A role for PKC epsilon and MAP kinase in bradykinin-induced arachidonic acid release in rabbit CCD cells.
Arachidonic acid (AA) release is the rate-limiting step in the production of prostaglandins, an important class of autocrine/paracrine factors that modulate collecting duct function. Previous results from this laboratory have established cytosolic phospholipase A2 (cPLA2) as the enzyme responsible for bradykinin (BK)-stimulated AA mobilization in rabbit cortical collecting duct (RCCD) cells, and the present study pursues the intracellular signaling mechanisms responsible for its activation. Pretreatment of cells with Ro-31-8220, an inhibitor of protein kinase C (PKC), or PD-98059, an inhibitor of the mitogen-activated protein kinase (MAPK) cascade, resulted in a 50-60% reduction in BK-stimulated AA release. Incubation of RCCD cells with a combination of both Ro-31-8220 and PD-98059 did not achieve a greater inhibition of either BK-stimulated AA release or cPLA2 activity, possibly indicating that MAPK activation was dependent upon prior activation of PKC. This was supported by the observation that BK-induced MAPK activation could be reversed by either inhibitor. Additional experiments dealing with immunoblots for PKC isozymes revealed that RCCD cells express PKC species alpha, gamma, epsilon, and zeta. Following BK stimulation, only PKC epsilon translocated to the particulate fraction. Based on these results, it appears that PKC is activated and involved in the sequential activation of MAPK and cPLA2 following BK treatment. The results also suggest that PKC epsilon may be the isozyme implicated in the process.
Lal
M A
MA
Department of Cellular and Molecular Medicine, Faculty of Medicine, University of Ottawa, Ontario, Canada.
Proulx
P R
PR
Hébert
R L
RL
D000818:Animals; D016718:Arachidonic Acid / Q000378:metabolism*; D001920:Bradykinin / Q000494:pharmacology*; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000502:physiology*; D002478:Cells, Cultured; D003600:Cytosol / Q000201:enzymology; D007527:Isoenzymes / Q000502:physiology*; D007672:Kidney Cortex; D007685:Kidney Tubules, Collecting / Q000166:cytology / Q000378:metabolism*; D010741:Phospholipases A / Q000378:metabolism; D054467:Phospholipases A2; D011493:Protein Kinase C / Q000502:physiology*; D051744:Protein Kinase C-epsilon; D011817:Rabbits; D015398:Signal Transduction / Q000502:physiology
D007527:Isoenzymes; D016718:Arachidonic Acid; D011493:Protein Kinase C; D051744:Protein Kinase C-epsilon; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D010741:Phospholipases A; D054467:Phospholipases A2; D001920:Bradykinin