Electrophysiological modelling of pulmonary artery smooth muscle cells in the rabbits—Special consideration to the generation of hypoxic pulmonary vasoconstriction

Hypoxic pulmonary vasoconstriction is caused by a rise in cytosolic Ca2+ ([Ca2+]cyt) in pulmonary artery smooth muscle cells (PASMC) via multiple mechanisms. PASMC consist of heterogeneous phenotypes defined by contractility, proliferation, and apoptosis as well as by differences in expression and function of various genes. In rat PASMC, hypoxia-mediated decrease in voltage-gated K+ (Kv) currents ( IK(V)) and increase in [Ca2+]cyt were not uniformly distributed in all PASMC tested. Acute hypoxia decreased IK(V) and increased [Ca2+]cyt in ∼46% and ∼53% of PASMC, respectively. Using combined techniques of single-cell RT-PCR and patch clamp, we show here that mRNA expression level of Kv1.5 in hypoxia-sensitive PASMC (in which hypoxia reduced IK(V)) was much greater than in hypoxia-insensitive cells (in which hypoxia negligibly affected IK(V)). These results demonstrate that 1) different PASMC express different Kv channel α- and β-subunits, and 2) the sensitivity of a PASMC to acute hypoxia partially depends on the expression level of Kv1.5 channels; hypoxia reduces whole-cell IK(V) only in PASMC that express high level of Kv1.5. In addition, the acute hypoxia-mediated changes in [Ca2+]cyt also vary in different PASMC. Hypoxia increases [Ca2+]cyt only in 34% of cells tested, and the different sensitivity of [Ca2+]cyt to hypoxia was not related to the resting [Ca2+]cyt. An intrinsic mechanism within each individual cell may be involved in the heterogeneity of hypoxia-mediated effect on [Ca2+]cyt in PASMC. These data suggest that the heterogeneity of PASMC may partially be related to different expression levels and functional sensitivity of Kv channels to hypoxia and to differences in intrinsic mechanisms involved in regulating [Ca2+]cyt.

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Diphenyleneiodonium inhibits both potassium and calcium currents in isolated pulmonary artery smooth muscle cells

Journal of Applied Physiology ◽

10.1152/jappl.1994.76.6.2611 ◽

1994 ◽

Vol 76 (6) ◽

pp. 2611-2615 ◽

Cited By ~ 61

Author(s):

E. K. Weir ◽

C. N. Wyatt ◽

H. L. Reeve ◽

J. Huang ◽

S. L. Archer ◽

...

Keyword(s):

Hydrogen Peroxide ◽

Smooth Muscle ◽

Nadph Oxidase ◽

Smooth Muscle Cells ◽

Hypoxic Pulmonary Vasoconstriction ◽

Muscle Cells ◽

Calcium Currents ◽

Pulmonary Vasoconstriction ◽

Pulmonary Artery Smooth Muscle ◽

Isolated Pulmonary Artery

Diphenyleneiodonium (DPI) blocks hypoxic vasoconstriction in the pulmonary vasculature. Because one of the actions of DPI is the inhibition of NADPH oxidase, this has led to the suggestion that NADPH oxidase acts as an oxygen tension sensor in pulmonary smooth muscle cells. We investigated the effects of DPI on potassium and calcium currents in freshly isolated pulmonary artery smooth muscle cells by using whole cell patch-clamp recordings, since these ionic currents are known to be involved in hypoxic pulmonary vasoconstriction. DPI (3 and 10 microM) reversibly inhibited potassium currents, and in its presence, residual currents appeared markedly more transient than under control conditions. The actions of DPI could not be reversed by 4.4 mM hydrogen peroxide, the product of NADPH oxidase. Calcium channel currents were also reversibly inhibited by 3 microM DPI. Thus DPI is a nonselective blocker of ionic channels in pulmonary smooth muscle cells, and its mechanism of action does not appear to involve inhibition of hydrogen peroxide formation. The ability of DPI to block calcium currents can explain its inhibition of hypoxic pulmonary vasoconstriction.

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A role for receptor-operated Ca2+ entry in human pulmonary artery smooth muscle cells in response to hypoxia

Physiological Research ◽

10.33549/physiolres.931875 ◽

2010 ◽

pp. 909-918 ◽

Cited By ~ 1

Author(s):

C Tang ◽

WK To ◽

F Meng ◽

Y Wang ◽

Y Gu

Keyword(s):

Smooth Muscle ◽

Pulmonary Artery ◽

Smooth Muscle Cells ◽

Hypoxic Pulmonary Vasoconstriction ◽

Muscle Cells ◽

Trpc Channels ◽

Hypoxia Response ◽

Compound C ◽

Pulmonary Artery Smooth Muscle ◽

Human Pulmonary Artery

Hypoxic pulmonary vasoconstriction (HPV) is an important homeostatic mechanism in which increases of [Ca2+]i are primary events. In this study, primary cultured, human pulmonary artery smooth muscle cells (hPASMC) were used to examine the role of TRPC channels in mediating [Ca2+]i elevations during hypoxia. Hypoxia (PO2 about 20 mm Hg) evoked a transient [Ca2+]i elevation that was reduced by removal of extracellular calcium. Nifedipine and verapamil, blockers of voltage-gated calcium channels (VGCCs), attenuated the hypoxia-induced [Ca2+]i elevation by about 30 %, suggesting the presence of alternate Ca2+ entry pathways. Expression of TRPC1 and TRPC6 in hPASMC were found by RT-PCR and confirmed by Western blot analysis. Antagonists for TRPC, 2APB and SKF96365, significantly reduced hypoxia-induced [Ca2+]i elevation by almost 60 %. Both TRPC6 and TRPC1 were knocked down by siRNA, the loss of TRPC6 decreased hypoxic response down to 21 % of control, whereas the knockdown of TRPC1 reduced the hypoxia response to 85 %, suggesting that TRPC6 might play a central role in mediating hypoxia response in hPASMC. However, blockade of PLC pathway caused only small inhibition of the hypoxia response. In contrast, AICAR, the agonist of AMP-activated kinase (AMPK), induced a gradual [Ca2+]i elevation, whereas compound C, an antagonist of AMPK, almost abolished the hypoxia response. However, co-immunoprecipitation revealed that AMPKα was not colocalized with TRPC6. Our data supports a role for TRPC6 in mediation of the [Ca2+]i elevation in response to hypoxia in hPASMC and suggests that this response may be linked to cellular energy status via an activation of AMPK.

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PLCγ1-PKCε-IP3R1 signaling plays an important role in hypoxia-induced calcium response in pulmonary artery smooth muscle cells

AJP Lung Cellular and Molecular Physiology ◽

10.1152/ajplung.00243.2017 ◽

2018 ◽

Vol 314 (5) ◽

pp. L724-L735 ◽

Cited By ~ 9

Author(s):

Vishal R. Yadav ◽

Tengyao Song ◽

Lin Mei ◽

Leroy Joseph ◽

Yun-Min Zheng ◽

...

Keyword(s):

Smooth Muscle ◽

Pulmonary Artery ◽

Smooth Muscle Cells ◽

Molecular Mechanisms ◽

Acute Hypoxia ◽

Muscle Cells ◽

Mesenteric Arteries ◽

Pulmonary Vasoconstriction ◽

Expression Activity ◽

Pulmonary Artery Smooth Muscle

Hypoxia-induced pulmonary vasoconstriction (HPV) is attributed to an increase in intracellular Ca2+ concentration ([Ca2+]i) in pulmonary artery smooth muscle cells (PASMCs). We have reported that phospholipase C-γ1 (PLCγ1) plays a significant role in the hypoxia-induced increase in [Ca2+]i in PASMCs and attendant HPV. In this study, we intended to determine molecular mechanisms for hypoxic Ca2+ and contractile responses in PASMCs. Our data reveal that hypoxic vasoconstriction occurs in pulmonary arteries, but not in mesenteric arteries. Hypoxia caused a large increase in [Ca2+]i in PASMCs, which is diminished by the PLC inhibitor U73122 and not by its inactive analog U73433 . Hypoxia augments PLCγ1-dependent inositol 1,4,5-trisphosphate (IP3) generation. Exogenous ROS, hydrogen peroxide (H2O2), increases PLCγ1 phosphorylation at tyrosine-783 and IP3 production. IP3 receptor-1 (IP3R1) knock-down remarkably diminishes hypoxia- or H2O2-induced increase in [Ca2+]i. Hypoxia or H2O2 increases the activity of IP3Rs, which is significantly reduced in protein kinase C-ε (PKCε) knockout PASMCs. A higher PLCγ1 expression, activity, and basal [Ca2+]i are found in PASMCs, but not in mesenteric artery smooth muscle cells from mice exposed to chronic hypoxia (CH) for 21 days. CH enhances H2O2- and ATP-induced increase in [Ca2+]i in PASMCs and PLC-dependent, norepinephrine-evoked pulmonary vasoconstriction. In conclusion, acute hypoxia uniquely causes ROS-dependent PLCγ1 activation, IP3 production, PKCε activation, IP3R1 opening, Ca2+ release, and contraction in mouse PASMCs; CH enhances PASM PLCγ1 expression, activity, and function, playing an essential role in pulmonary hypertension in mice.

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