Premium
Effect of Protein Kinase C Modulators on 14,15‐Epoxyeicosatrienoic Acid Incorporation into Astroglial Phospholipids
Author(s) -
Shivachar Amruthesh C.,
Willoughby Karen A.,
Ellis Earl F.
Publication year - 1995
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.1471-4159.1995.65010338.x
Subject(s) - epoxyeicosatrienoic acid , arachidonic acid , protein kinase c , biochemistry , phosphatidylinositol , epoxide hydrolase , phosphatidylcholine , metabolism , epoxide hydrolase 2 , eicosanoid metabolism , diacylglycerol kinase , chemistry , eicosanoid , kinase , biology , enzyme , phospholipid , membrane , microsome
Our previous studies have shown that 14,15‐epoxyeicosatrienoic acid (14,15‐EET) is a major product of arachidonic acid metabolism in astrocytes. The purpose of this study was to investigate cellular regulation of 14,15‐EET incorporation, distribution, and metabolism in primary cultures of rat brain cortical astrocytes. Incorporation of 14,15‐EET into astrocytes was lower (93,390 ± 11,121 dpm/5 × 10 6 cells) than incorporation of 8,9‐EET (226,500 ± 5,567 dpm/5 × 10 6 cells) and arachidonic acid (321,600 ± 1,200 dpm/5 × 10 6 cells). 14,15‐EET was distributed in the order neutral lipids and free fatty acids (solvent front) ≫ phosphatidylcholine (PC) > phosphatidylinositol (PI) > phosphatidylethanolamine. In contrast, 8,9‐EET and arachidonic acid were exclusively incorporated into PC. During incubation, astroglial epoxide hydrolase selectively metabolized 14,15‐EET, but not 8,9‐EET, to its vic ‐diol. Although 4‐phenylchalcone oxide, a potent inhibitor of epoxide hydrolase, completely inhibited 14,15‐EET metabolism, a large amount of cell‐incorporated radioactivity remained as free 14,15‐EET. Long‐term exposure of astrocytes to 4β‐phorbol 12‐myristate 13‐acetate (4β‐PMA) resulted in a time‐dependent incorporation of 14,15‐EET into PI but not in control cells exposed to 4α‐phorbol 12,13‐didecanoate. PKC down‐regulation completely inhibited epoxide hydrolase metabolism of 14,15‐EET. Following recovery of down‐regulated PKC, 1 week after treatment with 4β‐PMA, astrocytes regained their normal pattern of low incorporation of 14,15‐EET. Protein kinase C (PKC) inhibition by staurosporine enhanced 14,15‐EET incorporation without affecting its metabolism to 14,15‐dihydroxyeicosatrienoic acid. Incorporation of 14,15‐EET by PKC‐down‐regulated cells was inhibited by thimerosal, a known inhibitor of fatty acyl‐CoA synthase. Our results suggest that the lower incorporation of 14,15‐EET into astroglial cells may be due to modulation of PKC‐mediated cellular mechanism(s).