title: Type-3 ryanodine receptors mediate hypoxia-, but not neurotransmitter-induced calcium release and contraction in pulmonary artery smooth muscle cells.
abstract: In this study we examined the expression of RyR subtypes and the role of RyRs in neurotransmitter- and hypoxia-induced Ca2+ release and contraction in pulmonary artery smooth muscle cells (PASMCs). Under perforated patch clamp conditions, maximal activation of RyRs with caffeine or inositol triphosphate receptors (IP3Rs) with noradrenaline induced equivalent increases in [Ca2+]i and Ca2+-activated Cl- currents in freshly isolated rat PASMCs. Following maximal IP3-induced Ca2+ release, neither caffeine nor chloro-m-cresol induced a response, whereas prior application of caffeine or chloro-m-cresol blocked IP3-induced Ca2+ release. In cultured human PASMCs, which lack functional expression of RyRs, caffeine failed to affect ATP-induced increases in [Ca2+]i in the presence and absence of extracellular Ca2+. The RyR antagonists ruthenium red, ryanodine, tetracaine, and dantrolene greatly inhibited submaximal noradrenaline- and hypoxia-induced Ca2+ release and contraction in freshly isolated rat PASMCs, but did not affect ATP-induced Ca2+ release in cultured human PASMCs. Real-time quantitative RT-PCR and immunofluorescence staining indicated similar expression of all three RyR subtypes (RyR1, RyR2, and RyR3) in freshly isolated rat PASMCs. In freshly isolated PASMCs from RyR3 knockout (RyR3-/-) mice, hypoxia-induced, but not submaximal noradrenaline-induced, Ca2+ release and contraction were significantly reduced. Ruthenium red and tetracaine can further inhibit hypoxic increase in [Ca2+]i in RyR3-/- mouse PASMCs. Collectively, our data suggest that (a) RyRs play an important role in submaximal noradrenaline- and hypoxia-induced Ca2+ release and contraction; (b) all three subtype RyRs are expressed; and (c) RyR3 gene knockout significantly inhibits hypoxia-, but not submaximal noradrenaline-induced Ca2+ and contractile responses in PASMCs.
authors.affiliation: Center for Cardiovascular Sciences, Neuroscience, and Neuropharmacology, Albany Medical College, NY 12208, USA.
authors.forename: Yun-Min
authors.initials: YM
authors.lastname: Zheng
authors.forename: Qing-Song
authors.initials: QS
authors.lastname: Wang
authors.forename: Rakesh
authors.initials: R
authors.lastname: Rathore
authors.forename: Wan-Hui
authors.initials: WH
authors.lastname: Zhang
authors.forename: Joseph E
authors.initials: JE
authors.lastname: Mazurkiewicz
authors.forename: Vincenzo
authors.initials: V
authors.lastname: Sorrentino
authors.forename: Harold A
authors.initials: HA
authors.lastname: Singer
authors.forename: Michael I
authors.initials: MI
authors.lastname: Kotlikoff
authors.forename: Yong-Xiao
authors.initials: YX
authors.lastname: Wang
mesh_terms: D000818:Animals; D002110:Caffeine / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D015687:Cell Hypoxia / Q000502:physiology*; D002478:Cells, Cultured; D004305:Dose-Response Relationship, Drug; D009119:Muscle Contraction / Q000187:drug effects / Q000502:physiology*; D032389:Myocytes, Smooth Muscle / Q000187:drug effects / Q000502:physiology*; D018377:Neurotransmitter Agents / Q000378:metabolism*; D011651:Pulmonary Artery / Q000187:drug effects / Q000502:physiology*; D051381:Rats; D017207:Rats, Sprague-Dawley; D019837:Ryanodine Receptor Calcium Release Channel / Q000187:drug effects / Q000378:metabolism*
chemical_list: D018377:Neurotransmitter Agents; D019837:Ryanodine Receptor Calcium Release Channel; D002110:Caffeine; D002118:Calcium