The long term objective of this proposal is to characterize the autoregulatory humoral control mechanisms of bone blood flow, with particular emphasis on how these mechanisms may contribute to impaired bone flow with post-ischemic reperfusion. Prior work related to this project has indicated that both alpha1 and alpha2 adrenergic receptors are important for control of bone vascular smooth muscle tone. In adequate dosage, alpha1 receptor blockade may produce nearly complete vascular muscle relaxation and thus maximal vasodilation. The specific interests for this funding period will concern the role of endothelium, anorexia and bone storage techniques and circulating blood on vasodilation and bone blood flow with and without alpha adrenergic antagonism. The major portion of the study will utilize the cannulated canine tibia placed in an ex vivo perfusion apparatus using an asanguine oxygenated perfusate. This model permits precise control of flow rate, vascular resistance perfusate composition, pH and temperature and avoids intravascular thrombosis problems. The final year of this proposal will employ an in vivo model of microvascular canine tibia transfer for validation of data derived from the ex vivo studies. The investigation will be divided into four phases of one year each. The first will evaluate the role of endothelial-derived chemical mediators on vascular smooth muscle adrenergic receptors following varying periods of anoxia. The second phase will determine the role of optimal storage techniques on preservation of bone vascular endothelial integrity and smooth muscle receptor responsiveness. The third phase will identify the site of action within the bone vascular bed of vasoconstrictor responses and the relation of vasoconstriction to arteriovenous shunting and bone tissue bed capillary diffusion. The final phase will evaluate in vivo the influence of intraosseous vasodilation on blood flow, bone cell survival or repopulation and vascular anastomosis patency with and without prolonged anoxic storage. The data derived from this study is expected to be of particular relevance for understanding the important factors of blood flow regulation in free vascularized autografts and possibly future applications of vascularized bone allografts. This information may be useful for maximizing bone blood flow, intraosseous cell survival and bone healing. Many of the mechanisms ellucidated would be anticipated to be of relevance in understanding bone blood flow autoregulatory control under physiologic conditions as well.