Acutely, hypoxia increases pulmonary vasomotor tone, resulting in acute hypoxic pulmonary vasoconstriction (AHPV). Chronically, hypoxia causes pulmonary vascular remodeling and sustained increase in tone, resulting in chronic hypoxic pulmonary vasoconstriction (CHPV). The mechanisms of AHPV and CHPV remain unknown. Evidence suggests that calcium influx from extracellular fluid into pulmonary artery smooth muscle cells (PASMCs) plays a role. In many cells, depletion of Ca2+ stored in sarcoplasmic reticulum (SR)causes """"""""capacitative Ca2+ entry (CCE)"""""""" through store-operated Ca2+ channels (SOCCs) composed of proteins homologous to """"""""transient receptor potential"""""""" (TRP) proteins in Drosophila. Our preliminary data indicates that hypoxia increased resting [Ca2+]i and CCE in PASMCs and that TRPC1 and -6 gene expression in PASMCs was enhanced by hypoxia, perhaps due to activation of the transcription factor, hypoxia-inducible factor 1 (HIF-1). In this proposal, we test the hypotheses that in PASMCs acute hypoxia causes activation of CCE through sarcolemmal SOCCs composed of TRPC proteins, leading to increased [Ca2+]i and AHPV, while chronic hypoxia causes a sustained increase in CCE and resting [Ca2+]i in PASMCs due to HIF-1 dependent upregulation of TRPC protein expression, leading to CHPV. As initial tests of these hypotheses, we will determine: 1) if treatment of PASMCs with small interfering RNA (siRNA) specific for TRPC proteins blocks increases in [Ca2+]i and CCE caused by acute hypoxia; 2) the effects of chronic hypoxia on TRPC expression, CCE and pulmonary vascular resistance; 3) whether SOCC antagonists reverse hypoxia-induced changes in baseline [Ca2+]i, CCE and pulmonary vascular reactivity; 4) if treatment with specific TRPC siRNA prevents changes in basal [Ca2+]i and CCE in PASMCs induced by prolonged hypoxia and 5) whether the transcription factor, HIF-1is responsible for hypoxia-induced upregulation of TRPC expression. We hope that our results will provide new mechanistic information and that elucidating the factors involved in this process will lead to improved methods of pharmacological prevention and treatment of this lethal complication of chronic lung disease.
