Chronic Hypoxia–Induced Upregulation of Store-Operated and Receptor-Operated Ca ²⁺ Channels in Pulmonary Arterial Smooth Muscle Cells

Abstract

Abstract-Chronic hypoxic pulmonary hypertension is associated with profound vascular remodeling and alterations in Ca 2ϩ homeostasis in pulmonary arterial smooth muscle cells (PASMCs). Recent studies show that transient receptor potential (TRPC) genes, which encode store-operated and receptor-operated cation channels, play important roles in Ca 2ϩ regulation and cell proliferation. However, the influence of chronic hypoxia on TRPC channels has not been determined. Here we compared TRPC expression, and store-and receptor-operated Ca 2ϩ entries in PASMCs of normoxic and chronic hypoxic rats. Reverse-transcription polymerase chain reaction (RT-PCR), Western blot, and immunostaining showed consistently that TRPC1, TRPC3, and TRPC6 were expressed in intralobar pulmonary arteries (PAs) and PASMCs. Application of 1-oleoyl-2-acetyl-sn-glycerol (OAG) to directly activate receptor-operated channels, or thapsigargin to deplete Ca 2ϩ stores, caused dramatic increase in cation entry measured by Mn 2ϩ quenching of fura-2 and by Ca 2ϩ transients. OAG-induced responses were Ϸ700-fold more resistant to La 3ϩ inhibition than thapsigargin-induced responses. siRNA knockdown of TRPC1 and TRPC6 specifically attenuated thapsigargin-and OAG-induced cation entries, respectively, indicating that TRPC1 mediates store-operated entry and TRPC6 mediates receptor-operated entry. In hypoxic PAs, there were 2-to 3-fold increases in TRPC1 and TRPC6 expression. They were accompanied by significant increases in basal, OAG-induced, and thapsigargin-induced cation entries in hypoxic PASMCs. Moreover, removal of Ca 2ϩ or inhibition of store-operated Ca 2ϩ entry with La 3ϩ and SK&F-96365 reversed the elevated basal [Ca 2ϩ ] i in PASMCs and vascular tone in PAs of chronic hypoxic animals, but nifedipine had minimal effects. Our results for the first time to our knowledge show that both store-and receptor-operated channels of PASMCs are upregulated by chronic hypoxia and contribute to the enhanced vascular tone in hypoxic pulmonary hypertension. Key Words: pulmonary hypertension Ⅲ transient receptor potential channels Ⅲ store-operated Ca 2ϩ channels Ⅲ receptor-operated Ca 2ϩ channels P rolonged exposure to alveolar hypoxia causes pulmonary hypertension with profound vascular remodeling and increase in vasomotor tone. The increase in vascular tone is in part attributable to alterations in vasoconstricting and vasorelaxing influences imposed by the endothelially derived and circulating factors. 1 Recent evidence indicates that chronic hypoxia also causes intrinsic changes in ionic balance and Ca 2ϩ homeostasis in pulmonary arterial smooth muscle cells (PASMCs), including membrane depolarization, elevation in resting [Ca 2ϩ ] i , and changes in electrophysiological and Ca 2ϩ responses to vasoconstrictors and vasodilators. [2][3][4][5][6] The mechanism for alteration in Ca 2ϩ homeostasis in hypoxic PASMCs is controversial. Previous studies found significant suppression of voltage-gated K ϩ (K V ) currents and K V channel expression in PASMCs isolated fr...

Effects of chronic exposure to cigarette smoke on canonical transient receptor potential expression in rat pulmonary arterial smooth muscle

“…The upregulation of TRPC expression can increase the activity of SOCC, relating to enhanced SOCE, and also promotes the proliferation of PASMCs (5, 42). Additionally, upregulated TRPC1 and TRPC6 expression and elevated SOCE are also observed under chronic hypoxia in rat distal PASMCs (9,32). In the present study, similar to the effects of chronic hypoxia, chronic exposure to CS promotes the [Ca 2ϩ ] i and SOCE, as well as upregulated TRPC1 and TRPC6 expression in PASMCs.…”

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