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Effect of Protein Kinase C Activation on the Release of [ 3 H]Acetylcholine in the Presence of Vesamicol
Author(s) -
Barbosa J.,
Clarizia A. D.,
Gomez M. V.,
RomanoSilva M. A.,
Prado V. F.,
Prado M. A. M.
Publication year - 1997
Publication title -
journal of neurochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.75
H-Index - 229
eISSN - 1471-4159
pISSN - 0022-3042
DOI - 10.1046/j.1471-4159.1997.69062608.x
Subject(s) - vesicular acetylcholine transporter , protein kinase c , acetylcholine , calphostin c , calphostin , phosphorylation , stimulation , chemistry , phorbol , biochemistry , activator (genetics) , medicine , endocrinology , biology , gene , choline acetyltransferase
The present work tested whether pharmacological activation of protein kinase C (PKC) influences the release of [ 3 H]‐acetylcholine ([ 3 H]ACh) synthesized in the presence of vesamicol, an inhibitor of the vesicular acetylcholine transporter (VAChT). Newly synthesized [ 3 H]ACh was released from hippocampal slices by field stimulation (15 Hz) in the absence of vesamicol, but as expected [ 3 H]ACh synthesized during exposure to vesamicol was not released significantly by stimulation. Treatment of slices with the PKC activator phorbol myristate acetate (PMA) decreased the inhibitory effect of vesamicol on [ 3 H]ACh release. The effect of PMA was dose‐dependent, was sensitive to calphostin C, a PKC‐selective inhibitor, and could not be mimicked by α‐PMA, an inactive phorbol ester. PMA did not alter the release of [ 3 H]ACh in the absence of vesamicol, suggesting that the site of PKC action could be related to the VAChT. In agreement with this observation, immunoprecipitation of VAChT from 32 P‐labeled synaptosomes showed that phosphorylation occurs and that incorporation of 32 P in the VAChT protein increases in the presence of PMA. We suggest that PKC alters the output of [ 3 H]ACh formed in the presence of vesamicol and also provide circumstantial evidence for a role of phosphorylation of VAChT in this process.