. Preservation of serotonin-mediated contractility in adult sheep pulmonary arteries following long-term high-altitude hypoxia. High Alt. Med. Biol. 12:253-264.-Long-term hypoxia (LTH) can increase serotonin (5-HT) signaling as well as extracellular calcium entry in adult rodent pulmonary arteries (PA), and 5-HT is associated with pulmonary hypertension. Because LTH, 5-HT, and calcium entry are related, we tested the hypothesis that LTH increases 5-HT-mediated PA contractility and associated calcium influx through L-type Ca 2 + channels, nonselective cation channels (NSCC), and reverse-mode sodium-Ca 2 + exchange. We performed wire myography and confocal calcium imaging on pulmonary arteries from adult ewes that lived near sea level or were maintained at high-altitude (3801 m) for *110 days. LTH did not increase the arterial medial wall thickness, nor did it affect the potency or efficacy for 5-HT-induced PA contraction. Ketanserin (100 nM), a 5-HT 2A antagonist, shifted the 5-HT potency to a far greater extent than 1 lM GR-55562, a 5-HT 1B/D inhibitor. These influences were unaffected by LTH. The rank order for reducing 5-

Antenatal maternal long-term hypoxia (LTH) can alter serotonin (5-HT) and calcium (Ca 2þ ) signaling in fetal pulmonary arteries (PAs) and is associated with persistent pulmonary hypertension of the newborn (PPHN). In humans, the antenatal maternal hypoxia can be secondary to smoking, anemia, and chronic obstructive pulmonary disorders. However, the mechanisms of antenatal maternal hypoxia-related PPHN are unresolved. Because both LTH and 5-HT are associated with PPHN, we tested the hypothesis that antenatal maternal LTH can increase 5-HT-mediated PA contraction and associated extracellular Ca 2þ influx through L-type Ca 2þ channels (Ca L ), nonselective cation channels (NSCCs), and reverse-mode sodium-calcium exchanger (NCX) in the near-term fetus. We performed wire myography and confocal-Ca 2þ imaging approaches on fetal lamb PA (*140 days of gestation) from normoxic ewes or those acclimatized to high-altitude LTH (3801 m) for *110 days. Long-term hypoxia reduced the potency but not the efficacy of 5-HT-induced PA contraction. Ketanserin (100 nmol/L), a 5-HT 2A antagonist, shifted 5-HT potency irrespective of LTH, while GR-55562 (1 mmol/L), a 5-HT 1B/D inhibitor, antagonized 5-HT-induced contraction in normoxic fetuses only. Various inhibitors for Ca L , NSCC, and reverse-mode NCX were used in contraction studies. Contraction was reliant on extracellular Ca 2þ regardless of maternal hypoxia, NSCC was more important to contraction than Ca L , and reverse-mode NCX had little or no role in contraction. Long-term hypoxia also attenuated the effects of 2-APB and flufenamic acid and reduced Ca 2þ responses observed by imaging studies. Overall, LTH reduced 5HT 1B/D function and increased NSCC-related Ca 2þ -dependent contraction in ovine fetuses, which may compromise pulmonary vascular function in the newborn.

Background-Nitrite can be converted to nitric oxide (NO) by a number of different biochemical pathways. In newborn lambs, an aerosol of inhaled nitrite has been found to reduce pulmonary blood pressure, possibly acting via conversion to NO by reaction with intraerythrocytic deoxyhemoglobin. If so, the vasodilating effects of nitrite would be attenuated by free hemoglobin in plasma that would rapidly scavenge NO. Methods and Results-Pulmonary vascular pressures and resistances to flow were measured in anesthetized newborn lambs. Plasma hemoglobin concentrations were then elevated, resulting in marked pulmonary hypertension. This effect was attenuated if infused hemoglobin was first oxidized to methemoglobin, which does not scavenge NO. These results further implicate NO as a tonic pulmonary vasodilator. Next, while free hemoglobin continued to be infused, the lambs were given inhaled NO gas (20 ppm), inhaled sodium nitrite aerosol (0.87 mol/L), or an intravascular nitrite infusion (3 mg/h bolus, 5 mg ⅐ kg Ϫ1 ⅐ h Ϫ1 infusion). Inhaled NO and inhaled nitrite aerosol both resulted in pulmonary vasodilation. Intravascular infusion of nitrite, however, did not. Increases in exhaled NO gas were observed in lambs while breathing the nitrite aerosol (Ϸ20 ppb NO) but not during intravascular infusion of nitrite. Conclusions-We conclude that the pulmonary vasodilating effect of inhaled nitrite results from its conversion to NO in airway and parenchymal lung tissue and is not dependent on reactions with deoxyhemoglobin in the pulmonary circulation. Inhaled nitrite aerosol remains a promising candidate to reduce pulmonary hypertension in clinical application. (Circulation. 2011;123:605-612.)

is metabolized in plasma, in part by the ferroxidase ceruloplasmin (Cp), to form nitrite and nitrosothiols (SNOs), which are proposed to mediate protective responses to hypoxia and ischemia. We hypothesized that NO metabolism would be attenuated in fetal plasma due to low Cp activity. We measured Cp concentrations and activity in plasma samples collected from adults and fetuses of humans and sheep. We then added NO ([NO]: 1.5 or 100 M) to plasma and aqueous buffer and measured rates of NO disappearance and the production of nitrite and SNO. Cp concentrations in fetal plasma were Ͻ15% of adult levels. In aqueous buffer, 1.5 M NO disappeared with a half-life of 347 Ϯ 64 s (means Ϯ SE) but in plasma of humans the half-life was 19 Ϯ 2 s (adult) and 11 Ϯ 1 s (fetus, P ϭ 0.004) and in sheep it was 31 Ϯ 3 s (adult) and 43 Ϯ 5 s (fetus, P ϭ 0.04). Cp activity was not correlated with the overall elimination half-life of NO or with the amount of SNO ([NO]: 100 M) or nitrite ([NO]: 1.5 or 100 M) produced but correlated with SNO yields at 1.5 M [NO] (r ϭ 0.92, P ϭ 0.04). Our data demonstrate that Cp is not essential to the increased rate of metabolism of NO in plasma relative to aqueous buffers and that it is not essential to the production of nitrite from NO. Cp may be involved in the conversion of NO to SNO in plasma under near-physiological concentrations of NO.ceruloplasmin; nitrite; nitrosothiol; fetal plasma NITRIC OXIDE (NO) IS PRODUCED by the vascular endothelium and diffuses either abluminally into the adjacent vascular smooth muscle cells to cause vasodilation or luminally into the flowing blood (23). In the blood, NO is rapidly oxidized to nitrate by reaction with oxyhemoglobin (8) or binds with high affinity to the heme groups of deoxygenated hemoglobin to produce iron-nitrosyl hemoglobin (7). Both reactions proceed at nearly diffusion-limited rates and thus would severely limit the vasoactive effects of free NO. However, encapsulation of hemoglobin within the erythrocyte slows the diffusion of NO to the hemoglobin by a factor of ϳ800 (16,25,48). This results in increased plasma NO concentrations, where it is metabolized to bioactive products such as nitrite (NO 2 Ϫ ) and S-nitrosothiols (SNO; Refs. 38,43,49). Increasing evidence suggests that these NO metabolites play an important endocrine role in the regulation of blood flow and provide tissue protection during hypoxic and ischemic stress (28), yet questions remain about the specific mechanisms underlying their production from NO in plasma.In oxygenated aqueous buffer, NO reacts with O 2 to yield nitrite according to the following reaction:This reaction, which is second order with respect to NO, has an overall rate constant of 2 to 4 ϫ 10 6 M Ϫ2 ·s Ϫ1 (11, 24). However, at physiological concentrations of NO and O 2 , the reaction is too slow to account for the rate of NO disappearance from plasma (38, 43). One possibility is that plasma contains one or more metal-containing enzymes that would catalyze the oxidation of NO to nitrite or facilitate nitrosation c...