Activators of protein kinase A decrease the levels of free arachidonic acid in osteoblasts via stimulation of phosphatidylcholine and phosphatidylethanolamine synthesis

Abstract

In order to examine the role of protein kinase A (PKA) in the regulation of arachidonic acid availability, the interaction between cAMP agonists and the G protein activator AIF 4 − in their effects on phospholipid metabolism were measured in MC3T3-E1 osteoblasts. We show that forskolin and 8-brcAMP, activators of PKA, amplify the AIF 4 −-induced stimulation of phosphatidylinositol-specific phospholipase C (phosphatidylinositol inositolphosphohydrolase; EC 3.1.4.3), measured by the formation of [ 3H]inositol phosphates in prelabeled cells. However, the AIF 4 −-stimulated production of 1,2-diacylglycerols and the release of [ 3H] arachidonic acid ([ 3H]AA) were inhibited 50–75% by forskolin and 8-bromocAMP. Furthermore, pretreatment with PKA activators prevented much of the AIF 4 −-induced loss of [ 3H]AA from phosphatidylcholine and phosphatidylethanolamine in prelabeled osteoblasts. In addition, in the absence of AIF 4 −, forskolin was found to stimulate the incorporation of [ 3H]AA and [ 32P]orthophosphoric acid selectively into these two major phospholipids and selectively increased their mass. The effects of forskolin and 8-BrcAMP on the levels of free [ 3H]AA were completely reversed by pretreatment with the PKA inhibitor H-89. Therefore, our findings suggest that the activation of cAMP-dependent protein kinase can reduce the availability of free arachidonic acid for prostaglandin synthesis in osteoblast cells by stimulating its reesterification via phospholipid resynthesis.