Age and sex differences in brain gene expression in neonatal rats.

Gene expression in the central nervous system is highly region-specific. We tested the hypothesis that certain developmental biomarkers could be detected in the whole brain or in cortical, subcortical or cerebellar structures. Brain gene expressions of male and female rats at birth, 3 days, and 10 days of age were measured by microarray technique ( approximately 10 K genes; n=9/category). We found 53 significantly up-regulated and 8 down-regulated genes at 10 days of age, relative to birth and 3 days of age. The whole brain, however, showed no significant sex differences in gene expression patterns up to 10 days of age. Ten genes with the highest up-regulation, and 5 down-regulated genes were further confirmed by quantitative real-time PCR (Q-PCR), using the whole brain, cortices, subcortical structures, and cerebellum. The Q-PCR confirmed genes are known to be involved in neuronal differentiation, axonal myelination and growth, neurotransmission and glycolytic pathways. With a few exceptions, the expression levels of Q-PCR confirmed genes were significantly different in the whole brain, compared to other regions. In a separate study, we tested the potential utility of the Q-PCR confirmed genes, as whole brain biomarkers, after a six-hour exposure to hyperoxia (>98% oxygen breathing) in 10 days old rats. This relatively mild oxidative challenge created a 3.5-fold increase in the expression of T-cell receptor beta Variable 8.3b, known to have regulatory function during development. We suggest that genes displaying significant expression in the whole brain, regardless of their origin, could be used to screen normal brain development in neonatal rat models of experimental neurology.

Hyperbaric oxygen therapy protects against mitochondrial dysfunction and delays onset of motor neuron disease in Wobbler mice.

The Wobbler mouse is a model of human motor neuron disease. Recently we reported the impairment of mitochondrial complex IV in Wobbler mouse CNS, including motor cortex and spinal cord. The present study was designed to test the effect of hyperbaric oxygen therapy (HBOT) on (1) mitochondrial functions in young Wobbler mice, and (2) the onset and progression of the disease with aging. HBOT was carried out at 2 atmospheres absolute (2 ATA) oxygen for 1 h/day for 30 days. Control groups consisted of both untreated Wobbler mice and non-diseased Wobbler mice. The rate of respiration for complex IV in mitochondria isolated from motor cortex was improved by 40% (P<0.05) after HBOT. The onset and progression of the disease in the Wobbler mice was studied using litters of pups from proven heterozygous breeding pairs, which were treated from birth with 2 ATA HBOT for 1 h/day 6 days a week for the animals' lifetime. A "blinded" observer examined the onset and progression of the Wobbler phenotype, including walking capabilities ranging from normal walking to jaw walking (unable to use forepaws), and the paw condition (from normal to curled wrists and forelimb fixed to the chest). These data indicate that the onset of disease in untreated Wobbler mice averaged 36+/-4.3 days in terms of walking and 40+/-5.7 days in terms of paw condition. HBOT significantly delayed (P<0.001 for both paw condition and walking) the onset of disease to 59+/-8.2 days (in terms of walking) and 63+/-7.6 days (in terms of paw condition). Our data suggest that HBOT significantly ameliorates mitochondrial dysfunction in the motor cortex and spinal cord and greatly delays the onset of the disease in an animal model of motor neuron disease.

Plasma colloid osmotic pressure in healthy infants.

BACKGROUND: The plasma colloid osmotic pressure (COP) plays a major role in transcapillary fluid balance. There is no information on plasma COP of healthy infants beyond the first post-natal week. The normal COP in healthy adult subjects (25 mmHg) is currently also applied as a reference value for healthy infants. This study was designed to test whether plasma COP values in healthy infants are the same as those in normal adults. METHODS: Plasma COP was measured in 37 male and female healthy infants from 1 to 11 months old. For this purpose, 1 ml blood was collected during the patient's regularly scheduled visit if the patient required any type of blood test for routine laboratory analyses. RESULTS: Plasma COP levels correlated slightly with increasing age from 1 to 9 months old (linear regression analysis; r2 = 0.1, P < 0.049). We found no correlation between plasma COP and body weight at the same age (r2 = 0.05, P = 0.155). The mean and standard deviation of COP in all infants was 25.1 +/- 2.6 mmHg, which is almost identical to an average COP of 25 mmHg in healthy adult subjects. Arbitrary division of the infants into three different age groups (1-3 months [n = 11], 5-8 months [n = 13] and 9-11 months [n = 13]) showed an average increase of approximately 2 mmHg in COP of 9-month-old to 11-month-old infants, compared with 1-month-old to 3-month-old infants (one-way analysis of variance; P = 0.26). There was no gender difference in the COP level (unpaired t-test), with an average of 25.1 +/- 2.4 mmHg in 19 male infants compared with 25.2 +/- 2.9 in 18 female infants. The 95% confidence interval for COP in both male and female infants (n = 37) was between 24.3 to 26.0 mmHg, ranging from 19.5 to 30.3 mmHg, with a median value of 25.2 mmHg. CONCLUSIONS: The data accept the null hypothesis that the COP range in infants younger than 1 year old is similar to those observed in adult subjects. Our observations, compared with previously reported neonatal COP values, suggest that there is a sharp increase in COP within the first months after birth.

Pulmonary and extrapulmonary effects of increased colloid osmotic pressure during endotoxemia in rats.

OBJECTIVES: We tested the hypothesis that an increase in the blood colloid osmotic pressure (COP) that is maintained during early-stage endotoxemia may decrease fluid flux across capillaries and may reduce pulmonary and multiple-organ edema. DESIGN: Prospective study. SETTINGS: Research laboratory in a hospital. SUBJECTS: Male albino Sprague-Dawley rats. INTERVENTIONS: Rats were anesthetized with pentobarbital, underwent tracheotomies, were cannulated in the femoral vein and artery, and were randomly assigned to the following four groups comprising 11 rats each: group I, controls (saline solution treatment); group II, albumin treatment (three doses of 1 g/kg 25% human albumin every 2 h); group III, endotoxin treatment with a single IV dose of 4 mg/kg endotoxin; and group IV, endotoxin and albumin-treatment (4 mg/kg endotoxin plus albumin treatment). Experiments lasted for 6 h while fluid intake was equally maintained in all groups. MEASUREMENTS AND RESULTS: COP and other variables were measured every 2 h. To determine the water content of an organ, after the rat was killed, the lung, heart, kidney, intestine, and liver were removed. Albumin treatment alone (group II) generated significant increases in COP (maximum, 58% from the baseline measurement) but did not change the water content of the organ, compared with saline solution-treated controls. Endotoxin-treated rats (group III) developed significant reductions in COP, with significant increases in pulmonary, renal, and heart water content compared with controls. Albumin treatment in endotoxemic rats (group IV) significantly increased the COP without improving the endotoxemia-induced organ edema. Pulmonary edema, however, was increased further, compared with endotoxemia alone. CONCLUSIONS: COP elevation by albumin administration during the early stage of endotoxemia does not ameliorate pulmonary or multiple-organ edema and may aggravate pulmonary edema.

Effects of arteriovenous extracorporeal therapy on hemodynamic stability, ventilation, and oxygenation in normal lambs.

OBJECTIVE: To evaluate hemodynamic stability and gas exchange in a neonatal animal model of pumpless arteriovenous extracorporeal membrane oxygenation (AV-ECMO) with extracorporeal shunt flow of up to 15% of cardiac output during variable ventilation and oxygenation. DESIGN: Prospective study. SETTING: Research laboratory in a hospital. SUBJECTS: Seven lambs (5.5 +/- 0.6 kg, mean +/- sd). INTERVENTIONS: The lambs initially were anesthetized by 50 mg/kg ketamine intravenously. After tracheostomy, the lambs were mechanically ventilated and paralyzed by using 1 mg/kg vecuronium bromide followed by 0.1 mg.kg(-1).hr(-1). One femoral vein was cannulated with a pulmonary artery flotation catheter and used for cardiac output and pulmonary artery pressure measurements. A femoral artery was cannulated for measuring mean arterial blood pressure, measuring heart rate, and blood sampling for gas exchange analyses. Finally, the right internal jugular vein and carotid artery were cannulated and used for the AV-ECMO. Normothermia (38 +/- 0.5 degrees C), fluid balance (5 mL.kg(-1).hr(-1) normal saline), and anesthesia (5 mg.kg(-1).hr(-1), intravenous ketamine) were maintained. Ventilator settings were adjusted to establish a baseline Paco2 (25-35 mm Hg) at an Fio2 of 0.4. The AV-ECMO circuit was established by using a hollow fiber oxygenator, primed with maternal sheep blood (150-200 mL). MEASUREMENTS AND MAIN RESULTS: The physiologic effects of the AV-ECMO shunt were evaluated at 15, 25, and 40 mL.kg(-1).hr(-1) ECMO flow, corresponding roughly to 4%, 8%, and 15% of the cardiac output values. The baseline minute volume was maintained during stepwise increases in arteriovenous shunt. A significant increase in endogenous cardiac output occurred at arteriovenous shunt of 25 and 40 mL.kg(-1).hr(-1) (analysis of variance followed by Tukey-Kramer multiple comparisons test), which was attributed to a significant increase of 30% in the heart rate. Effective cardiac output (difference between the thermodilution value and the AV-ECMO flow rate) and mean arterial blood pressure were not significantly changed. CO2 removal, measured at 15% arteriovenous shunt, was significantly increased with decreasing ventilation to 25% and 50% of the baseline (analysis of variance and Tukey-Kramer test). Oxygenation through the membrane was measured after reducing inspired Fio2 from 0.4 to 0.21, 0.15, and 0.10 with 15% arteriovenous shunt and baseline minute ventilation. Oxygen delivery by the oxygenator was significantly increased at Fio2 of 0.10, providing a maximum of 19.5% of the total oxygen consumption at an arterial hemoglobin-oxygen saturation of 60%. CONCLUSIONS: Healthy lambs are capable of maintaining effective cardiac output in the presence of moderate arteriovenous shunts (15%). AV-ECMO may provide efficient ventilatory support in the neonatal population with hypercapnia. The amount of oxygen delivery with AV-ECMO depends on arterial desaturation.

Intratracheal pulmonary ventilation versus conventional mechanical ventilation: continuous carinal pressure monitoring at low and high flows and frequencies.

We continuously measured proximal and carinal pressures at low and high flow rates and frequencies during conventional mechanical ventilation (CMV) and intratracheal pulmonary ventilation (ITPV), using an artificial lung. The proximal peak inspiratory pressure (PIP), carinal PIP, proximal positive end expiratory pressure (PEEP), and carinal PEEP, or negative end expiratory pressure (NEEP), were measured during simulated CMV and ITPV. Two levels of frequency (30 and 90 per min) and two gas flow rates (3 and 6 L/min) were examined, in both dry and humid states (four combinations of gas flow and frequency at each state). The gas flow and inspiratory time were held constant throughout the CMV and ITPV trials. Humidification of the ventilatory circuit during ITPV prevented the accurate measurement of carinal pressures. This problem was solved by introducing a continuous "bias flow" of 11 ml/min into the pressure monitoring line. A combination of low gas flow and low frequency with CMV showed no significant differences between the proximal and carinal PIP, as well as the proximal and carinal PEEP. The same combination with ITPV, however, resulted in a significantly lower carinal PIP and PEEP, compared to proximal PIP and PEEP. Carinal PIP and PEEP during ITPV were also significantly lower than those observed during CMV with a low flow and low frequency rates. During both CMV and ITPV, using a combination of a high flow rate with a high breathing frequency, carinal PIPs were significantly lower than proximal PIPs. ITPV, however, generated much larger differences between proximal and carinal PIPs than the CMV. A significant NEEP was generated at the carinal level during ITPV with high flow rates, both with high and low frequencies. The NEEP did not occur with a low gas flow, in combination with either a low frequency or a high frequency. The "bias flow" had no significant effect on carinal pressures. In conclusion, ITPV, compared with CMV, generates a significantly lower carinal PIP, but it may also generate carinal NEEP. For safety reasons, therefore, it is essential to monitor carinal pressures continuously in patients treated with ITPV.

Effect of hypothermia on ventilation in anesthetized, spontaneously breathing rats: theoretical implications for mechanical ventilation.

OBJECTIVE: To test if hypothermia, induced by a sustained pentobarbital anesthesia, in rats can reduce ventilatory demands without compromising pulmonary gas-exchange efficiency. DESIGN: Prospective study. SETTING: Research laboratory in a hospital. SUBJECTS: One group of 11 female Sprague Dawley rats. INTERVENTIONS: The rats were anesthetized with 45 mg/kg pentobarbital, tracheostomized and intubated; their femoral veins and arteries were cannulated. After surgery, anesthesia and fluid balance were maintained (10 mg/kg per h pentobarbital, and 5 ml/kg per h saline, i.v.). Rectal temperature, mean arterial blood pressure (MAP), and heart rate (HR) were continuously monitored. The respiratory variables and gas-exchange profiles were determined at 38 degrees C (normothermia), and during stepwise hypothermia at 37, 35, 33, 31 and 29 degrees C. The arterial pressure of carbon dioxide (PaCO2), pH and arterial pressure of oxygen (PaO2) during hypothermia were corrected at body temperature. MEASUREMENTS AND RESULTS: Graded systemic hypothermia, with maintained anesthesia, produced a strong correlation between reduction in the respiratory frequency and rectal temperature (r2 = 0.55; p < 0.0001; n = 66). The minute volume was significantly reduced, starting at 35 degrees C, without significant changes in the tidal volume (repeated measures of analyses of variance followed by Dunnett multiple comparisons test). No significant changes occurred in the PaCO2, pH, PaO2, hemoglobin oxygen saturation, the calculated arterial oxygen content and estimated alveolar-arterial oxygen difference during mild hypothermia (37-33 degrees C). The PaO2, however, was significantly reduced below 31 degrees C. The MAP remained stable at different levels of hypothermia, whereas HR was significantly reduced below 33 degrees C. CONCLUSIONS: Mild hypothermia in rats, induced by a sustained pentobarbital anesthesia, reduces ventilation without compromising arterial oxygenation or acid-base balance, as measured at body temperature. Theoretically, our observations in spontaneously breathing rats imply that a combination of mild hypothermia with anesthesia could be safely utilized to maintain adequate ventilation, using relatively low minute ventilation. We speculate that such a maneuver, if applied during mechanical ventilation, may prevent secondary pulmonary damage by allowing the use of lower ventilator volume-pressure settings.

Intratracheal pulmonary ventilation in a rabbit lung injury model: continuous airway pressure monitoring and gas exchange efficacy.

OBJECTIVES: To compare carinal pressures vs. proximal airway pressures, and gas exchange efficacy with a constant minute volume, in lung-injured rabbits during conventional mechanical ventilation (CMV) and intratracheal pulmonary ventilation (ITPV); and to evaluate performance of a prototype ITPV gas delivery and continuous airway pressure monitoring system. DESIGN: Prospective controlled study. SETTING: Animal research laboratory at a teaching hospital. SUBJECTS: Sixteen adult female rabbits. INTERVENTIONS: Anesthetized rabbits were tracheostomized with a multilumen endotracheal tube. Anesthesia and muscle relaxation were maintained continuously throughout the study. Proximal airway pressures and carinal pressures were recorded continuously. The injection port of the multilumen endotracheal tube was used for the carinal pressure monitoring. To prevent obstruction of the port, it was flushed with oxygen at a rate of 11 mL/min. CMV was initiated with a pressure-limited, time-cycled ventilator set at an FiO2 of 1.0 and at a flow of 1.0 L/kg/min. The pressure limit of the ventilator was effectively disabled. A normal baseline for arterial blood gases was achieved by adjusting the inspiratory/expiratory time ratios. ITPV was established using a flow of 1.0 L/kg/min through a reverse thrust catheter, at the same baseline and inspiratory/expiratory ratio. Carinal positive end-expiratory pressure was maintained at a constant value of 2 cm H2O by adjusting the expiratory resistance of the ventilator circuit Lung injury was achieved over a 30-min period by three normal saline lavages of 5 mL/kg each. After lung injury, all animals were consecutively ventilated for 1 hr with CMV, for 1 hr with ITPV, and again for 1 hr with CMV. Six rabbits were ventilated at 30 breaths/min (group 1), and ten rabbits were ventilated at 80 breaths/min (group 2). Four rabbits in group 2 were subjected, 1 hr after return to CMV from ITPV, to another session of ITPV, with positive end-expiratory pressure gradually being increased to 4, 6, and 8 cm H2O for 15 mins each. RESULTS: No significant differences were observed in carinal peak inspiratory pressure between CMV and ITPV modes, at both low and high frequencies of breathing, indicating that the inspired tidal volume remained constant during both modes of ventilation. Significant gradients were noted between proximal airway and carinal peak inspiratory pressure during ITPV but not during CMV. Initiation of ITPV, at a flow of 1.0 L/kg/min, required an increase in the ventilator expiratory resistance to maintain a constant level of positive end-expiratory pressure (2 cm H2O) as measured at the carina. During ITPV, the PaCO2 was significantly reduced by 20% at 30 breaths/min (p < .05) and by 22% at 90 breaths/min (p < .01), compared with CMV. Arterial oxygenation was significantly enhanced with a positive end-expiratory pressure of 6 and 8 cm H2O (p < .05 and .001, respectively), compared with a positive end-expiratory pressure of 2 cm H2O during ITPV. All components of the new prototype gas delivery and airway pressure monitoring system functioned without failure, at least for 3 hrs of the CMV, ITPV, and CMV trials. CONCLUSIONS: ITPV in saline-lavaged, lung-injured rabbits at breathing frequencies of 30 and 80 breaths/min, compared with CMV at the same minute ventilation, can improve CO2 exchange. During ITPV, significant pressure gradients can develop between carinal and proximal airway pressures. Continuous carinal pressure monitoring is therefore necessary for the safe clinical application of ITPV. Reliable carinal pressure monitoring can be achieved by adding a small bias flow through the carinal pressure monitoring port. Although ITPV can remove CO2 from injured lungs efficiently, simultaneous addition of positive end-expiratory pressure can further improve arterial oxygenation.

Oxygen-carrying capacity during 10 hours of hypercapnia in ventilated dogs.

OBJECTIVE: To test if a relatively long-term exogenous hypercapnia, equivalent to those maintained during permissive hypercapnia, can persistently increase oxygen-carrying capacity in ventilated dogs. DESIGN: Prospective study. SETTING: Research laboratory in a hospital. SUBJECTS: Six mongrel dogs (3 males; 3 females). INTERVENTIONS: The dogs were anesthetized (30 mg/kg pentobarbital, i.v.), intubated, and cannulated in one femoral artery, one femoral vein, and the right jugular vein. The mean arterial blood pressure, heart rate, and mean pulmonary artery pressure were continuously recorded. Anesthesia, fluid balance, and normothermia were maintained. Arterial hypercapnia was generated by the addition of 60 torr dry CO2 (8 kPa) to the inspired air for 10 hrs, continuously. All subjects were paralyzed (vecuronium bromide) and ventilated with room air, while the ventilator settings were kept constant. MEASUREMENTS AND MAIN RESULTS: Arterial and venous gas exchange profiles, hemoglobin concentration, oxygen saturation, oxygen content, cardiac output, and oxygen consumption were determined, before, during, and after 10 hrs of hypercapnia, periodically. Both hemoglobin concentration and oxygen content were gradually increased during hypercapnia and reached significant levels at 8 and 10 hrs of hypercapnia, respectively. These increases continued up to 2 hrs after termination of hypercapnia. The PaO2/FIO2, as an index of arterial oxygenation, was significantly increased during the first 3 hrs of hypercapnia and then remained at the normoxic level up to 10 hrs of hypercapnia. No significant changes occurred in the mean arterial blood pressure and oxygen consumption. The heart rate and cardiac output were significantly reduced at 4 and 8 hrs of hypercapnia, respectively. The mean pulmonary artery pressure was increased throughout the hypercapnic trial. CONCLUSIONS: A relatively long-term exogenous hypercapnia can significantly increase oxygen-carrying capacity in normal ventilated dogs. Whether this effect can occur during permissive hypercapnia because of controlled ventilation in patients warrants investigation.

Experimental critical care in rats: gender differences in anesthesia, ventilation, and gas exchange.

OBJECTIVE: To compare normative ventilatory and gas-exchange data and anesthetic requirements in male and female rats subjected to critical care conditions. DESIGN: Prospective study. SETTING: Critical care research laboratory in a hospital. SUBJECTS: Twenty-two age-matched young male and female rats (Sprague-Dawley, Long Evans strain). INTERVENTIONS: Anesthesia was induced with 65 and 45 mg/kg pentobarbital in male and female rats, respectively. The rats were then tracheostomized and cannulated in one femoral vein and artery. Anesthesia was maintained using 8-15 mg/kg/hr pentobarbital (iv) and controlled by continuous hemodynamic monitoring. MEASUREMENTS AND MAIN RESULTS: Normoxic baselines for breathing frequency, tidal volume, minute volume, inspiratory-to-expiratory ratio, inspiratory drive (tidal volume/inspiratory time), respiratory system compliance, peak airway pressure, and gas-exchange profiles were established. Ventilatory and gas-exchange responses to oxygen and CO2 were then determined by exposure to 10 mins of hyperoxia (100% oxygen), two levels of mild and severe hypercapnic hyperoxia (inspired Pco2 of 30 and 60 torr; 4 and 8 kPa), and two levels of mild and severe normocapnic hypoxia (inspired PO2 of 81 and 48 torr; 10.7 and 6.3 kPa). The average anesthetic requirement (during a 5- to 6-hr experiment) was 30% less in the female rats than in the male rats (p < .05). Female rats showed significantly lower breathing frequency, minute volume (mL/min/kg), and inspiratory drive (mL/kg/sec) during hyperoxia, mild and severe hypercapnia, and mild hypoxia. Pulmonary peak airway pressure was significantly lower in the female rats, consistent with a significantly higher weight-indexed compliance during all exposures. The female rats also had significantly higher inspiratory-to-expiratory ratio and higher PaCO2 with lower pH during normoxia, hyperoxia, and mild hypercapnia. These gender differences had no effect on PaO2, which was similar in all exposures. CONCLUSIONS: There are significant gender differences in ventilation, gas exchange, and anesthetic requirements in rats subjected to critical care conditions. The gas-exchange values observed in these spontaneously breathing rats may represent the optimal levels attainable during pentobarbital anesthesia with normal lungs. They may serve as standards for ventilator settings in the rat models used for critical care studies.

Experimental critical care in ventilated rats: effect of hypercapnia on arterial oxygen-carrying capacity.

PURPOSE: We have previously demonstrated an increased arterial O2-carrying capacity in normal ventilated dogs subjected to both acute and prolonged exogenous hypercapnia. In the present study, we tested if arterial hypercapnia, during controlled ventilation, can increase O2-carrying capacity also in rats. MATERIALS AND METHODS: Twenty young male Sprague Dawley rats were anesthetized (60 mg/kg pentobarbital), tracheostomized, intubated, and one femoral vein and artery were cannulated. Anesthesia and paralysis were maintained using 15 mg/kg/h pentobarbital intravenously, and 2 mg/kg/h vecuronium bromide. The fluid balance (5 mL/kg/h saline), normothermia, and minute volume were maintained. The mean arterial blood pressure and heart rate were continuously monitored. Experiments included the following: (1) a control group, ventilated with normoxic air for 150 minutes (n = 5); (2) mild hypercapnia, a group of eight rats ventilated with normoxic air for 30 minutes and then ventilated with a mixture of normoxic air at 60 mm Hg CO2 (8 kPa) for 1 hour; and (3) severe hypercapnia, a group of seven rats were treated exactly as in group II, except a 90 mm Hg (12 kPa) CO2 during hypercapnia. Gas-exchange profile, arterial hemoglobin (Hb) concentration, arterial Hb-oxygen saturation (Hb-O2), and arterial O2 content were periodically determined during normocapnia and 1 hour of hypercapnia. RESULTS: Exposures to mild and severe hypercapnia, in rats with maintained ventilation, significantly reduced the arterial O2 content by 20% and 33%, respectively, without significant changes in the arterial Hb concentration (-2%). Severe hypercapnia generated a significant reduction of -14% in the PaO2, but not in PaO2/ FiO2 ratio. CONCLUSION: Rats subjected to controlled ventilation and permissive hypercapnia, unlike dogs and perhaps humans, show no augmentation of Hb concentration. Hypercapnia in rats also provokes much stronger Bohr effect than in dogs. Hypercapnia-induced Bohr effect in rats is accompanied with extreme desaturations of Hb-O2, and substantial reduction in the O2-carrying capacity. We speculate that the strong hypercapnia-induced Bohr effect in rats may prevent hypoxia at the tissue level. However, to maintain a stable oxygen-carrying capacity in rats used for pulmonary critical care studies with hypercapnia, we suggest to use hyperoxia, with or without a mild hypothermia.

Acute hypercapnia increases the oxygen-carrying capacity of the blood in ventilated dogs.

OBJECTIVE: To test the hypothesis that PaCO2 levels generated during permissive hypercapnia may enhance arterial oxygenation, when ventilation is maintained. DESIGN: Prospective study. SETTING: Research laboratory in a hospital. SUBJECTS: One group of eight mongrel dogs (four male; four female). INTERVENTIONS: The dogs were anesthetized (30 mg/kg iv pentobarbital), intubated, and cannulated in one femoral artery and vein. While paralyzed with 0.1 mg/kg/hr iv vecouronium bromide, all subjects were ventilated with room air. Anesthesia was maintained, using 2 to 3 mg/kg/hr iv pentobarbital. Arterial hypercapnia at the levels generated during permissive hypercapnia was produced by stepwise increases in the dry, inspired Pco2 (PiCO2) (0, 30, 45, 60 and 75 torr [0, 4, 6, 8, and 10 kPa]; 15 mins each). MEASUREMENTS AND MAIN RESULTS: Blood gas profiles were determined at each level of hypercapnia. The minute volume was maintained at the baseline level during all exposures. Arterial hypercapnia produced gradual and significant increases in the hemoglobin concentration. These increases were approximately 6%, 7%, 11%, and 14% at PiCO2 of 30, 45, 60, and 75 torr (4, 6, 8, and 10 kPa), respectively (p < .05; repeated analysis of variance followed by Dunnett multiple comparisons test). In parallel, the oxygen content increased by approximately 6%, 7%, 11%, and 13%, respectively. During hypercapnic trials, the PaO2 remained at the normal range, whereas the dry, inspired PO2 (PiO2) was reduced from 150 to 138 torr (20 to 18.4 kPa). The average PaO2 at the highest investigated level of arterial hypercapnia was at a normal range. The hemoglobin concentration and oxygen content returned to baseline values 30 mins after hypercapnic trials. The PaCO2 and pH became normalized 15 mins after hypercapnic trials. Indirect evidence for a similar response to hypercapnia in humans is presented. CONCLUSIONS: Permissive hypercapnia due to inhaled CO2 increases oxygen-carrying capacity in dogs. The PaO2 remains at normal range even at a PiCO2 of 75 torr (10 kPa). The benefits of these effects during permissive hypercapnia, due to controlled hypoventilation, warrants investigation.

Oxidative stress following traumatic brain injury in rats.

BACKGROUND: Free radicals may be involved in the pathophysiology of traumatic brain injury (TBI) through oxidative damage of neurovascular structures. Endogenous antioxidants, such as ascorbate and alpha-tocopherol, may play a critical role in combating these oxidative reactions and their oxidized products can serve as an important index of oxidative stress. METHODS: We used electron spin resonance (ESR) spectroscopy and in vivo spin trapping (reaction of an organic compound with free radical species) to detect the possible generation of free radicals after TBI. Injury was inflicted by a weight drop technique over the head (5.7 kg-cm). Rats were intravenously infused with either 1 mL, 0.1 M of the spin trap, alpha-phenyl-N-tert-butyl nitrone (PBN), or an equivalent volume of saline immediately before TBI or sham-injury. Animals were divided into four groups: (1) Group I: PBN-infused sham-injured, (2) Group II: PBN-infused injured, (3) Group III: saline-infused sham-injured, and (4) Group IV: saline-infused injured. Additional groups of saline-infused uninjured, saline-infused, and PBN-infused injured animals were used for histopathology. Sixty minutes after TBI or sham-injury, rats were again anesthetized and decapitated. The brains were removed within 1 minute, homogenized, and extracted for lipids. The extracts were analyzed by ESR spectroscopy. Brain ascorbic acid (AA) concentration was determined spectrophotometrically, using the ascorbate oxidase assay. RESULTS: No PBN spin adduct signals (indicating trapped free radical species) were visible 60 minutes after TBI. All groups of rats showed an ascorbyl free radical signal. The ascorbyl signal intensity (AI) was, however, significantly higher in the injured rats, while the brain (AA) was significantly reduced. In addition, the ratio of AI/AA, which eliminates the effect of variable ascorbate concentrations in the brain, was also significantly higher in the injured animals. CONCLUSIONS: We conclude that 60 minutes following TBI there was a significantly increased level of oxidative stress in the brain. This may reflect formation of free radical species with subsequent interaction with ascorbate (antioxidant) during the 60 minute period. The lack of PBN spin adduct signals 1 hour after TBI may indicate that free radical generation is time dependent and might be detectable earlier or later than the 60 minute period.

Modulation of sensitivity to hyperbaric oxygen by CO2 in newborn rats.

We examined the modifying effects of CO2 on the CNS and pulmonary manifestations of hyperbaric oxygenation (HBO) in newborn rats. Four- to seven-day-old rats were exposed to HBO with inspired PCO2 (PICO2) of approximately 0, 60, 90, 140, 190, and 380 mmHg at a total pressure of 5 atm abs. The PCO2 values studied corresponded with 0, 8, 12, 18, 25, and 50 kPa, respectively, at a total pressure of 507 kPa. The O2-CO2 exposures lasted for 2-8 h. Hypercapnia at PICO2 of 60 and 90 mmHg with HBO produced extensive pulmonary damage with a high post-decompression mortality, compared to HBO alone. In contrast, PICO2 at the anesthetic levels of 140 and 190 mmHg attenuated the visible pulmonary and neurologic manifestations of O2 toxicity, and significantly reduced post-decompression mortality, compared to moderate hypercapnia of 60-90 mmHg. Supercapnia, at PICO2 of 380 mmHg, with HBO also produced no visible neurologic effect, but it caused an early apnea with severe pulmonary damage. These data indicate unique and dose-dependent cerebral and pulmonary responses to hyperoxic-hypercapnia in newborn rats. It is speculated that anesthetic levels of hypercapnia may be utilized to improve tissue oxygenation during O2 therapy in newborns, without increasing the risk of pulmonary and CNS O2 poisoning.

Jejunoileal bypass-induced liver dysfunction and bacterial translocation: effect of intraluminal glutamine-infusion.

We tested the effect of long-term intraluminal administration of glutamine on jejunoileal bypass (JIB) induced abnormalities in the plasma-liver profile in rats. Male Sprague Dawley rats (200-250 g) were subjected to an end to side JIB followed by daily intraluminal infusions of either 8 ml saline (n = 5), infused over a 4-hour period, or 8 ml saline containing 1g/Kg body weight glutamine (n = 7) for 3 weeks. Thirteen unoperated rats and four JIB rats without infusions served as controls. At the conclusion of the experiment, a cardiac blood sample was removed and analyzed for plasma cholesterol, albumin, total protein, gamma glutaril transferase, lactic dehydrogenase, glutamic-oxaloacetic transaminase, glutamic pyruvic transaminase, alkaline phosphatase, and bilirubin. Tissue samples from various segments of bowel, liver, mesenteric lymph nodes, and spleen underwent histopathologic examination. Bacteriological cultures were prepared from jejunum, ileum, mesenteric lymph nodes, liver, and spleen. Bacterial translocation occurred in both JIB-saline and JIB-glutamine infused rats. Glutamine-infused rats developed a significant decrease in the plasma cholesterol levels. However, glutamine did not prevent the JIB-induced alterations in the plasma-liver profile and bowel histopathology. It is suggested that experimental JIB procedure can be used as a model of bacterial translocation consequent to mucosal permeability and intestinal inflammatory diseases.

Experimental retinopathy by hyperbaric oxygenation.

Retinopathy of prematurity (ROP) usually occurs after a prolonged exposure to normobaric hyperoxia in newborn mammals and infants. We hypothesized that experimental ROP also could develop after acute exposures to hyperbaric oxygenation (HBO), providing that a severe and maintained retinal vasoconstriction occurred during HBO exposure. Five- to seven-day-old, Long Evans Sprague-Dawley rats were exposed for 5 h either to 5 atm abs oxygen or to 5 atm abs O2 with 190 mmHg inspired PCO2 (hypercapnia). Control rats breathed air at atmospheric pressure. Two months after exposures, rats were anesthetized, perfused intraventricularly with India ink, and retinal images were obtained. Retinal vascular density (RVD) in each image was calculated as the number of pixels in the retinal vessel area divided by the total number of pixels in the image (retinal tissue and vessels). The RVD was significantly increased from 0.0112 +/- 0.004 in the air-exposed controls to 0.0417 +/- 0.029 in the HBO-exposed rats (mean +/- SD; n = 4 in each group). HBO with hypercapnia produced a nonsignificant increase in RVD (0.0255 +/- 0.007; n = 4), reducing the HBO-induced increase in RVD by 39%. These results are consistent with the hypothesis that a sustained HBO-induced retinal vasoconstriction in newborn rats, followed by a hypoxic-ischemic injury, might result in vascular proliferation, thereby initiating ROP development on return to air. Hypercapnia does not completely prevent HBO-induced retinal vasoproliferation, probably because possible vasodilation, induced by hypercapnia, can greatly elevate retinal tissue PO2 and promote oxidative damage.

Hyperbaric oxygen tolerance in newborn mammals--hypothesis on mechanisms and outcome.

Newborn mammals, compared to adults, are extremely resistant to the CNS effects of hyperbaric oxygenation (HBO) induced by excessive generation of reactive oxygen species. This tolerance to HBO may be related to either physiological responses or the chemical characteristics of the immature brain, including a low cerebral blood flow and energy metabolism, and a low concentration of polyunsaturated fatty acids. In adult mammals the main protective mechanism against CNS oxygen toxicity, besides endogenous antioxidants, is a transient HBO-induced cerebral vasoconstriction. How cerebral vasculature reacts to HBO in the immature brain is not known. We present indirect evidence suggesting that HBO in newborn rats induces a persistent cerebral vasoconstriction concurrently with a severe and maintained reduction in ventilation. It is speculated that the outcome of these physiologic responses to hyperoxic exposures may be: (a) extension of tolerance to both CNS and pulmonary oxygen poisoning; (b) creation of a profound hypoxic-ischemic condition in vulnerable neural structures; and (c) impairment of the circulatory and ventilatory responses to hypoxic stimuli on return to air with consequent development of a secondary hypoxic-ischemic condition. These hypothetical pre- and post-HBO events may set the stage for the development of some delayed neurological disorders, including the retinopathy of prematurity and the retardation of brain development in fetuses or prematurely-born infants subjected to oxygen therapy.

Hyperbaric oxygenation alters carotid body ultrastructure and function.

We previously demonstrated that chronic normobaric hyperoxia (NH) for 60-67 h attenuated the carotid chemosensory response to hypoxia, probably initiated by the generation of reactive oxygen species (ROS). Since biological systems are affected by oxygen in a dose-dependent manner, we hypothesized that hyperbaric oxygenation (HBO) would affect the cellular mechanisms of oxygen chemoreception in a shorter time. To test the hypothesis, we studied the effects of oxygen at 5 atmospheres absolute (ATA) on cats (n = 7) carotid body ultrastructure and chemosensory responses to hypoxia, hypercapnia, and to bolus injections of cyanide, nicotine and dopamine. Four control cats breathed room air at 1 ATA. At the termination of the experiments, carotid bodies from 4 cats in each group were fixed and prepared for electron microscopy and morphometry. On the average, HBO diminished the chemosensory responsiveness to hypoxia (P < 0.01, unpaired t-test) within about 2 h, supporting the hypothesis. The responses to hypercapnia or bolus injections of cyanide, nicotine and dopamine were normal. HBO did not diminish the distribution of the dense-cored vesicles but significantly increased the mean volume-density of mitochondria and decreased the cristated area per mitochondrion in the glomus cells. The latter suggests a link between oxidative metabolism and chemosensing, and the former excludes availability of neurotransmitters being the cause of the blunted chemosensory response to hypoxia.

Cerebral cardiovascular and respiratory variables after an experimental brain missile wound.

Brain missile wounding (BMW) affects brainstem and medullary cadiorespiratory functions leading to immediate systemic hypertension, bradycardia, and apnea. Secondary complications may also occur because of subsequent changes in systemic and intracranial physiological variables. To delineate the immediate and secondary effects of BMW, we monitored changes in several cerebral and cardiorespiratory parameters in pentobarbital-anesthetized spontaneously breathing cats before wounding and up to 90 min afterward. Total and regional cerebral blood flow (rCBF) and cardiac output (CO) were measured (microsphere technique) and arterial blood was sampled for pH, PO2 and PCO2 once before BMW and one to four times afterward. Mean arterial blood pressure (MABP), intracranial pressure (ICP), cerebral perfusion pressure (CPP = MABP - ICP), electrocardiogram (ECG), heart rate (HR), and electroencephalogram (EEG) were continuously recorded. Respiratory frequency (f), tidal volume (Vt), and ventilation (V) were recorded during each flow measurement and periodically throughout the experiment. Four unwounded cats served as controls and 15 cats were wounded at 1.4 J fronto-occipitally through an intact cranium. Unwounded cats showed no significant changes in any physiological variable measured during a 100 min experimentation period. Four wounded cats survived a 90 min post-BMW period and had only a transient brainstem effect including a 50% increase in MABP concurrently with 50% reductions in the f and HR. Nonsurvivors (11 of 15) lived from 1 to 41 min after wounding. These cats initially demonstrated similar changes in MABP, f, and HR to survivors, but these variables remained unstable, possibly indicating a persisting brainstem damage. Apnea accounted for death in 10 of 11 nonsurvivors. Although the primary brainstem effect might have existed in all nonsurvivors, it appears that only one cat died from reduced respiration alone. Others had one or several postwounding secondary complications: abruptly increased ICP producing a negative CPP, extreme reductions in CO or CBF and ventilation. Cardiac arrest occurred once. Thus, post-BMW mortality cannot be consistently ascribed to the impairment of a single physiological variable.

Free radical generation in the brain precedes hyperbaric oxygen-induced convulsions.

We tested the hypothesis that hyperbaric oxygenation (HBO) generates free radicals in the brain before the onset of neurological manifestations of central nervous system (CNS) oxygen poisoning. Chronically cannulated, conscious rats were individually placed in a transparent pressure chamber and exposed to (1) 5 atmospheres absolute (ATA) oxygen for 15 min (n = 4); (2) 5 ATA oxygen for 30 min (n = 5), during which no visible convulsions occurred; (3) 5 ATA oxygen for 30 min with recurrent convulsions (n = 6); (4) 5 ATA oxygen until the appearance of the first visible convulsions (n = 5); (5) 4 ATA oxygen for 60 min during which no convulsions occurred (n = 5); and (6) 5 ATA air for 30 min (n = 5, controls). Immediately before compression, 1 mL of 0.1 M of alpha-phenyl-N-tert-butyl nitrone (PBN) was administered intravenously (iv) for spin trapping. At the termination of each experiment, rats were euthanized by pentobarbital iv and decompressed within 1 min. Brains were rapidly removed for preparation of lipid extracts (Folch). The presence of PBN spin adducts in the lipid extracts was examined by electron spin resonance (ESR) spectroscopy. ESR spectra from unconvulsed rats exposed to 5 ATA oxygen for 30 min revealed both oxygen-centered and carbon-centered PBN spin adducts in three of the five brains. One of the five rats in this group showed an ascorbyl signal in the ESR spectrum.(ABSTRACT TRUNCATED AT 250 WORDS)