IN VIVO REGULATION OF cAMP/PKA SIGNALING BY PKIgamma

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DESCRIPTION (provided by applicant): PTH exerts both anabolic and catabolic actions on bone turnover primarily by activating cAMP/PKA signaling in osteoblasts. The catabolic action is primarily mediated by genes that are regulated in a sustained fashion. In contrast, the anabolic action is primarily mediated by immediate-early (I-E) genes that are rapidly, but transiently, up-regulated by PTH. Our overall hypothesis is that PKI( inhibits the transient anabolic action of cAMP/PKA signaling in osteoblasts. This overall hypothesis is based on our recent demonstration that endogenous PKI( inhibits the transient effects of PTH in an osteoblastic cell line. This inactivation of PKA terminates both expression of I-E genes and the anti-apoptotic effect of PTH. The anti-apoptotic effect of PTH was used as an indicator of the anabolic action of PTH since otherwise the anabolic action is difficult to study in cell culture. Since these results were obtained in an osteoblastic cell line, it is important to test their relevance in a more physiological setting. We are therefore proposing to determine whether endogenous PKI( regulates the anabolic actions of PTH in vivo and in primary osteoblast cultures. We have also demonstrated that the (-adrenergic agonist, isoproterenol, acts like PTH in osteoblastic cell lines to transiently induce nuclear PKA activity and I-E gene expression and to transiently inhibit apoptosis. These results provide the underpinning for our secondary hypothesis that intermittent administration of (-adrenergic agonists is anabolic for bone. Also like PTH, the transient effects of isoproterenol are terminated by endogenous PKI(, which further supports our overall hypothesis. Each of the Specific Aims will contribute to testing of our overall hypothesis by measuring the ability of endogenous PKI( to inhibit responses to PTH or (-adrenergic agonists in mice and in primary osteoblast cultures. The (-adrenergic agonist portions of the Specific Aims will also test our secondary hypothesis. Aim 1 will determine whether endogenous PKI( inhibits PKA signaling induced by PTH or by (-adrenergic agonists in mice and in primary osteoblast cultures. Aim 2 will determine whether endogenous PKI( inhibits I-E gene expression induced by PTH or by (-adrenergic agonists in mice and in primary osteoblast cultures. Aim 3 will determine whether endogenous PKI( inhibits the anabolic action of PTH or by (-adrenergic agonists in mice and in primary osteoblast cultures. Intermittent PTH is the only anabolic agent approved by the FDA for treatment of osteoporosis. However, a substantial proportion of patients are unresponsive to intermittent PTH. A better understanding of the mechanisms responsible for regulating the anabolic and catabolic actions of PTH might allow (1) identification of patients who are likely to be unresponsive, (2) improved monitoring of the effectiveness of PTH therapy, or (3) development of co-therapies that increase the anabolic actions of PTH, especially for patients who respond poorly to PTH by itself.