, chlorophenylthio-cAMP, dibutyryl-cAMP, and 3-is obutyl-1-methylxanthine. Phosphorylation at the second PKA site (serin e-573) was necessary and sufficient to diminish sodium current amplitu de, Phosphorylation at the third and fourth positions (serine-610 and serine-623) reduced current amplitude, but the effect was considerably smaller at those positions. Introduction of a negative charge at site 2 by substitution of serine-573 with an aspartate constitutively redu ced the basal level of sodium current, indicating that the attenuation of sodium current by phosphorylation of site 2 by PKA results from th e introduction of a negative charge at this site.