Increase In Oxygen Tension Research Articles

The Effect of Hyperoxia on Reactive Oxygen Species (ROS) in Rat Petrosal Ganglion Neurons During Development Using Organotypic Slices

Hyperoxia, during development in rats, results in hypoxic chemosensitivity ablation, carotid body hypoplasia, and reduced chemoafferents. We hypothesized that hyperoxia increases reactive oxygen species (ROS) in cell bodies of chemoafferents. Organotypic slices of petrosal-nodose ganglia from rats at day of life (DOL) 5-6 and 17-18 were exposed to 8%, 21%, or 95% O(2) for 4 h in the presence or absence of the ROS-sensitive fluorescent indicator, CM-H(2)DCFDA, and propidium iodide was used to determine the relationship between cell death and oxygen tension. In tissue slices from DOL 5-6 rats, fluorescence intensity was 182.5 +/- 2.9 for hypoxia, 217.5 +/- 3.3 for normoxia, and 336.6 +/- 3.8 for hyperoxia, (mean +/- SEM, p < 0.001, ANOVA). Normoxia increased ROS levels by 19.2% from hypoxia (p < 0.01) with a further increase of 54.8% from normoxia to hyperoxia (p < 0.001). In tissue slices from DOL 17-18 rats, ROS levels increased with increasing oxygen tension but were less than in younger animals (p < 0.01, ANOVA). The antioxidants, NAC and TEMPO-9-AC, attenuated ROS levels and cell death. Electron microscopy demonstrated that hyperoxia damages the ultrastructure within petrosal ganglion neurons. Hyperoxic-induced increased levels of ROS in petrosal ganglion neurons may contribute to loss of hypoxic chemosensitivity during early postnatal development.

Cutaneous Tissue Flap Viability following Partial Venous Obstruction

Venous outflow obstruction is the most common cause of tissue failure after microvascular reconstructive surgery. If it is not recognized early, there is an increased risk of tissue damage and loss. Currently, however, there are no adequate models for the study of this clinical problem. The purpose of this study was to develop a partial congestion model for the study of skin flap physiology in response to varying levels of occluded venous outflow. Nine mixed-breed pigs were equally divided into three experimental groups (0 percent, 20 percent, and 50 percent venous outflow) to determine the effects of varying venous outflow on cutaneous flap color, oxygen tension, and edema. A cutaneous pedicle flap model and a partial congestion system were used to observe changes in variable venous obstruction. Only 0 percent venous outflow resulted in progressive color change across time. In addition, 0 percent venous outflow demonstrated significantly different oxygen tension levels relative to the other groups. Twenty percent venous outflow resulted in significant edema formation relative to the other groups. The 50 percent group showed an increase in oxygen tension from the second hour of venous obstruction to the end of the experiment. Tissue flap color is the clinical standard on which flap health is measured. After 8 hours, only complete venous occlusion resulted in significant color change. However, physiological changes that could affect tissue flap health were noted with only partial venous occlusion, including the development of edema formation. Accordingly, subtle color change could indicate partial venous congestion and may warrant intervention by the surgeon.

Dynamic HIF1A Regulation During Human Placental Development1

The human placenta is a unique organ in terms of oxygenation as it undergoes a transition from a low to a more oxygenated environment. This physiological switch in oxygen tension is a prerequisite for proper placental development and involves the hypoxia inducible factor (HIF). HIF is stable and initiates gene transcription under hypoxia, whereas in normoxia, interaction with the von Hippel-Lindau tumor suppressor protein (VHL) leads to rapid degradation of the HIF1A subunit. The degradation requires formation of a multiprotein complex (VHLCBC) and hydroxylation of HIF1A proline residues via members of the egg-laying-defective nine (EGLN) family. Herein, we have investigated the regulatory mechanisms of HIF1A expression during human placental development. Expression of HIF1A and VHL was high at 7-9 wk of gestation, when oxygen tension is low, and decreased when placental oxygen tension increases (10-12 wk of gestation). During early placentation, HIF1A localized in cytotrophoblasts, while VHL was present in syncytiotrophoblasts. At 10-12 wk, VHL appeared in cytotrophoblast cells, which coincided with the disappearance of HIF1A. At the same time the association of VHL and Cullin 2 as well as ubiquitination of HIF1A was maximal. EGLN1, EGLN2, and EGLN3 were also temporally expressed in an oxygen-dependent fashion, with greatest mRNA expression at 10-12 wk of gestation. Inhibition of EGLN activity increased HIF1A stability in villous explants and stimulated transforming growth factor beta 3 (TGFB3) expression consistent with promoter analyses showing that HIF1A transactivates TGFB3. These data demonstrate that during placental development, HIF1A is regulated by temporal and spatial changes in expression and association of molecules forming the multi-protein VHLCBC complex as well as prolyl hydroxylase activities.

HIF-1 mediates adaptation to hypoxia by actively downregulating mitochondrial oxygen consumption

The HIF-1 transcription factor drives hypoxic gene expression changes that are thought to be adaptive for cells exposed to a reduced-oxygen environment. For example, HIF-1 induces the expression of glycolytic genes. It is presumed that increased glycolysis is necessary to produce energy when low oxygen will not support oxidative phosphorylation at the mitochondria. However, we find that while HIF-1 stimulates glycolysis, it also actively represses mitochondrial function and oxygen consumption by inducing pyruvate dehydrogenase kinase 1 (PDK1). PDK1 phosphorylates and inhibits pyruvate dehydrogenase from using pyruvate to fuel the mitochondrial TCA cycle. This causes a drop in mitochondrial oxygen consumption and results in a relative increase in intracellular oxygen tension. We show by genetic means that HIF-1-dependent block to oxygen utilization results in increased oxygen availability, decreased cell death when total oxygen is limiting, and reduced cell death in response to the hypoxic cytotoxin tirapazamine.

Nitric oxide augments fetal pulmonary artery endothelial cell angiogenesis in vitro

Growth and development of the lung normally occur in the low oxygen environment of the fetus. The role of this low oxygen environment on fetal lung endothelial cell growth and function is unknown. We hypothesized that low oxygen tension during fetal life enhances pulmonary artery endothelial cell (PAEC) growth and function and that nitric oxide (NO) production modulates fetal PAEC responses to low oxygen tension. To test this hypothesis, we compared the effects of fetal (3%) and room air (RA) oxygen tension on fetal PAEC growth, proliferation, tube formation, and migration in the presence and absence of the NO synthase (NOS) inhibitor N(omega)-nitro-l-arginine (LNA), and an NO donor, S-nitroso-N-acetylpenicillamine (SNAP). Compared with fetal PAEC grown in RA, 3% O(2) increased tube formation by over twofold (P < 0.01). LNA treatment reduced tube formation in 3% O(2) but had no affect on tube formation in RA. Treatment with SNAP increased tube formation during RA exposure to levels observed in 3% O(2). Exposure to 3% O(2) for 48 h attenuated cell number (by 56%), and treatment with LNA reduced PAEC growth by 44% in both RA and 3% O(2). We conclude that low oxygen tension enhances fetal PAEC tube formation and that NO is essential for normal PAEC growth, migration, and tube formation. Furthermore, we conclude that in fetal cells exposed to the relative hyperoxia of RA, 21% O(2), NO overcomes the inhibitory effects of the increased oxygen, allowing normal PAEC angiogenesis and branching. We speculate that NO production maintains intrauterine lung vascular growth and development during exposure to low O(2) in the normal fetus. We further speculate that NO is essential for pulmonary angiogenesis in fetal animal exposed to increased oxygen tension of RA and that impaired endothelial NO production may contribute to the abnormalities of angiogenesis see in infants with bronchopulmonary dysplasia.

Portal Flow and Arterioportal Shunting after Transjugular Intrahepatic Portosystemic Shunt Creation

It was postulated that a transjugular intrahepatic portosystemic shunt (TIPS) produces arterioportal shunting and accounts for reversed flow in the intrahepatic portal veins (PVs) after creation of the TIPS. This study sought to quantify this shunting in patients undergoing TIPS creation and/or revision with use of a direct catheter-based technique and by measuring changes in blood oxygenation within the TIPS and the PV. This prospective study consisted of 26 patients. Median Model for End-stage Liver Disease and Child-Pugh scores were 13 and 9, respectively. Primary TIPS creation was attempted in 21 patients and revision of failing TIPS was undertaken in five. In two patients, TIPS creation was unsuccessful. All TIPS creation procedures but one were performed with use of polytetrafluoroethylene-covered stent-grafts. Flow within the main PV (Q(portal)) was measured with use of a retrograde thermodilutional catheter before and after TIPS creation/revision, and TIPS flow (Q(TIPS)) was measured at procedure completion. The amount of arterioportal shunting was assumed to be the increase between final Q(portal) and Q(TIPS), assuming Q(TIPS) was equivalent to the final Q(portal) plus the reversed flow in the right and left PVs. Oxygen saturation within the TIPS and the PV was determined from samples obtained during TIPS creation and revision. Mean Q(portal) before TIPS creation was 691 mL/min; mean Q(portal) after TIPS creation was 1,136 mL/min, representing a 64% increase (P = .049). Mean Q(TIPS) was 1,631 mL/min, a 44% increase from final Q(portal) (P = .0009). Among cases of revision, baseline Q(portal) was 1,010 mL/min and mean Q(portal) after TIPS revision was 1,415 mL/min, a 40% increase. Mean Q(TIPS) was 1,693 mL/min, a 20% increase from final Q(portal) (P = .42). Arterioportal shunting rates were 494 mL/min after TIPS creation and 277 mL/min after TIPS revision, representing 30% of total Q(TIPS) after TIPS creation and 16% of Q(TIPS) after TIPS revision. No increase in oxygen tension or saturation was seen in the PV or TIPS compared with initial PV levels. Q(TIPS) did not correlate with the portosystemic gradient. TIPS creation results in significant arterioportal shunting, with less arterioportal shunting seen among patients who undergo TIPS revision. Further work is necessary to correlate Q(TIPS) with the risk of hepatic encephalopathy and liver failure.

Perflubron emulsion increases subcutaneous tissue oxygen tension in rats

ABSTRACTOxygen plays a central role in wound healing. The hypothesis of this study was that the administration of a perfluorocarbon emulsion, which dissolves oxygen will increase subcutaneous tissue oxygen tension (PsqO2) in normovolemic and hemorrhaged animals. In the first set of experiments, PsqO2 was measured with a polarographic oxygen electrode along the dorsum of normovolemic‐anesthetized rats (n=20) breathing supplemental oxygen. After baseline equilibration of tissue oxygen tension, perflubron emulsion (n=12) or saline control (n=8) was administered. Perflubron administration increased PsqO2 by 32.0 ± 7.2 mmHg, whereas saline administration had no effect (−1.0 ± 6.6 mmHg). In a second set of experiments, PsqO2 (n=12) was measured in rats breathing 100 percent oxygen after being bled 20 percent of their blood volume. Creating hypovolemic rats allowed for the study of the effect of perflubron emulsion administration on subcutaneous oxygen tension in tissues with compromised blood flow. Perflubron emulsion was only effective at increasing oxygen tension in tissues with oxygen extractions less then 2.0 vol%. These findings agree with those results analytically predicted based on the oxygen solubility of the perflubron emulsion.

Intraretinal Oxygen Distribution in the Monkey Retina and the Response to Systemic Hyperoxia

To measure the intraretinal oxygen distribution and consumption in the fovea, the parafovea, and the inferior retina in the monkey eye and determine the influence of graded systemic hyperoxia. Oxygen sensitive microelectrodes were used to measure the oxygen tension as a function of depth through the retina in anesthetized monkeys (n = 8) under normoxic and hyperoxic conditions. Oxygen consumption rates in the avascular regions of retina were determined by fitting the oxygen profiles to established oxygen consumption models. Under normoxic conditions, in the foveal area, the intraretinal oxygen distribution reflected the absence of retinal capillaries and the predominantly choroidal origin of retinal oxygenation. A similar shape of oxygen distribution was seen in the parafoveal retina with the addition of local perturbations in the inner retina attributed to the presence of retinal capillaries. In the inferior retina the same general shape was found. Oxygen consumption in the outer retina was higher in the parafoveal region, and the minimum oxygen tension was lower. During hyperoxia, choroidal oxygen levels in all areas increased dramatically, but the increase in oxygen tension in the inner retina was much less. The avascular nature of the foveal area allowed oxygen consumption analysis of both the inner and outer retina and showed that inner retinal oxygen consumption increased significantly during hyperoxic ventilation to a level equivalent to that of the outer retina. In the outer retina of the monkey the P(O2) minimum is lower, and the oxygen consumption rate is higher in the parafoveal region. During systemic hyperoxia, outer retinal oxygen consumption is unaffected, but in the foveal area, total oxygen consumption increases. This regulation of oxygen consumption in the monkey retina is comparable to that reported in lower mammals and may represent an important mechanism in retinal homeostasis.

Enhanced Survival of MDS Progenitor Cells under Hypoxic Conditions.

The myelodysplastic syndromes (MDS) are a diverse group of primary bone marrow disorders characterized by persistent cytopenias and a variable rate of progression to acute myelogenous leukemia (AML). A consistent finding in the bone marrow of patients with MDS is increased apoptosis and a defect in terminal differentiation of maturing cells. We hypothesized that MDS cells are unable to tolerate the increasing oxygen tension experienced during hematopoiesis as cells move out of the hypoxic stem cell niche and this intolerance leads to increased apoptosis. In order to test this hypothesis, we have studied the growth in methylcellulose colonies of bone marrow progenitors in eight samples from patients with MDS at normoxia (21% O2) and at hypoxia (3% O2). In 5/8 samples there is a greater than 8-fold (range 8.5–15.9) increase in colony number under hypoxic conditions. The remaining three MDS samples, while only demonstrating an average increase in colony number of 2.4-fold, demonstrated significantly larger colonies of various lineages. Comparatively, in six normal patient samples the augmentation averaged 1.5-fold and never exceeded 2-fold, with a statistically significant (p=0.01) difference in the mean fold-change between normal and MDS marrows. These results suggest that MDS cells have an impaired ability to tolerate increasing levels of oxygen and also suggests a methodology for expansion of MDS cells for further biochemical analysis of this phenotype. The cellular response to hypoxia is modulated in part through the mitochondrion and significant genetic data suggests that a mitochondrial abnormality is associated with MDS. In order to examine mitochondrial potential in MDS progenitor cells, we have adapted a multiparameter flow cytometric strategy for the simultaneous analysis of mitochondrial potential (ψm) in defined hematopoietic progenitors using the potentiometric dye tetramethyl rhodamine ethyl ester (TMRE). These studies confirm that in normal bone marrow samples, ψm increases with development from the hematopoietic stem cell (HSC) to differentiated progenitor cells. Analysis of marrow from a patient with early MDS (refractory cytopenia with multilineage dysplasia, IPSS Low) and a 10-fold increase in colony number under hypoxic conditions demonstrates the absence of this progressive increase in ψm through myeloid development when compared to normal controls. Additionally, two samples from patients with progression of their MDS to frank AML demonstrated a markedly elevated ψm in the blast cells, consistent with the elimination of the mitochondrial pathology during the genesis of the leukemic clone. Analysis of mitochondrial potential in additional MDS samples is ongoing and will be presented. These data suggest that a defect in the mitochondrial adaptation to increasing oxygen tension experienced during hematopoietic differentiation may explain the impaired growth of MDS progenitor cells.

Hyperoxia and angiogenesis

We hypothesized that tissue hyperoxia would enhance and hypoxia inhibit neovascularization in a wound model. Therefore, we used female Swiss-Webster mice to examine the influence of differential oxygen treatment on angiogenesis. One milliliter plugs of Matrigel, a mixture of matrix proteins that supports but does not itself elicit angiogenesis, were injected subcutaneously into the mice. Matrigel was used without additive or with added vascular endothelial growth factor (VEGF) or anti-VEGF antibody. Animals were maintained in hypoxic, normoxic, or one of four hyperoxic environments: hypoxia -- 13 percent oxygen at 1 atmosphere absolute (ATA); normoxia -- 21 percent oxygen at 1 ATA; hyperoxia -- (groups a-d) 100 percent oxygen for 90 minutes twice daily at the following pressures: Group a, 1 ATA; Group b, 2 ATA; Group c, 2.5 ATA; Group d, 3.0 ATA. Subcutaneous oxygen tension was measured in all groups. The Matrigel was removed 7 days after implantation. Sections were graded microscopically for the extent of neovascularization. Angiogenesis was significantly greater in all hyperoxic groups and significantly less in the hypoxic group compared with room air-exposed controls. Anti-VEGF antibody abrogated the angiogenic effect of both VEGF and increased oxygen tension. We conclude that angiogenesis is proportional to ambient pO(2) over a wide range. This confirms the clinical impression that angiogenesis requires oxygen. Intermittent oxygen exposure can satisfy the need for oxygen in ischemic tissue.

Cerebral Blood Flow and Brain Oxygenation in Rats Breathing Oxygen under Pressure

Hyperbaric oxygen (HBO(2)) increases oxygen tension (PO(2)) in blood but reduces blood flow by means of O(2)-induced vasoconstriction. Here we report the first quantitative evaluation of these opposing effects on tissue PO(2) in brain, using anesthetized rats exposed to HBO(2) at 2 to 6 atmospheres absolute (ATA). We assessed the contribution of regional cerebral blood flow (rCBF) to brain PO(2) as inspired PO(2) (PiO(2)) exceeds 1 ATA. We measured rCBF and local PO(2) simultaneously in striatum using collocated platinum electrodes. Cerebral blood flow was computed from H(2) clearance curves in vivo and PO(2) from electrodes calibrated in vitro, before and after insertion. Arterial PCO(2) was controlled, and body temperature, blood pressure, and EEG were monitored. Scatter plots of rCBF versus PO(2) were nonlinear (R(2)=0.75) for rats breathing room air but nearly linear (R(2)=0.88-0.91) for O(2) at 2 to 6 ATA. The contribution of rCBF to brain PO(2) was estimated at constant inspired PO(2), by increasing rCBF with acetazolamide (AZA) or decreasing it with N-nitro-L-arginine methyl ester (L-NAME). At basal rCBF (78 mL/100 g min), local PO(2) increased 7- to 33-fold at 2 to 6 ATA, compared with room air. A doubling of rCBF increased striatal PO(2) not quite two-fold in rats breathing room air but 13- to 64-fold in those breathing HBO(2) at 2 to 6 ATA. These findings support our hypothesis that HBO(2) increases PO(2) in brain in direct proportion to rCBF.

143 Estrogen Attenuates Oligodendrocyte Apoptosis Caused by Hyperoxia

Background: In the developing human brain, periventricular leukomalacia (PVL) is the predominant white matter injury underlying the development of cerebral palsy. Our group recently showed that a rise of oxygen tissue tension is a powerful trigger to initiate apoptosis in premature and immature, but not in mature oligodendrocytes (OL). Estrogen plays an important role in the development and function of the cerebral nervous system, and estrogen receptors show a specific distribution with high density arround the third ventricle, the region where OL progenitors are generated. We therefore examined the effects of estrogen on hyperoxia-induced cell death in cultured rat immature oligodendroglia cells. Methods: OLN-93 cells, derived from spontaneously transformed cells of 5- to 10-day-old primary rat brain oligodendrocytes were subjected to 24–72 h of hyperoxia (80% O2) in the presence or absence of estrogen (17beta-estradiol) at various concentrations. These immature oligodendrocytes resemble an intermediate stage between oligodendrocyte precursors and mature oligodendrocytes. Flow cytometry was used to assess apoptosis via annexin-V, anti-activated caspase-3 antibody, and propidium iodide staining. Oligodendrocyte maturation status and caspase-3 activation were confirmed by immunochemistry. Results: In immature rat oligodendrocytes (OLN-93), apoptosis was detected at various stages (early: annexin-V, effector: caspase-3) after 24–72 h of hyperoxia (80% O2). Cell death was reduced up to 30% by preincubation with estrogen at a concentration optimum of 10–7 M estrogen. Conclusion: Estrogen reduces apoptosis in immature oligodendrocytes exposed to hyperoxia. The sudden drop in circulating estrogen of placental origin after mammalian birth may aggravate the susceptibility of immature oligodendrocytes to increased oxygen tension. Supported by grants from the German Federal Department of Education and Research (BMBF, # 01 ZZ 0101), Bonn, and the Ernst Schering Research Foundation, Berlin.

Lung Fluid Balance During Development

After completing this article, readers should be able to: 1. Describe the major driving forces for liquid formation in and liquid removal from the lumen of the fetal lung. 2. Delineate the role of a normal volume of liquid within the lumen of the fetal lung for normal lung growth during development. 3. Describe the effects of labor on Na,K-ATPase activity in lung epithelial cells and clearance of lung liquid after birth. 4. Explain the route by which most residual liquid drains from the lung. 5. List the conditions associated with preterm birth that may contribute to delayed removal of fetal lung liquid. Curious students of mammalian development recognized more than a century ago that the lungs are filled with liquid during fetal life, (1) but the origin of that liquid remained obscure until 1948, when an unexpected discovery by two French scientists dispelled the prevailing view that fetal lung liquid derived from intrauterine inhalation of amniotic fluid. In studies designed to explore the ontogeny of the pituitary-adrenal axis, tracheal ligation of fetal rabbits for 9 days resulted in fluid distention of the lungs. (2) This serendipitous finding, subsequently confirmed and embellished by others, (3)(4)(5) established that the fetal lung itself, rather than the amniotic sac, is the source of the liquid that fills the lung during development. This liquid forms a slowly expanding structural template that prevents collapse and promotes growth of the fetal lung. Subsequent studies, conducted mostly with sheep, showed that liquid in the lumen of the fetal lung forms as a result of chloride secretion in the respiratory epithelium, (6)(7) a process that can be inhibited by diuretics that block Na,K-2Cl cotransport. (8)(9) In vitro experiments using cultured explants of lung tissue and monolayers of epithelial cells harvested from human fetal lung have …

Dynamics of tumour cords following changes in oxygen availability: A model including a delayed exit from quiescence

In this paper, we investigate the formation of tumour cords (cylindrical arrangements of tumour cells around blood vessels) within a tumour tissue. The shrinkage and the expansion of the cord following a stepwise decrease or, respectively, increase of oxygen tension in the blood vessel are also described. The model represents the tumour cell population as a continuum and includes two novel aspects with respect to our previous works: cell death does not occur instantaneously, but is preceded by a reversible prenecrotic state; the recruitment of cells from quiescence into proliferation may require a recovery period. Model predictions are compared with experimental data reported by Hirst et al..

Effects of isovolaemic haemodilution on oxygenation of liver and skeletal muscle

Hydroxyethyl starch is frequently used for volume substitution during surgical procedures and for isovolaemic haemodilution. Haemodilution has also been shown to improve tissue oxygen tension in skeletal muscle: However, effects of this volume substitute on tissue oxygen tension of the liver during haemodilution remains unknown. Fourteen foxhounds were anaesthetized with fentanyl/midazolam and mechanically ventilated with 30% oxygen. Following splenectomy animals were randomly assigned to a control group without haemodilution but fluid substitution with Ringer's lactate (Group C) or underwent isovolaemic haemodilution to a haematocrit of 25% with hydroxyethyl starch 70/0.5 (Group H). Haemodynamic parameters and oxygen transport during 100 min following isovolaemic haemodilution were measured. Liver oxygen tension was recorded using a flexible polarographic electrode tonometer, whereas in the muscle a polarographic needle probe was used. Animal characteristics and baseline haematocrit were similar in both groups. At baseline the tissue oxygen tension of liver and skeletal muscle were not different between groups. Haemodilution with hydroxyethyl starch 70/0.5 provided augmentation of mean liver tissue oxygen tension (baseline: 46 +/- 13 mmHg; 20 min: 60.3 +/- 12 mmHg; 60 min: 60 +/- 16 mmHg; 100 min: 63 +/- 16 mmHg; P < 0.05 vs. baseline), while oxygen tensions in Group C remained unchanged (baseline: 48 +/- 16 mmHg; 20 min: 52 +/- 19 mmHg; 60 min: 49 +/- 12 mmHg; 100 min: 52 +/- 16 mmHg) and no differences could be detected between groups. Oxygen tension in skeletal muscle changed as follows: Group H - baseline: 24 +/- 32 mmHg; 20 min: 32 +/- 3 mmHg; 60 min: 33 +/- 7 mmHg; 100 min: 33 +/- 11 mmHg. Group C - baseline: 22 +/- 6 mmHg; 20 min: 21 +/- 3 mmHg; 60 min: 24 +/- 4 mmHg; 100 min: 18 +/- 4 mmHg (P < 0.05 vs. baseline, p < 0.05 vs. Group C). In this animal model, isovolaemic haemodilution with hydroxyethyl starch 70/0.5 increased tissue oxygen tension in liver and skeletal muscle in comparison with baseline values. However, when compared between groups haemodilution only resulted in an increase of tissue oxygen tension in the muscle but not in the liver.

Oxygen increases ductus arteriosus smooth muscle cytosolic calcium via release of calcium from inositol triphosphate-sensitive stores

In utero, blood shunts away from the lungs via the ductus arteriosus (DA) and the foramen ovale. After birth, the DA closes concomitant with increased oxygen tension. The present experimental series tests the hypothesis that oxygen directly increases DA smooth muscle cell (SMC) cytosolic calcium ([Ca(2+)](i)) through inactivation of a K(+) channel, membrane depolarization, and entry of extracellular calcium. To test the hypothesis, DA SMC were isolated from late-gestation fetal lambs and grown to subconfluence in primary culture in low oxygen tension (25 Torr). DA SMC were loaded with the calcium-sensitive fluorophore fura-2 under low oxygen tension conditions and studied using microfluorimetry while oxygen tension was acutely increased (120 Torr). An acute increase in oxygen tension progressively increased DA SMC [Ca(2+)](i) by 11.7 +/- 1.4% over 40 min. The effect of acute normoxia on DA SMC [Ca(2+)](i) was mimicked by pharmacological blockade of the voltage-sensitive K(+) channel. Neither removal of extracellular calcium nor voltage-operated calcium channel blockade prevented the initial increase in DA SMC [Ca(2+)](i). Manganese quenching experiments demonstrated that acute normoxia initially decreases the rate of extracellular calcium entry. Pharmacological blockade of inositol triphosphate-sensitive, but not ryanodine-sensitive, intracellular calcium stores prevented the oxygen-induced increase in [Ca(2+)](i). Endothelin increased [Ca(2+)](i) in acutely normoxic, but not hypoxic, DA SMC. Thus acute normoxia 1) increases DA SMC [Ca(2+)](i) via release of calcium from intracellular calcium stores, and subsequent entry of extracellular calcium, and 2) potentiates the effect of contractile agonists. Prolonged patency of the DA may result from disordered intracellular calcium homeostasis.

Intraretinal oxygenation and oxygen consumption in the rabbit during systemic hyperoxia.

To determine the ability of the retina and choroid in the avascular region of the rabbit retina to regulate intraretinal oxygen levels during periods of acute systemic hyperoxia. Oxygen-sensitive microelectrodes were used to measure oxygen tension as a function of depth through the retina and choroid in anesthetized rabbits before and after stepwise incremental increases in inspired oxygen level. The extent of any oxygen increases throughout the retina and choroid was determined, and retinal oxygen consumption was determined by analyzing intraretinal oxygen distribution. Increases in systemic arterial oxygen in the rabbit resulted in significant increases in oxygen tension in the choroid and in all retinal layers. There was no apparent regulation of choroidal oxygen tension, as seen in the avascular retina of the guinea pig. Neither was there any increase in oxygen consumption within the retina, which has been shown to dampen the extent of raised oxygen levels in the inner retina in other mammals. Consequently, all retinal layers in the avascular area of the rabbit retina were exposed to high oxygen levels during systemic hyperoxia. The magnitude of the oxygen changes in all retinal layers was consistent with that in the preretinal vitreous. Unlike other mammals studied to date, the rabbit does not possess any regulatory mechanisms for controlling the intraretinal oxygen environment during acute increases in systemic arterial oxygen levels. This may well account for the reported vulnerability of the rabbit retina to toxic damage during extended periods of systemic hyperoxia.

Oxygen and the copper chaperone CCS regulate posttranslational activation of Cu,Zn superoxide dismutase

Oxidative stress leads to the up-regulation of many antioxidant enzymes including Cu,Zn superoxide dismutase (SOD1) via transcriptional mechanisms; however, few examples of posttranslational regulation are known. The copper chaperone for SOD1 (CCS) is involved in physiological SOD1 activation, and its primary function is thought to be delivery of copper to the enzyme. Data presented here are consistent with a previously uncharacterized function for CCS in the SOD1 pathway, namely mediating enzyme activation in response to increases in oxygen tension. Activity assays with pure proteins and cell extracts reveal that O(2) (or superoxide) is required for activation of SOD1 by CCS. Dose-response studies with a translational blocking agent demonstrate that the cellular oxidative response to O(2) is multitiered: existing apo-pools of SOD1 are activated by CCS in the early response, followed by increasing expression of SOD1 protein with persistent oxidative stress. This CCS function provides oxidant-responsive posttranslational regulation of SOD1 activity and may be relevant to a wide array of physiological stresses that involve a sudden elevation of oxygen availability.

Hemodynamic Response to Sildenafil, Nitric Oxide, and Iloprost in Primary Pulmonary Hypertension

Different vasodilators and different routes of application are used for the treatment of primary pulmonary hypertension (PPH). Recently, sildenafil, a phosphodiesterase-V inhibitor, has been shown to have beneficial hemodynamic effects in PPH. However, the hemodynamic effects of sildenafil have not been characterized and compared to other vasodilators such as inhaled nitric oxide (iNO) or iloprost in PPH in the same group of patients. We investigated prospectively 10 consecutive patients with PPH using iNO, iloprost aerosol, and oral sildenafil to test acute hemodynamic response during right-heart catheterization. iNO, iloprost aerosol, and sildenafil caused a significant fall of mean pulmonary artery pressure and pulmonary vascular resistance (PVR) [p < 0.05]. Correspondingly, cardiac output and mixed venous saturation increased slightly in all groups. Systemic arterial pressure and vascular resistance were mainly unaltered. Using a PVR reduction of > or =20% to define a significant response, 7 of 10 patients were responders to iloprost aerosol, whereas 4 of 10 patients responded to iNO and oral sildenafil. Improvement of oxygenation as indicated by an increase of arterial oxygen tension was observed with iloprost aerosol (p < 0.01). All of the three substances, iNO, iloprost aerosol, and oral sildenafil, significantly improved pulmonary hemodynamics in patients with PPH. The most prominent hemodynamic effects and improvement of oxygenation were observed with iloprost aerosol.

In situ monitoring of heart rates in shore crabs Carcinus maenas in two tidal estuaries: Effects of physico-chemical parameters on tidal and diel rhythms

Heart rates were monitored in situ in the shore crab, Carcinus maenas, in relation to variations in depth, salinity, oxygen tension, temperature, light intensity and pH. Experiments were performed in the Looe Estuary, Cornwall, England and in Batson Creek in the Salcombe-Kingsbridge Estuary, Devon, England. Experiments in the Looe Estuary were conducted in the vicinity of a storm water storage discharge whereas the experiments in Batson Creek were performed on a clean site. Tidal rhythms in heart rates were commonly detected but diel rhythms in heart rate were also observed frequently. Both types of rhythm were more evident in animals from Batson Creek than from Looe. In Batson Creek, 12 out of 15 crabs expressed tidal rhythms in heart rate, whereas 6 out of 15 crabs expressed diel rhythms. In the two studies in the Looe Estuary, 6 out of 15 crabs and 3 out of 15 crabs expressed tidal and diel rhythm in heart rate, respectively. At both experimental sites, heart rates were positively correlated with increasing changes in depth and salinity, whereas heart rates were negatively correlated with light intensity. In addition, heart rates appeared to be positively correlated with increasing oxygen tension in the experiments performed in the Looe Estuary. The study suggests that depth and oxygen availability are more important to in situ heart rates in shore crabs within tidal estuaries than are salinity, light intensity and pH. Also, sewage discharge appears to cause an acute increase in heart rate, which may affect expression of biological rhythms in shore crabs.