Unexpected maturation of PI3K and MAPK-ERK signaling in fetal ovine cardiomyocytes.

In the first two-thirds of gestation, ovine fetal cardiomyocytes undergo mitosis to increase cardiac mass and accommodate fetal growth. Thereafter, some myocytes continue to proliferate while others mature and terminally differentiate into binucleated cells. At term (145 days gestational age; dGA) about 60% of cardiomyocytes become binucleated and exit the cell cycle under hormonal control. Rising thyroid hormone (T3) levels near term (135 dGA) inhibit proliferation and stimulate maturation. However, the degree to which intracellular signaling patterns change with age in response to T3 is unknown. We hypothesized that in vitro activation of ERK, Akt, and p70(S6K) by two regulators of cardiomyocyte cell cycle activity, T3 and insulin like growth factor-1 (IGF-1), would be similar in cardiomyocytes at gestational ages 100 and 135 dGA. IGF-1 and T3 each independently stimulated phosphorylation of ERK, Akt, and p70(S6K) in cells at both ages. In the younger mononucleated myocytes, the phosphorylation of ERK and Akt was reduced in the presence of IGF-1 and T3. However, the same hormone combination led to a dramatic twofold increase in the phosphorylation of these signaling proteins in the 135 dGA cardiomyocytes-even in cells that were not proliferating. In the older cells, both mono- and binucleated cells were affected. In conclusion, fetal ovine cardiomyocytes undergo profound maturation-related changes in signaling in response to T3 and IGF-1, but not to either factor alone. Differences in age-related response are likely to be related to milestones in fetal cardiac development as the myocardium prepares for ex utero life.

Insignificant response of the fetal placental circulation to arterial hypotension in sheep.

Infusion of the angiotensin-converting enzyme inhibitor enalaprilat into fetal sheep caused a profound arterial hypotension within days. Five fetal lambs were infused with enalaprilat for 8 days starting at day 128 of gestation. Total accumulated dose was 0.30 ± 0.11 mg/kg. Arterial pressure decreased from 43.6 to 25.6 mmHg; venous pressure did not change. Biventricular output was not statistically significantly changed; placental blood flow decreased almost in proportion to the decrease in pressure but the increase in somatic flow was not statistically significant. There were no significant changes in pressure 30 min after the initial 50-µg loading dose of enalaprilat. However, the arterial pressure responses to test doses of ANG I were largely abolished. After 1 day, however, there was a significant decrease in somatic vascular resistance, which became stronger with time, but almost no decrease in the placental resistance. We conclude that the fetal somatic circulation exhibits a slow but strong decrease in resistance but that the response to hypotension is weak or absent in the fetal placenta, possibly because it is already fully relaxed.

Atrial natriuretic peptide inhibits angiotensin II-stimulated proliferation in fetal cardiomyocytes.

The role of atrial natriuretic peptide (ANP) in regulating fetal cardiac growth is poorly understood. Angiotensin II (Ang II) stimulates proliferation in fetal sheep cardiomyocytes when growth is dependent on the activity of the mitogen-activated protein kinase (MAPK) and phosphoinositol-3-kinase (PI3K) pathways. We hypothesized that ANP would suppress near-term fetal cardiomyocyte proliferation in vitro and inhibit both the MAPK and PI3K pathways. Forty-eight hour 5-bromodeoxyuridine (BrdU) uptake (used as an index of proliferation) was measured in cardiomyocytes isolated from fetal sheep (135 day gestational age) in response to 100 nm Ang II with or without ANP (0.003-100 nm) or 1 microm 8-bromo-cGMP. The effects of these compounds on the MAPK and PI3K pathways were assessed by measuring extracellular signal-regulated kinase (ERK) and AKT phosphorylation following 10 min of treatment with Ang II, ANP or 8-bromo-cGMP. In right ventricular myocytes (RV), the lowest dose of ANP (0.003 nm) inhibited Ang II-stimulated BrdU uptake by 68%. Similarly, 8-bromo-cGMP suppressed Ang II-stimulated proliferation by 62%. The same effects were observed in left ventricular (LV) cardiomyocytes but the RV was more sensitive to the inhibitory effects of ANP than the LV (P < 0.0001). Intracellular cGMP was increased by 4-fold in the presence of 100 nm ANP. Ang II-stimulated ERK and Akt phosphorylation was inhibited by 100 nm ANP. The activity of ANP may in part be cGMP dependent, as 8-bromo-cGMP had similar effects on the cardiomyocytes.

Reduced systolic pressure load decreases cell-cycle activity in the fetal sheep heart.

The fetal heart is highly sensitive to changes in mechanical load. We have previously demonstrated that increased cardiac load can stimulate cell cycle activity and maturation of immature cardiomyocytes, but the effects of reduced load are not known. Sixteen fetal sheep were given either continuous intravenous infusion of lactated Ringer solution (LR) or enalaprilat, an angiotensin-converting enzyme inhibitor beginning at 127 days gestational age. After 8 days, fetal arterial pressure in the enalaprilat-infused fetuses (23.8 +/- 2.8 mmHg) was lower than that of control fetuses (47.5 +/- 4.7 mmHg) (P < 0.0001). Although the body weights of the two groups of fetuses were similar, the heart weight-to-body weight ratios of the enalaprilat-infused fetuses were less than those of the LR-infused fetuses (5.6 +/- 0.5 g/kg vs. 7.0 +/- 0.6 g/kg, P < 0.0001). Dimensions of ventricular myocytes were not different between control and enalaprilat-infused fetuses. However, there was a significant decrease in cell cycle activity in both the right ventricle (P < 0.005) and the left ventricle (P < 0.002) of the enalaprilat-infused fetuses. Thus, we conclude a sustained reduction in systolic pressure load decreases hyperplastic growth in the fetal heart.

Effect of fetal anaemia on myocardial ischaemia-reperfusion injury and coronary vasoreactivity in adult sheep.

AIMS: We investigated whether chronic fetal anaemia affects myocardial infarct in adulthood and elicits functional modifications in adult coronary vasoreactivity. METHODS: Seven-month-old sheep that were made anaemic in utero and transfused to normal haematocrit before birth were studied. Infarct size was determined by tetrazolium after 1-h ischaemia (occlusion of the mid of left anterior descending artery) and 2-h reperfusion. The dose-response to vasoconstrictors and vasodilators was assessed in small resistance coronary arteries. RESULTS: There were no significant differences between the animals previously subjected to in utero anaemia and the control animals regarding the percentage infarct size and the area-at-risk to the left ventricle. The ventricular function (dP/dt) was preserved. The percentage infarct size of the area-at-risk (70.7 +/- 3.5%) was larger than that in the controls (49.8 +/- 4.5%) (P = 0.006). The vascular responses were not altered. Endothelium-dependent relaxation to bradykinin (96.0 +/- 2.6% vs. 98.8 +/- 1.0%) was not affected by PGI(2) inhibitor (94.6 +/- 2.6% vs. 98.5 +/- 1.0%) but significantly reduced by the inhibition of nitric oxide (NO) in both anaemic (P < 0.05) and control (P < 0.001) groups with a significant right shift of EC(50) (P < 0.01). The non-NO-non-PGI(2)-mediated relaxation was slightly potentiated in anaemic animals. CONCLUSIONS: Exposing fetal sheep to in utero anaemia in late gestation for 3 weeks may increase the susceptibility of adult hearts to ischaemia-reperfusion injury without major alterations in coronary vasomotor responsiveness. The impact of in utero anaemia at earlier period of pregnancy and on the earlier or later life of the adult is yet to be further investigated.

Persistent changes in arterial blood gases in fetal sheep.

Two anaesthetic protocols were compared using pregnant sheep. In both groups of animals, anaesthesia was induced using an intravenous (i.v.) injection of diazepam and ketamine. The ewes were then intubated for positive pressure ventilation using 0.8 L/min of nitrous oxide and 2 L/min oxygen with 1.1-1.8% halothane. If the ewe showed any signs of awakening, one of two protocols was followed. First, the halothane concentration was increased to 2-3% until the ewe was completely anaesthetized. Second, the halothane concentration was not altered, but the ewe was given doses of i.v. diazepam (0.1 mg/kg) and ketamine (1 mg/kg) until again completely anaesthetized. At the completion of surgery, maternal recovery was rapid and similar between the two groups. However, five days after surgery, the fetal arterial Po(2) and oxygen content of the fetuses receiving additional halothane (1.9 +/- 0.2 kPa and 4.4 +/- 1.0 mL/100 mL) were statistically significantly depressed when compared with the fetuses receiving additional diazepam and ketamine (2.9 +/- 0.1 kPa and 7.0 +/- 0.5 mL/100 mL). These results led us to conclude that certain anaesthetic protocols, in spite of good maternal recovery, can lead to deleterious effects upon the fetus that persist for at least five days after surgery.

Thyroid hormone inhibits proliferation of fetal cardiac myocytes in vitro.

Thyroid hormone (T(3)) is a key regulator of fetal organ maturation. Premature elevations of thyroid hormone may lead to a 'mature' cardio-phenotype. Thyroid hormone will stimulate maturation of ovine fetal cardiomyocytes in culture by decreasing their proliferative capacity. Group 1 fetal cardiomyocytes (approximately 135 days gestation) were incubated with T3 (1.5, 3, 10, and 100 nM) and bromodeoxyuridine (BrdU; 10 microM) for 24 and 48 h. Group 2 cardiomyocytes were cultured with T3 alone for later protein analysis of cell cycle regulators. At all concentrations, T3 decreased BrdU uptake fourfold in serum media (P<0.001 versus serum, n=5). Following serum-free (SF) T3 treatment, BrdU uptake was inhibited when compared with serum (P<0.001 versus serum, n=5). p21 expression increased threefold (P<0.05 versus serum free, n=4) and cyclin D1 expression decreased twofold (P<0.05 versus serum, n=4) in T3-treated cardiomyocytes. (1) T3 inhibits fetal cardiomyocyte proliferation, while (2) p21 protein levels increase, and (3) cyclin D1 levels decrease. Thus, T3 may be a potent regulator of cardiomyocyte proliferation and maturation in the late gestation fetus.

Cortisol stimulates cell cycle activity in the cardiomyocyte of the sheep fetus.

The role of cortisol in regulating cardiac myocyte growth in the near-term fetal sheep is unknown. We hypothesized that cortisol would suppress cardiomyocyte proliferation and stimulate cardiomyocyte binucleation and enlargement, signs of terminal differentiation. Cardiomyocyte dimensions and percent binucleation were determined in isolated cardiac myocytes from seven cortisol-treated and seven control fetuses; percentage of myocytes positive for Ki-67 was determined in an additional four cortisol-treated and four control hearts. Cortisol was infused into the circumflex coronary artery at subpressor rates (0.5 microg/kg.min, 7 d). Cortisol infusion had no hemodynamic effects, compared with controls or pretreatment conditions. Cortisol treatment increased heart weight (44.0 +/- 8.7 g vs. control, 34.9 +/- 9.1 g, P < 0.05). Heart to body weight ratio was greater in treated hearts, compared with controls (10.3 +/- 1.9 vs. 7.7 +/- 0.9 g/kg, P < 0.01). Ventricular myocyte length, width, and percent binucleation were not different between groups. The proportion of treated myocytes in the cell cycle staining for Ki-67 was higher in the left ventricle (5.5 +/- 0.1 vs. 2.7 +/- 0.4%, P < 0.005) and right ventricle (4.4 +/- 0.4 vs. 3.7 +/- 0.7%, P < 0.05), compared with controls. Wet weight to dry weight ratios from cortisol-treated and control hearts were not different. In conclusion, whereas cortisol infused into the fetal sheep heart has no effect on cardiomyocyte size or maturational state, it stimulates entry of cardiomyocytes in the cell cycle. Thus, increases in fetal heart mass associated with subpressor doses of cortisol are due to cardiomyocyte proliferation and not hypertrophic growth.

Fetal infusions of plasma cause an increase in umbilical vascular resistance in sheep.

Earlier studies suggested that the fetal placental circulation is relatively inert with fetal placental flow increasing or decreasing with perfusion pressure. Subsequent studies have demonstrated that the placenta may not be an unreactive vascular bed. The present study was undertaken to determine if plasma infusion-induced hypertension increased fetal placental flow in proportion to the driving pressure across the fetal placental circulation. Six fetal sheep were operated on at 118-122 days to place intravascular catheters and a flow sensor on the common umbilical artery. Starting 6 days later, the fetuses were infused with adult sheep plasma. During the 7-day-long infusion period, they received a total of 1515+/-217 (SD) ml of fluid and 93.2+/-12.0 g of protein. Fetal plasma protein concentrations increased from 34.2+/-2.3 to 77.0+/-9.7 g/l (P<0.0001). Fetal arterial blood pressures rose from 42+/-3 to 59+/-4 mmHg (P<0.01) and venous pressures rose from 2.2+/-0.5 to 4.8+/-0.8 mmHg (P<0.01). In spite of the large increase in driving pressure, fetal placental blood flow remained (statistically) constant (627+/-299 ml/min and 552+/-221 ml/min) while fetal umbilical resistance increased from 0.077+/-0.038 to 0.115+/-0.053 mmHg min/ml (P<0.01). On day 7, plasma renin activity had fallen from 6.7+/-4.2 ng/(ml/h) at preinfusion control to 0.6+/-0.6 ng/(ml/h) (P<0.05) and plasma angiotensin-II concentration had fallen from 33.2+/-26.6 to 6.2+/-3.9 pg/ml, although this fall was not statistically significant (P=0.07). Fetal placental flow did not increase with increased driving pressure across the fetal placental circulation. The increase in fetal placental resistance may be a response to the increase in arterial pressure since there was no increase in flow.

The effects of anaemia as a programming agent in the fetal heart.

The intrauterine environment plays a powerful role in determining the life-long risk of cardiovascular disease. A number of stressors are well known to affect the development of the cardiovascular system in utero including over/under maternal nutrition, excess glucocorticoid and chronic hypoxia. Chronic fetal anaemia in sheep is a complex stressor that alters cardiac loading conditions, causes hypoxic stress and stimulates large changes in flow to specific tissues, including large increases in resting coronary blood flow and conductance. Decreased viscosity can account for approximately half of the increased flow. It appears that immature hearts are 'plastic' in that increases in coronary conductance with fetal anaemia persist into adulthood even if the anaemia is corrected before birth. These large changes in conductance are possible only through extensive remodelling of the coronary tree. Adult hearts that were once anaemic in utero are more resistant to hypoxic stress as adults but it is not known whether such an adaptation would be deleterious in later life. These studies indicate the need for investigation into the basic mechanisms of coronary tree remodelling in the immature myocardium. New information on these mechanisms is likely to lead to better prevention of and therapies for adult-onset coronary disease.

Anaemia stimulates aquaporin 1 expression in the fetal sheep heart.

Interstitial fluid fluxes are much greater in the fetus than in the adult, and filtration rates are increased over control in most tissues of the anaemic fetus. Increased capillary filtration may lead to cardiac oedema which, in turn, severely impacts cardiac function. Mechanisms that underlie these differences in flux are incompletely understood. One possible mechanism is an increase in capillary water permeability. Therefore, the goal of our study was to determine the level of expression of the water channel aquaporin 1 (AQP1) during cardiac development and in the anaemic fetal sheep heart. Hearts from chronically instrumented anaemic sheep fetuses and hearts from normal early fetal, late fetal, neonatal and adult sheep were used for Northern and Western analyses and immunohistochemistry. We found that AQP1 mRNA levels were lower in the young fetal left ventricle than in the adult left ventricle (P < 0.05). We also found that cardiac AQP1 expression was increased in anaemic fetuses compared to age-matched controls (P < 0.05). Expression of AQP1 in all groups was greatest in the microvascular endothelium. These data suggest that AQP1 plays an important role in the physiological accommodation to fetal anaemia.

Angiotensin II stimulates hyperplasia but not hypertrophy in immature ovine cardiomyocytes.

Rat and sheep cardiac myocytes become binucleate as they complete the 'terminal differentiation' process soon after birth and are not able to divide thereafter. Angiotensin II (Ang II) is known to stimulate hypertrophic changes in rodent cardiomyocytes under both in vivo and in vitro conditions via the AT1 receptor and intracellular extracellular regulated kinase (ERK) signalling cascade. We sought to develop culture methods for immature sheep cardiomyocytes in order to test the hypothesis that Ang II is a hypertrophic agent in the immature myocardium of the sheep. We isolated fetal sheep cardiomyocytes and cultured them for 96 h, added Ang II and phenylephrine (PE) for 48 h, and measured footprint area and proliferation (5-bromo-2'-deoxyuridine (BrdU) uptake) separately in mono- vs. binucleate myocytes. We found that neither Ang II nor PE changed the footprint area of mononucleated cells. PE stimulated an increase in footprint area of binucleate cells but Ang II did not. Ang II increased myocyte BrdU uptake compared to serum free conditions, but PE did not affect BrdU uptake. The MAP kinase kinase (MEK) inhibitor UO126 prevented BrdU uptake in Ang II-stimulated cells and prevented cell hypertrophy in PE-stimulated cells. This paper establishes culture methods for immature sheep cardiomyocytes and reports that: (1) Ang II is not a hypertrophic agent; (2) Ang II stimulates hyperplastic growth among mononucleate myocytes; (3) PE is a hypertrophic agent in binucleate myocytes; and (4) the ERK cascade is required for the proliferation effect of Ang II and the hypertrophic effect of PE.

Augmentation of coronary conductance in adult sheep made anaemic during fetal life.

Maximal coronary conductance with adenosine in anaemic fetal sheep is twice that of non-anaemic fetuses. To investigate whether this increase in conductance persists into adulthood we studied twin sheep as fetuses and again as adults. Nine anaemic fetuses (118 days gestation) underwent isovolaemic haemorrhage for 18.0 +/- 4.6 days (means +/- S.D.) during which time the haematocrit was reduced from 39.9 +/- 5.2 % to 16.3 +/- 3.4 % and oxygen content from 8.6 +/- 1.3 to 2.3 +/- 0.2 ml dl-1. At 138 days the anaemic fetuses were transfused; at delivery the haematocrit was 29.3 +/- 6.8 % compared to nine control fetuses in which the haematocrit was 38.5 +/- 4.3 %. The weight at delivery was 3.5 +/- 0.36 kg in the anaemic fetuses vs. 4.2 +/- 0.83 kg in controls. Twenty-eight weeks later, we placed an occluder on the descending thoracic aorta and inferior vena cava, a flow probe around the proximal left circumflex coronary artery, and catheters in the left atrial appendage, jugular and carotid vessels. Maximal coronary conductance was determined in the adults by recording coronary blood flow as driving pressure was altered by inflating the occluders while adenosine was infused into the left atrium. Right atrial, left atrial, systolic and mean arterial pressures, systemic vascular resistance and haematocrit were not different between 'in utero anaemic' and control adults. The adults that were anaemic in utero weighed less than the controls 39.4 +/- 4.6 kg vs. 45.0 +/- 5.6 kg. Maximal conductance was greater in the adults that were anaemic in utero: 11.2 +/- 4.0 ml min(-1) (100 g)(-1) mmHg-1 as compared to 6.1 +/- 1.8 ml min(-1) (100 g)(-1) mmHg(-1) in the controls. Vascular reactivity of the mesenteric arteries was not different. These data suggest that coronary conductance can be modified in utero by anaemia (high flow and hypoxaemia) and that the remodelled coronary tree persists to adulthood.

Does nitric oxide contribute to the basal vasodilation of pregnancy in conscious rabbits?

Pregnancy produces marked systemic vasodilation, but the mechanism is unknown. Experiments were performed in conscious rabbits to test the hypotheses that increased nitric oxide (NO) production contributes to the increased vascular conductance, but that the contribution varies among vascular beds. Rabbits were instrumented with aortic and vena caval catheters and ultrasonic flow probes implanted around the ascending aorta, superior mesenteric artery, terminal aorta, and/or a femoral artery. Hemodynamic responses to intravenous injection of N(omega)-nitro-L-arginine (L-NA; 20 mg/kg or increasing doses of 2, 5, 10, 15, and 20 mg/kg) were determined in rabbits first before pregnancy (NP) and then at the end of gestation (P). L-NA produced similar increases in arterial pressure between groups, but the following responses were larger (P < 0.05) when the rabbits were pregnant: 1) decreases in total peripheral conductance [-3.7 +/- 0.3 (NP), -5.0 +/- 0.5 (P) ml x min(-1) x mmHg(-1)], 2) decreases in mesenteric conductance [-0.47 +/- 0.05 (NP), -0.63 +/- 0.07 (P) ml x min(-1) x mmHg(-1)], 3) decreases in terminal aortic conductance [-0.43 +/- 0.05 (NP), -0.95 +/- 0.19 ml x min(-1) x mmHg(-1) (P)], and 4) decreases in heart rate [-41 +/- 4 (NP), -62 +/- 5 beats/min (P)]. Nevertheless, total peripheral and terminal aortic conductances remained elevated in the pregnant rabbits (P < 0.05) after L-NA. Furthermore, decreases in cardiac output and femoral conductance were not different between the reproductive states. We conclude that the contribution of NO to vascular tone increases during pregnancy, but only in some vascular beds. Moreover, the data support a role for NO in the pregnancy-induced increase in basal heart rate. Finally, unknown factors in addition to NO must also underlie the basal vasodilation observed during pregnancy.

Hemodynamic effects of S-nitrosocysteine, an intravenous regional vasodilator.

BACKGROUND: S-nitrosocysteine is a carrier form of nitric oxide that can be delivered intravenously. S-nitrosocysteine is rapidly metabolized by plasma (half-life = 2-3 seconds), forming nitric oxide and cysteine. With its short half-life and potent vasodilatory properties, S-nitrosocysteine may be useful as a pulmonary vasodilating agent in cases of postoperative and chronic pulmonary hypertension. OBJECTIVE: Our objective was to determine the hemodynamic properties of S-nitrosocysteine on the pulmonary and systemic circulations to assess its potential utility as a pulmonary vasodilatory agent. METHODS: Eleven adult swine were anesthetized. Thermodilution (Swan-Ganz; Baxter International, Inc, Deerfield, Ill) and arterial catheters were inserted. Flow probes were placed around the coronary, renal, superior mesenteric, and iliac arteries. Incremental infusion doses of S-nitrosocysteine (5-80 nmol. kg(-1). min(-1)) were delivered into the right atrium. Cardiac output, right and left heart pressures, heart rate, Pao(2), and iliac, renal, coronary, and mesenteric blood flow rates were recorded at baseline and at each infusion dose of S-nitrosocysteine. RESULTS: Low-dose S-nitrosocysteine infusion decreased mean pulmonary artery pressure (15%, P =.013) without a significant reduction in mean systemic artery pressure. Higher dose infusions produced further dose-dependent declines in pulmonary vascular resistance and measurable reductions in systemic vascular resistance (P =.01). At an S-nitrosocysteine dosage of 40 nmol. kg(-1). min(-1), there was a significant reduction in renal (P <.001) and mesenteric (P =.003) blood flow but no change in iliac (P >.2) or coronary (P >.2) blood flow. Cardiac output remained constant up to infusion rates of 40 nmol. kg(-1). min(-1) (P >.2). Doses higher than 5 nmol. kg(-1). min(-1) resulted in a substantial dose-dependent reduction in Pao(2) (P <.001), suggesting dilation of atelectatic areas of the lung. CONCLUSION: S-nitrosocysteine is a potent vasodilatory agent capable of overcoming the hypoxic vasoconstrictive response of the lung. Our results suggest it may prove useful as a pulmonary vasodilatory agent at low doses. Higher dose infusions reduce mean systemic pressure and lead to compensatory reductions in renal and mesenteric blood flow without a decrease in cardiac output.

Right ventricular systolic pressure load alters myocyte maturation in fetal sheep.

The effects of right ventricular (RV) systolic pressure (RVSP) load on fetal myocyte size and maturation were studied. Pulmonary artery (PA) pressure was increased by PA occlusion from mean 47.4 +/- 5.0 (+/-SD) to 71 +/- 13.6 mmHg (P < 0.0001) in eight RVSP-loaded near-term fetal sheep for 10 days. The maximal pressure generated by the RV with acute PA occlusion increased after RVSP load: 78 +/- 7 to 101 +/- 15 mmHg (P < 0.005). RVSP-load hearts were heavier (44.7 +/- 8.4 g) than five nonloaded hearts (31.8 +/- 0.2 g; P < 0.03); heart-to-body weight ratio (10.9 +/- 1.1 and 6.5 +/- 0.9 g/kg, respectively; P < 0.0001). RVSP-RV myocytes were longer (101.3 +/- 10.2 microm) than nonloaded RV myocytes (88.2 +/- 8.1 microm; P < 0. 02) and were more often binucleated (82 +/- 13%) than nonloaded myocytes (63 +/- 7%; P < 0.02). RVSP-loaded myocytes had less myofibrillar volume than did nonloaded hearts (44.1 +/- 4.4% and 56. 1 +/- 2.6%; P < 0.002). We conclude that RV systolic load 1) leads to RV myocyte enlargement, 2) has minor effects on left ventricular myocyte size, and 3) stimulates maturation (increased RV myocyte binucleation). Myocyte volume data suggest that RV systolic loading stimulates both hyperplastic and hypertrophic growth.

Hemodynamic changes in pregnancy.

The basic mechanisms that underlie alterations in the physiology of pregnancy are virtually unknown. Basal oxygen consumption increases by some 50 mL/min in pregnant women at term. Blood volume increases gradually over gestation as does red cell mass. Cardiac output increases by some 50% by mid-third trimester. Stroke volume and heart rate increase over the course of pregnancy with heart rate increasing gradually until term. The heart of the pregnant woman remodels dramatically in the first few weeks of pregnancy; end diastolic volume increases. Stroke volume is augmented by the increase in end diastolic volume and maintenance of ejection fraction through a possible increase in contractile force. Systolic and diastolic blood pressures drop during normal pregnancy. There is evidence of blood vessel remodeling in all vessels. Venous compliance and venous blood volume are increased. Renal plasma flow increases by some 70% in pregnancy with glomerular filtration rate increasing by 50% by unknown mechanisms. The complex hormonal environment is changing throughout pregnancy. In summary, under the influence of circulating chemical mediators blood flow is redistributed to the uterus, breast, and kidney.

Nasal administration of the nitric oxide synthase inhibitor L-NAME induces daytime somnolence.

In preliminary studies, human subjects complained of drowsiness after aerosolization of NG-nitro-L-arginine methyl ester (L-NAME) into the nasal passages. We compared the effects of a nasal aerosol of L-NAME (0.5 M, 4 ml) to those of saline on sleep onset latency and exhaled nasal nitric oxide (NO). L-NAME decreased sleep onset latency and exhaled nasal NO. Vasoconstriction and local effects of L-NAME on NO synthesis are unlikely to explain this effect since oxymetazoline, a vasoconstrictor, decreased exhaled NO but had no effect on sleep onset latency. We conclude that aerosolization of L-NAME to the nasal passages induces daytime sleepiness.

Temperature conditioning of nasal air: effects of vasoactive agents and involvement of nitric oxide.

Nitric oxide (NO) is released into nasal air, but its function is unknown. We hypothesized that nasal vascular tone and/or flow influences temperature conditioning of nasal air and that NO participates in this process. We measured nasal air temperature (via a thermocouple) and exhaled nasal NO release (by chemiluminescence) in five humans and examined the effects of an aerosolized vasoconstrictor (oxymetazoline), a vasodilator (papaverine), N(G)-nitro-L-arginine methyl ester, an inhibitor of NO synthase, or saline (control). Compared with saline (which caused no changes in nasal air temperature or exhaled NO release), oxymetazoline (0.05%) reduced nasal air temperature and NO release (130.8 +/- 15.1 to 81.3 +/- 12.8 nl. min(-1). m(-2); P < 0.01). Papaverine (0.01 M) increased nasal air temperature and NO release (131.8 +/- 13.1 to 157.2 +/- 17.4 nl. min(-1). m(-2); P < 0.03). N(G)-nitro-L-arginine methyl ester reduced nasal air temperature and NO release (123.7 +/- 14.2 to 44.2 +/- 23.7 nl. min(-1). m(-2); P < 0.01). The results suggest that vascular tone and/or flow modulates temperature conditioning and that NO may participate in that function.

Angiotensin type 1 receptor antagonism with irbesartan inhibits ventricular hypertrophy and improves diastolic function in the remodeling post-myocardial infarction ventricle.

To evaluate the role of angiotensin II (AII) on diastolic function during post-myocardial infarction (MI) ventricular remodeling, coronary ligation or sham operation was performed in male Sprague-Dawley rats. Experimental animals were maintained on either irbesartan, a selective AT1-receptor antagonist, or no treatment. Measurement of cardiac hypertrophy, diastolic function, and sarcoendoplasmic reticulum adenosine triphosphatase (ATPase; SERCA) and phospholamban (PLB) gene expression was assessed at 6 weeks after MI. Myocardial infarction caused a significant increase in myocardial mass and left ventricular (LV) filling pressure, whereas LV systolic pressure and +dP/dt were reduced. The time constant of isovolumic relaxation (tau) was markedly prolonged after MI. Post-MI hypertrophy was associated with substantial increases in the messenger RNA (mRNA) expression of atrial natriuretic peptide (ANP), but no significant changes in SERCA or PLB levels. Although irbesartan treatment did not significantly alter post-MI LV systolic or filling pressures, it nevertheless effectively decreased ventricular hypertrophy, improved tau, and normalized ANP expression. These results demonstrate that AT1-receptor antagonism has important effects on myocardial hypertrophy and ANP gene expression, which are independent of ventricular loading conditions. In addition, the improvement in diastolic function was not related to changes in SERCA and PLB gene expression, suggesting that enhanced myocardial relaxation was related to the blockade of AII effects on myocyte function or through a reduction of ventricular hypertrophy itself or both.