This study examined whether serotonin can activate protein kinase C in rat
heart endothelial cells. Protein kinase C isozyme translocation was examine
d by Western blot analysis with isozyme-specific anti-protein kinase C anti
body. In this study, only alpha protein kinase C isozyme was found to be tr
anslocated from the cytosolic to the particulate fractions after serotonin
stimulation. The effect of serotonin on the incorporation of P-32 from [gam
ma-P-32]ATP into peptide substrate was studied as another indicator of prot
ein kinase C activation. The experiments in this study demonstrated that th
e Ca2+-phospholipid-dependent protein kinase, protein kinase C, was activat
ed by serotonin. By investigating [H-3]phorbol 12,13-dibutyrate binding to
protein kinase C and trypsin-treated protein kinase C activity, we demonstr
ated that the site of action of serotonin is probably the regulatory domain
of protein kinase C. Finally, we also demonstrated that serotonin had no e
ffect on the intracellular concentration of cyclic nucleotides (cAMP, cGMP)
. These findings support the hypothesis that protein kinase C may be an imp
ortant participant in serotonin-induced endothelial cell contraction and ba
rrier dysfunction. (C) 2000 Elsevier Science B.V. All rights reserved.