When traversing microvascular beds, such as in the lung, red blood cells (RBCs) are subjected to mechanical deformation. Our previous findings that RBCs are required for flow-induced nitric oxide (NO) synthesis in the lung and that mechanical deformation of RBCs results in the release of adenosine triphosphate (ATP), a stimulus for endothelial NO synthesis, suggested a novel mechanism for the control of vascular resistance in the pulmonary circulation. In this construct, as the RBC is increasingly deformed by increments in the velocity of blood flow through a vessel and/or by reductions in vascular caliber, it releases ATP which stimulates the synthesis of NO resulting in relaxation of vascular smooth muscle and, thereby, an increase in vascular caliber. We propose that RBC-derived ATP contributes to the low vascular resistance of the healthy lung. Moreover, if deformation-induced ATP release from RBCs is an important determinant of vascular resistance in the lung, then a signal- transduction pathway for ATP release in response to this stimulus should be present in that cell. Here, we address the hypothesis that mechanical deformation of RBCs sets into motion specific signal transduction pathways which culminate in release of adenosine triphosphate (ATP). In this proposal we intend to 1) demonstrate that heterotrimeric G proteins are components of a signal transduction pathway for deformation-induced release of ATP from RBCs of rabbits and healthy humans, 2) establish that increases in intracellular cAMP are required for deformation- induced ATP release from these RBCs, 3) demonstrate that the activity of protein kinase A (PKA) is required for deformation- induced ATP release from RBCs of rabbits and healthy humans, 4) establish that ATP release from these RBCs requires the activity of the nucleoside transporter and 5) establish that deformation- induced ATP release from RBCs is decreased in humans with primary pulmonary hypertension and determine the associated defect(s) in the signal-transduction pathway for ATP release in these patients. The successful completion of the studies described in this proposal will define a new role for the RBC as a regulator of vascular resistance in the pulmonary circulation and may provide new insights into the pathophysiology of pulmonary hypertension. This hypothesis is the logical extension of our previous work and is consistent with a major focus of this group, namely, identification and characterization of those mechanisms responsible for the control of pulmonary vascular resistance.
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