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This Article Full Text Full Text (PDF) All Versions of this Article: 292/3/L654    most recent 00229.2006v1 Alert me when this article is cited Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in ISI Web of Science Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via Google Scholar Google Scholar Articles by Hinton, M. Articles by Dakshinamurti, S. Search for Related Content PubMed PubMed Citation Articles by Hinton, M. Articles by Dakshinamurti, S.

Thromboxane hypersensitivity in hypoxic pulmonary artery myocytes: altered TP receptor localization and kinetics

Departments of ¹Physiology and ²Pediatrics, University of Manitoba, Biology of Breathing Group, and ³Manitoba Institute of Child Health, Winnipeg, Manitoba, Canada

Submitted 20 June 2006 ; accepted in final form 3 November 2006

Hypoxia-induced neonatal persistent pulmonary hypertension (PPHN) is characterized by sustained vasospasm and increased thromboxane (TxA₂)-to-prostacyclin ratio. We previously demonstrated that moderate hypoxia induces myocyte TxA₂ hypersensitivity. Here, we examined TxA₂ prostanoid receptor (TP-R) localization and kinetics following hypoxia to determine the mechanism of hypoxia-induced TxA₂ hypersensitivity. Primary cultured neonatal pulmonary artery myocytes were exposed to 10% O₂ (hypoxic myocytes; HM) or 21% O₂ (normoxic myocytes; NM) for 3 days. PPHN was induced in neonatal piglets by in vivo exposure to 10% FI_O2 for 3 days. TP-R was studied in whole lung sections from pigs with hypoxic PPHN- and age-matched controls; intracellular localization was studied by immunocytochemistry. TP-R affinity was studied in cultured myocytes by saturation binding kinetics using ³H-SQ-29548 and competitive binding kinetics by coincubation with U-46619. Phosphorylation and coupling were examined in immunoprecipitated TP-R. We report distal propagation of TP-R expression in PPHN, extending to pulmonary arteries <50 µm. In HM, intracellular TP-R moves towards the perinuclear region, mirroring a change in endoplasmic reticulum (ER) morphology. TP-R kinetics also alter in HM membranes, with decreased K_d and B_max (maximal binding sites). Additionally, in hypoxia, ³H-SQ-29548 is displaced at lower concentration of U-46619 than in normoxia, suggesting increased agonist affinity. Phosphorylation of serine residues on HM TP-R was significantly decreased compared with NM; this difference correlated with increased G_q coupling in hypoxia and was ablated by incubation with PKA. We conclude that the TP-R is normally desensitized in the neonatal pulmonary circuit by PKA-mediated regulatory phosphorylation, decreasing ligand affinity and coupling to G_q; this protection is lost following hypoxic exposure. Also, the appearance of TP-R in resistance arteries after development of hypoxic PPHN may contribute to increased pulmonary arterial pressure.

smooth muscle; persistent pulmonary hypertension of the newborn; Scatchard analysis