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An experimental ovine fetal model for perinatal pulmonary hypertension of the neonate (PPHN) was characterized by altered pulmonary vasoreactivity and structure. Because past studies had suggested impaired nitric oxide-cGMP cascade in this experimental model, we hypothesized that elevated phosphodiesterase (PDE) activity may contribute to altered vascular reactivity and structure in experimental PPHN. Therefore, we studied the effects of the PDE inhibitors zaprinast and dipyridamole on fetal pulmonary vascular resistance and PDE5 activity, protein, mRNA, and localization in normal and pulmonary hypertensive fetal lambs. Infusion of dipyridamole and zaprinast lowered pulmonary vascular resistance by 55 and 35%, respectively, in hypertensive animals. In comparison with control animals, lung cGMP PDE activity was elevated in hypertensive fetal lambs (150%). Increased PDE5 activity was not associated with either an increased PDE5 protein or mRNA level. Immunocytochemistry demonstrated that PDE5 was localized to vascular smooth muscle. We concluded that PDE5 activity was increased in experimental PPHN, possibly by posttranslational phosphorylation. We speculated that these increases in cGMP PDE activity contributed to altered pulmonary vasoreactivity in experimental perinatal pulmonary hypertension.

During transitional circulation, the pulmonary vascular bed undergoes a rapid and profound reduction in both tone and vascular smooth-muscle (VSM) content. 3',5'-Guanylate cyclic monophosphate (cGMP) is a crucial mediator in the regulation of pulmonary vascular resistance (PVR) and VSM proliferation. Hydrolysis of cGMP is achieved predominately by cGMP-specific phosphodiesterases (PDEs). Among the cGMP-specific PDEs, PDE5 is quantitatively prevalent in lung tissue. We have investigated the levels of pulmonary PDE5 enzymatic activity, protein, and messenger RNA (mRNA) in ovine and mouse lung during perinatal development. We report that within 1 h following birth, PDE5 activity, protein, and mRNA levels decrease in both species, in a manner that correlates with known decreases in PVR in early transition. However, from 4 to 7 d following birth, a secondary increase in PDE5 activity, protein, and mRNA occurs in both ovine and mouse lung, suggesting a complex regulation of PVR and VSM proliferation in late perinatal development. Our data imply that PDE5 may be an important mediator in the regulation of PVR in normal and possibly in pathologic states, and may ultimately provide a basis for PDE5 inhibitors as a treatment for pulmonary hypertension.

Nitric oxide (NO) modulates pulmonary vascular resistance (PVR) in the normal fetus by increasing the cyclic guanosine 3',5'-monophosphate (cGMP) content of pulmonary vascular smooth muscle cells. Although several vasodilator stimuli, including acetylcholine, decrease fetal PVR through the release of endogenous NO, fetal pulmonary vasodilation is often transient despite prolonged treatment. Because cGMP is hydrolyzed and inactivated by cGMP-specific (type 5) phosphodiesterases (PDE5), we hypothesized that PDE5 activity contributes to high fetal PVR and limits the capability of the fetal pulmonary circulation to dilate or sustain vasodilation in response to cGMP-dependent stimuli. To test this hypothesis, we studied the hemodynamic effects of dipyridamole in 19 late-gestation fetal lambs. To determine whether dipyridamole-induced vasodilation is dependent upon basal NO release, we measured the response to dipyridamole before and after pretreatment with the NO synthase antagonist nitro-L-arginine (L-NA) in five fetal lambs. L-NA completely blocked dipyridamole-induced pulmonary vasodilation. To evaluate the effect of dipyridamole on pulmonary vasodilation due to the stimulated release of NO, we studied effects of prolonged intrapulmonary acetylcholine infusions, with and without concomitant administration of low-dose dipyridamole, in six fetal lambs. During prolonged (2-h) infusions, acetylcholine and dipyridamole individually caused transient pulmonary vasodilation. When administered together, pulmonary vasodilation was of greater magnitude and was sustained for the entire study period. To determine the effects of dipyridamole on endothelium-independent pulmonary vasodilation, we investigated the hemodynamic effects of inhaled NO (5 and 20 ppm) alone and in combination with dipyridamole during mechanical ventilation with low FlO2. The combination of dipyridamole with inhaled NO resulted in a greater degree of pulmonary vasodilation than that achieved with inhaled NO alone. We conclude that dipyridamole-induced pulmonary vasodilation is dependent on endogenous (basal) NO production and that dipyridamole potentiates vasodilator responses to endothelium-dependent and -independent dilators in the ovine fetal pulmonary circulation. We speculate that PDES activity opposes vasodilation and maintains high PVR in the normal fetal lung.

The effects of propofol and its intralipid vehicle on increased and normal pulmonary vascular resistances (PVR) were studied in an in situ perfused rabbit lung model which controlled all major determinants of PVR. Propofol at both 5 and 10 micrograms/mL significantly reduced PVR increases by the thromboxane mimetic, U46619. In contrast, similar volumes of intralipid further increased PVR. Neither propofol nor intralipid had any effect on normal PVR. We conclude that propofol at 5 and 10 micrograms/mL is able to reduce increased PVR but has no effect on normal PVR.

In 12 pentobarbital anesthetized dogs the lungs were independently ventilated with a double piston ventilator. The right lung was ventilated throughout with 100% oxygen. Blood was drawn from the right atrium and pumped through a bubble oxygenator to a cannula in the ligated left main pulmonary artery. The pressures in the left main pulmonary artery and the left atrium were recorded during constant flow while the oxygen tension in the left lung alveolar gas and the perfusate were varied either to match each other (Protocol 1) or differ (Protocol 2) over the range from "zero" to "100%" oxygen. From the combined data a three dimensional response surface for hypoxic pulmonary vasoconstriction was derived. The maximum increase of pulmonary vascular resistance (r%PVRmax) was defined at a stimulus oxygen tension (PSO2) of 10 mmHg amounting to a 3.15 +/- (0.18)-fold increase of the vascular resistance on "100%" oxygen. The stimulus oxygen tension was shown to be PSO2 = PVO2(0.41) x PAO2(0.59) and the dose-response sigmoid for hypoxic pulmonary vasoconstriction in canine lungs was derived as r%PVRmax = 100 (PSO2(-2.616))/(6.683 x 10(-5) + PSO2(-2.616)) These results appear to reconcile observations from a number of laboratories and to be of quite general application.

The effects of amrinone on pulmonary vascular resistance (PVR) were studied in an isolated, perfused rabbit lung model where all the major determinants of PVR were controlled. In this preparation, the alveolar oxygen and carbon dioxide tensions, vascular pH and vascular oxygen and carbon dioxide tensions, and zonal conditions of the lung and phasic variations of pulmonary artery pressures could be precisely measured and controlled. Measurements of PVR were made by a complete determination of the pulmonary pressure-flow curve and determination of the PVR under identical flow conditions for all studies. This approach allowed a more precise determination of the primary effects of amrinone on normal and elevated PVR than has been previously possible. We found that amrinone in final concentrations of either 4 or 8 micrograms/ml had no effect on basal PVR and no effect on lung water weight to dry ratios. When PVR was elevated by the addition of the thromboxane A2 mimetic U46619, amrinone reduced the PVR by 27% at a final concentration of 4 micrograms/ml and by 74% at a final concentration of 8 micrograms/ml. We conclude that in the doses tested, amrinone has no effects on basal PVR but is able to reduce elevated PVR in a dose-dependent manner. These results are the first to demonstrate clearly that amrinone has the ability to reduce elevated pulmonary vascular tone through a direct mechanism and not through secondary effects on other determinants of PVR such as left atrial pressure (Pla), increased cardiac output with resultant vascular recruitment, or increases in mixed venous oxygen tension. The possible implications for the clinical use of amrinone in situations of elevated PVR are discussed.

The isolated perfused lower left lung lobe of the exteriorized fetal lamb was used to define quantitatively the relationship between pulmonary perfusate oxygen tension and pulmonary vascular resistance (PVR) in the fetus at multiple oxygen tensions over the range from 8.3 to 433 mm Hg. This allowed variation of the perfusate PO2 over the range of partial pressures from less than 10 mm Hg to over 400 mm Hg while constant values of PCO2, temperature and perfusate flow were maintained. In all animals, calculated pulmonary vascular resistance varied in an inverse manner with the perfusate PO2. The relationship between PVR and perfusate oxygen tension is described by the equation: PVR = 7.67 - 1.54 (log PO2) R2 = 0.70. While others have shown that a single, large increase in blood oxygen tension will decrease the PVR in fetal lambs, these data present the first quantitative description of the role of oxygen tension in the modulation of fetal pulmonary vascular resistance as determined at multiple perfusate oxygen tensions over a fifty-fold range.