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Timothy J McMahon

Publications and source records attributed to Timothy J McMahon.

14 recordsLinked to original sources
PubMed2006

Excitation-contraction coupling in airway smooth muscle.

Excitation-contraction (EC) coupling in striated muscles is mediated by the cardiac or skeletal muscle isoform of voltage-dependent L-type Ca(2+) channel (Ca(v)1.2 and Ca(v)1.1, respectively) that senses a depolarization of the cell membrane, and in response, activates its corresponding isoform of intracellular Ca(2+) release channel/ryanodine receptor (RyR) to release stored Ca(2+), thereby initiating muscle contraction. Specifically, in cardiac muscle following cell membrane depolarization, Ca(v)1.2 activates cardiac RyR (RyR2) through an influx of extracellular Ca(2+). In contrast, in skeletal muscle, Ca(v)1.1 activates skeletal muscle RyR (RyR1) through a direct physical coupling that negates the need for extracellular Ca(2+). Since airway smooth muscle (ASM) expresses Ca(v)1.2 and all three RyR isoforms, we examined whether a cardiac muscle type of EC coupling also mediates contraction in this tissue. We found that the sustained contractions of rat ASM preparations induced by depolarization with KCl were indeed partially reversed ( approximately 40%) by 200 mum ryanodine, thus indicating a functional coupling of L-type channels and RyRs in ASM. However, KCl still caused transient ASM contractions and stored Ca(2+) release in cultured ASM cells without extracellular Ca(2+). Further analyses of rat ASM indicated that this tissue expresses as many as four L-type channel isoforms, including Ca(v)1.1. Moreover, Ca(v)1.1 and RyR1 in rat ASM cells have a similar distribution near the cell membrane in rat ASM cells and thus may be directly coupled as in skeletal muscle. Collectively, our data implicate that EC-coupling mechanisms in striated muscles may also broadly transduce diverse smooth muscle functions.

Amino Acid Sequence↗
PubMed2006

Extrapulmonary effects of inhaled nitric oxide: role of reversible S-nitrosylation of erythrocytic hemoglobin.

Early applications of inhaled nitric oxide (iNO), typically in the treatment of diseases marked by acute pulmonary hypertension, were met by great enthusiasm regarding the purported specificity of iNO: vasodilation by iNO was specific to the lung (without a change in systemic vascular resistance), and within the lung, NO activity was said to be confined spatially and temporally by Hb within the vascular lumen. Underlying these claims were classical views of NO as a short-lived paracrine hormone that acts largely through the heme groups of soluble guanylate cyclase, and whose potential activity is terminated on encountering the hemes of red blood cell (RBC) Hb. These classical views are yielding to a broader paradigm, in which NO-related signaling is achieved through redox-related NO adducts that endow NO synthase products with the ability to act at a distance in space and time from NO synthase itself. Evidence supporting the biological importance of such stable NO adducts is probably strongest for S-nitrosothiols (SNOs), in which NO binds to critical cysteine residues in proteins or peptides. The circulating RBC is a major SNO reservoir, and RBC Hb releases SNO-related bioactivity peripherally on O2 desaturation. These new paradigms describing NO transport also provide a plausible mechanistic understanding of the increasingly recognized peripheral effects of inhaled NO. An explanation for the peripheral actions of inhaled NO is discussed here, and the rationale and results of attempts to exploit the "NO delivery" function of the RBC are reviewed.

Administration, Inhalation↗
PubMed2005

A nitric oxide processing defect of red blood cells created by hypoxia: deficiency of S-nitrosohemoglobin in pulmonary hypertension.

The mechanism by which hypoxia [low partial pressure of O(2) (pO(2))] elicits signaling to regulate pulmonary arterial pressure is incompletely understood. We considered the possibility that, in addition to its effects on smooth muscle, hypoxia may influence pulmonary vascular tone through an effect on RBCs. We report that exposure of native RBCs to sustained hypoxia is accompanied by a buildup of heme iron-nitrosyl (FeNO) species that are deficient in pO(2-)governed intramolecular transfer of NO to cysteine thiol, yielding a deficiency in the vasodilator S-nitrosohemoglobin (SNO-Hb). S-nitrosothiol (SNO)-deficient RBCs produce impaired vasodilator responses in vitro and exaggerated pulmonary vasoconstrictor responses in vivo and are defective in oxygenating the blood. RBCs from hypoxemic patients with elevated pulmonary arterial pressure (PAP) exhibit a similar FeNO/SNO imbalance and are thus deficient in pO(2)-coupled vasoregulation. Chemical restoration of SNO-Hb levels in both animals and patients restores the vasodilator activity of RBCs, and this activity is associated with improved oxygenation and lower PAPs.

Animals↗
PubMed2005

S-nitrosoglutathione inhibits alpha1-adrenergic receptor-mediated vasoconstriction and ligand binding in pulmonary artery.

Endogenous nitric oxide donor compounds (S-nitrosothiols) contribute to low vascular tone by both cGMP-dependent and -independent pathways. We have reported that S-nitrosoglutathione (GSNO) inhibits 5-hydroxytryptamine (5-HT)-mediated pulmonary vasoconstriction via a cGMP-independent mechanism likely involving S-nitrosylation of its G protein-coupled receptor (GPCR) system. Because catecholamines, like 5-HT, constrict lung vessels via a GPCR coupled to G(q), we hypothesized that S-nitrosothiols modify the alpha1-adrenergic GPCR system to inhibit pulmonary vasoconstriction by receptor agonists, e.g., phenylephrine (PE). Rat pulmonary artery rings were pretreated for 30 min with and without an S-nitrosothiol, either GSNO or S-nitrosocysteine (CSNO), and constricted with sequential concentrations of PE (10(-8)-10(-6) M). Effective cGMP-dependence was tested in rings pretreated with soluble guanylate cyclase inhibitors {either 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one (ODQ) or LY-83583} or G kinase inhibitor (KT-5823), and a thiol reductant [dithiothreitol (DTT)] was used to test reversibility of S-nitrosylation. Both S-nitrosothiols attenuated the PE dose response. The GSNO effect was not prevented by LY-83583, ODQ, or KT-5823, indicating cGMP independence. GSNO inhibition was reversed by DTT, consistent with S-nitrosylation or other GSNO-mediated cysteine modifications. In CSNO-treated lung protein, the alpha1-adrenergic receptor was shown to undergo S-nitrosylation in vitro using a biotin switch assay. Studies of alpha1-adrenergic receptor subtype expression and receptor density by saturation binding with 125I-HEAT showed that GSNO decreased alpha1-adrenergic receptor density but did not alter affinity for antagonist or agonist. These data demonstrate a novel cGMP-independent mechanism of reversible alpha1-adrenergic receptor inhibition by S-nitrosothiols.

Adrenergic alpha-Agonists↗
PubMed2005

Oxygen regulation of tumor perfusion by S-nitrosohemoglobin reveals a pressor activity of nitric oxide.

In erythrocytes, S-nitrosohemoglobin (SNO-Hb) arises from S-nitrosylation of oxygenated hemoglobin (Hb). It has been shown that SNO-Hb behaves as a nitric oxide (NO) donor at low oxygen tensions. This property, in combination with oxygen transport capacity, suggests that SNO-Hb may have unique potential to reoxygenate hypoxic tissues. The present study was designed to test the idea that the allosteric properties of SNO-Hb could be manipulated to enhance oxygen delivery in a hypoxic tumor. Using Laser Doppler flowmetry, we showed that SNO-Hb infusion to animals breathing 21% O2 reduced tumor perfusion without affecting blood pressure and heart rate. Raising the pO2 (100% O2) slowed the release of NO bioactivity from SNO-Hb (ie, prolonged the plasma half-life of the SNO in Hb), preserved tumor perfusion, and raised the blood pressure. In contrast, native Hb reduced both tumor perfusion and heart rate independently of the oxygen concentration of the inhaled gas, and did not elicit hypertensive effects. Window chamber (to image tumor arteriolar reactivity in vivo) and hemodynamic measurements indicated that the preservation of tissue perfusion by micromolar concentrations of SNO-Hb is a composite effect created by reduced peripheral vascular resistance and direct inhibition of the baroreceptor reflex, leading to increased blood pressure. Overall, these results indicate that the properties of SNO-Hb are attributable to allosteric control of NO release by oxygen in central as well as peripheral issues.

Animals↗
PubMed2005

A novel inhaled organic nitrate that affects pulmonary vascular tone in a piglet model of hypoxia-induced pulmonary hypertension.

Persistent pulmonary hypertension of the newborn is characterized by elevated pulmonary vascular resistance after birth leading to right-to-left shunting and systemic arterial hypoxemia. Inhaled nitric oxide (NO) is effective in reducing the need for extracorporeal membrane oxygenation, but it has potential toxicities, especially in an oxygen-rich environment. A number of other NO-based molecules have been given by inhalation, but their structure-function relationships have not been established. Recent studies have raised the idea that toxic and beneficial properties can be separated. We synthesized a novel organic nitrate [ethyl nitrate (ENO2)], tested it in vitro, and administered it to hypoxic piglets. ENO2 lowered pulmonary artery pressure and raised the Po2 in arterial blood but did not alter systemic vascular resistance or methemoglobin levels. In addition, we tested the effect of ENO2 in the presence of the thiol glutathione, both in vivo and in vitro, and found its action to be enhanced. Although ENO2 is less potent than inhaled NO on a dose-equivalency basis, pretreatment of hypoxic animals with glutathione, which may be depleted in injured lungs, led to a markedly enhanced effect (largely mitigating the difference in potency). These results suggest that ENO2 may hold promise as a safe alternative to NO, particularly in hypoxemic conditions characterized by thiol depletion.

Administration, Inhalation↗
PubMed2005

Redox activation of intracellular calcium release channels (ryanodine receptors) in the sustained phase of hypoxia-induced pulmonary vasoconstriction.

Hypoxia-induced pulmonary vasoconstriction (HPV) is an important adaptive process that remains incompletely understood. In preconstricted rat pulmonary arteries (inner diameter, 250 to 400 microm), hypoxia (pO2 approximately 10 mm Hg) induces an initial transient phase and a more slowly developing sustained phase of vasoconstriction. Since the release of calcium ions (Ca2+) from intracellular stores by redox-sensitive intracellular Ca2+ release channels known as ryanodine receptors (RyRs) in pulmonary arterial smooth-muscle cells (PASMCs) may play a role in HPV, and considerable evidence now supports that levels of reactive oxygen species (ROS) are paradoxically increased in PASMC under hypoxia, we investigated whether redox activation of RyRs by ROS may transduce HPV. By reverse transcriptase-polymerase chain reaction, we found that all three RyR isoforms are expressed in rat pulmonary arteries and in PASMCs. The sustained phase, but not the transient phase, of HPV can be prevented by pretreating pulmonary arteries with RyR inhibitors ryanodine (200 micromol/L) or dantrolene (50 micromol/L). The addition of dantrolene, ryanodine or the thiol-reducing agent dithiothreitol (1 mmol/L) during the sustained phase of HPV reversed the hypoxic vasoconstriction. In contrast, the superoxide scavenger nitroblue tetrazolium (500 nmol/L) prevented further hypoxic pulmonary vasoconstriction during the sustained phase of HPV but did not reverse it. Taken together, our data suggest that redox activation of RyRs by ROS has an important role in transducing the sustained contraction of pulmonary arteries under hypoxia.

Animals↗
PubMed2004

Essential roles of S-nitrosothiols in vascular homeostasis and endotoxic shock.

The current perspective of NO biology is formulated predominantly from studies of NO synthesis. The role of S-nitrosothiol (SNO) formation and turnover in governing NO-related bioactivity remains uncertain. We generated mice with a targeted gene deletion of S-nitrosoglutathione reductase (GSNOR), and show that they exhibit substantial increases in whole-cell S-nitrosylation, tissue damage, and mortality following endotoxic or bacterial challenge. Further, GSNOR(-/-) mice have increased basal levels of SNOs in red blood cells and are hypotensive under anesthesia. Thus, SNOs regulate innate immune and vascular function, and are cleared actively to ameliorate nitrosative stress. Nitrosylation of cysteine thiols is a critical mechanism of NO function in both health and disease.

Alcohol Dehydrogenase↗
PubMed2003

S-nitrosylation in health and disease.

S-nitrosylation is a ubiquitous redox-related modification of cysteine thiol by nitric oxide (NO), which transduces NO bioactivity. Accumulating evidence suggests that the products of S-nitrosylation, S-nitrosothiols (SNOs), play key roles in human health and disease. In this review, we focus on the reaction mechanisms underlying the biological responses mediated by SNOs. We emphasize reactions that can be identified with complex (patho)physiological responses, and that best rationalize the observed increase or decrease in specific classes of SNOs across a spectrum of disease states. Thus, changes in the levels of various SNOs depend on specific defects in both enzymatic and non-enzymatic mechanisms of nitrosothiol formation, processing and degradation. An understanding of these mechanisms is crucial for the development of an integrated model of NO biology, and for effective treatment of diseases associated with dysregulation of NO homeostasis.

Copper↗
PubMed2002

Nitric oxide in the human respiratory cycle.

Interactions of nitric oxide (NO) with hemoglobin (Hb) could regulate the uptake and delivery of oxygen (O(2)) by subserving the classical physiological responses of hypoxic vasodilation and hyperoxic vasconstriction in the human respiratory cycle. Here we show that in in vitro and ex vivo systems as well as healthy adults alternately exposed to hypoxia or hyperoxia (to dilate or constrict pulmonary and systemic arteries in vivo), binding of NO to hemes (FeNO) and thiols (SNO) of Hb varies as a function of HbO(2) saturation (FeO(2)). Moreover, we show that red blood cell (RBC)/SNO-mediated vasodilator activity is inversely proportional to FeO(2) over a wide range, whereas RBC-induced vasoconstriction correlates directly with FeO(2). Thus, native RBCs respond to changes in oxygen tension (pO2) with graded vasodilator and vasoconstrictor activity, which emulates the human physiological response subserving O(2) uptake and delivery. The ability to monitor and manipulate blood levels of NO, in conjunction with O(2) and carbon dioxide, may therefore prove useful in the diagnosis and treatment of many human conditions and in the development of new therapies. Our results also help elucidate the link between RBC dyscrasias and cardiovascular morbidity.

Erythrocytes↗
PubMed2002

Pulmonary vasoconstriction by serotonin is inhibited by S-nitrosoglutathione.

Nitric oxide (NO) functions as an endothelium-derived relaxing factor by activating guanylate cyclase to increase cGMP levels. However, NO and related species may also regulate vascular tone by cGMP-independent mechanisms. We hypothesized that naturally occurring NO donors could decrease the pulmonary vascular response to serotonin (5-HT) in the intact lung through chemical interactions with 5-HT(2) receptors. In isolated rabbit lung preparations and isolated pulmonary artery (PA) rings, 50-250 microM S-nitrosoglutathione (GSNO) inhibited the response to 0.01-10 microM 5-HT. The vasoconstrictor response to 5-HT was mediated by 5-HT(2) receptors in the lung, since it could be blocked completely by the selective inhibitor ketanserin (10 microM). GSNO inhibited the response to 5-HT by 77% in intact lung and 82% in PA rings. In PA rings, inhibition by GSNO could be reversed by treatment with the thiol reductant dithiothreitol (10 mM). 3-Morpholinosydnonimine (100-500 microM), which releases NO and O simultaneously, also blocked the response to 5-HT. Its chemical effects, however, were distinct from those of GSNO, because 5-HT-mediated vasoconstriction was not restored in isolated rings by dithiothreitol. In the intact lung, neither NO donor altered the vascular response to endothelin, which activates the same second-messenger vasoconstrictor system as 5-HT. These findings, which did not depend on guanylate cyclase, are consistent with chemical modification by NO of the 5-HT(2) G protein-coupled receptor system to inhibit vasoconstriction, possibly by S-nitrosylation of the receptor or a related protein. This study demonstrates that GSNO can regulate vascular tone in the intact lung by a reversible mechanism involving inhibition of the response to 5-HT.

Animals↗