Depolarization‐Dependent Contraction Increase after Birth and Preservation following Long‐Term Hypoxia in Sheep Pulmonary Arteries

Abstract

Membrane depolarization is critical to pulmonary arterial (PA) contraction. Both L-type Ca2+ channels (CaL) and Rho-kinase are important signaling components of this process and mitochondrial and non-mitochondrial generated superoxides can be part of the signaling process. Maturation and long-term hypoxia (LTH) each can modify depolarization-dependent contraction and the role of superoxides. By the use of wire myography, we tested the hypothesis that maturation and LTH increase pulmonary arterial reactivity to high-K+-induced membrane depolarization through enhancements in the importance of CaLand Rho-kinase-dependent pathways. The data show that maturation, but not LTH, increases contraction to 125 mM KCl (high-K+) without altering the EC50. High-K+-dependent contraction was inhibited to a similar extent in fetal and adult PA by multiple CaL blockers, including 10 μM diltiazem, 10 μM verapamil, and 10 μM nifedipine. Postnatal maturation increased the role for 10 μM nifedipine-sensitive CaL, and decreased that for 10 μM Y-27632-sensitive Rho-kinase. In all groups, the combination of nifedipine and Y-27632 effectively inhibited high-K+ contraction. Tempol (3 mM) but not 100 μM apocynin slightly reduced contraction in arteries from fetal hypoxic and adult normoxic and hypoxic sheep, indicating a limited role for non-mitochondrial derived superoxide to high-K+-induced contraction. Western immunoblot for alpha smooth muscle actin indicated small increases in relative abundance in the adult. The data suggest that while CaL therapies more effectively vasodilate PA in adults and rho-kinase therapies are more effective in newborns, combination therapies would provide greater efficacy in both young and mature patients regardless of normoxic or hypoxic conditions.

Developmental acceleration of bradykinin-dependent relaxation by prenatal chronic hypoxia impedes normal development after birth

“…Furthermore, PKG inhibits RhoA activity, which is a key modulator of the Rho kinase pathway (27,49). This is directly relevant to the current studies, as we previously showed that Rho kinase accounts for roughly half of the arterial tension due to serotonin or membrane depolarization in pulmonary arteries of sheep (10,68). Future studies are therefore needed to discriminate the interaction between PKG and various pathways.…”

Muscarinic Receptor Activation Affects Pulmonary Artery Contractility in Sheep: The Impact of Maturation and Chronic Hypoxia on Endothelium-Dependent and Endothelium-Independent Function

“…Many of the experimental procedures and protocols are based on our previously published methodology Papamatheakis et al, 2011Papamatheakis et al, , 2012Papamatheakis et al, , 2013Blood et al, 2013). All animals were purchased from Nebeker Ranch (Lancaster, CA; 720 m elevation).…”

“…The normoxic control animals were brought to LLU (353 m elevation; ewe arterial PaO 2 = 95 -5 Torr) for experimental study. The chronic hypoxic experimental animals were acclimatized to high altitude (3801 m elevation, ewe PaO 2 = 60 -5 Torr) at the Barcroft Laboratory, White Mountain Research Station (Bishop, CA), for *110 days (Longo et al, 1996;Goyal et al, 2011;Papamatheakis et al, 2012). …”

“…In vasodilation studies, special care was taken during arterial isolation and wire mounting to avoid endothelial disruption. In experiments in which pulmonary arterial contraction to muscarinic stimulation was studied, the endothelium was intentionally disrupted by inserting a wire through the arterial lumen and gently rotating, as previously described Papamatheakis et al, 2012;Blood et al, 2013).…”

“…Our previously published Ca 2+ signaling and vasoreactivity findings led us to postulate that CH and maturation each would differentially modify mAChR-dependent pulmonary arterial vasoreactivity (Xue et al, 2008;Goyal et al, 2011;Papamatheakis et al, 2011Papamatheakis et al, , 2012Hadley et al, 2012;Blood et al, 2013). The potential loss of endothelium-dependent pulmonary arterial vasodilation and acetylcholine-dependent vasoconstriction led us to test the hypothesis that mAChR vasoconstrictor responses predominate in the pulmonary vasculature of high-altitude sheep compared to normoxic controls.…”