Central Nervous System Oxygen Toxicity Research Articles

Two faces of nitric oxide: implications for cellular mechanisms of oxygen toxicity

Recent investigations have elucidated some of the diverse roles played by reactive oxygen and nitrogen species in events that lead to oxygen toxicity and defend against it. The focus of this review is on toxic and protective mechanisms in hyperoxia that have been investigated in our laboratories, with an emphasis on interactions of nitric oxide (NO) with other endogenous chemical species and with different physiological systems. It is now emerging from these studies that the anatomical localization of NO release, which depends, in part, on whether the oxygen exposure is normobaric or hyperbaric, strongly influences whether toxicity emerges and what form it takes, for example, acute lung injury, central nervous system excitation, or both. Spatial effects also contribute to differences in the susceptibility of different cells in organs at risk from hyperoxia, especially in the brain and lungs. As additional nodes are identified in this interactive network of toxic and protective responses, future advances may open up the possibility of novel pharmacological interventions to extend both the time and partial pressures of oxygen exposures that can be safely tolerated. The implications of a better understanding of the mechanisms by which NO contributes to central nervous system oxygen toxicity may include new insights into the pathogenesis of seizures of diverse etiologies. Likewise, improved knowledge of NO-based mechanisms of pulmonary oxygen toxicity may enhance our understanding of other types of lung injury associated with oxidative or nitrosative stress.

Recovery from central nervous system oxygen toxicity in the rat at oxygen pressures between 100 and 300 kPa

No symptoms related to central nervous system (CNS) oxygen toxicity have been reported when diving with oxygen rebreathers at depths shallower than 3 msw. We hypothesised that recovery from CNS oxygen toxicity will take place when the PO(2) is less than 130 kPa. We exposed rats to a high PO(2) (mainly 608 kPa) to produce CNS oxygen toxicity. The latency to the first electrical discharge (FED) preceding convulsions was determined as the animal's control latency. Thereafter, the rat was exposed to the same PO(2) for 60% of its latency, then to a lower PO(2) for 15 min (sufficient time for full recovery in normoxia), and finally to the high PO(2) again until appearance of the FED. If recovery from CNS oxygen toxicity takes place during the interim period, the latency for the final exposure to the high oxygen pressure should not be shorter than the control. The latencies to CNS oxygen toxicity for exposure to the high oxygen pressure after a 15-min interim period at 21, 101, 132, 203, 304, 405, and 456 kPa were 110, 110, 125, 94, 85, 54 and 38% of the control value, respectively. Only after the last two interim pressures were the latencies significantly shorter than control values. The remaining latencies were not significantly different from 100%. Recovery from CNS oxygen toxicity in the rat takes place at a PO(2) anywhere between 21 and 304 kPa. The present findings support our previous suggestion that recovery from CNS oxygen toxicity in humans will take place at a PO(2) below 130 kPa. If our findings are corroborated by further human studies, this will justify including recovery in the algorithm for CNS oxygen toxicity in closed-circuit oxygen divers.

Contributions of nitric oxide synthase isoforms to pulmonary oxygen toxicity, local vs. mediated effects

Reactive species of oxygen and nitrogen have been collectively implicated in pulmonary oxygen toxicity, but the contributions of specific molecules are unknown. Therefore, we assessed the roles of several reactive species, particularly nitric oxide, in pulmonary injury by exposing wild-type mice and seven groups of genetically altered mice to >98% O2 at 1, 3, or 4 atmospheres absolute. Genetically altered animals included knockouts lacking either neuronal nitric oxide synthase (nNOS(-/-)), endothelial nitric oxide synthase (eNOS(-/-)), inducible nitric oxide synthase (iNOS(-/-)), extracellular superoxide dismutase (SOD3(-/-)), or glutathione peroxidase 1 (GPx1(-/-)), as well as two transgenic variants (S1179A and S1179D) having altered eNOS activities. We confirmed our earlier finding that normobaric hyperoxia (NBO2) and hyperbaric hyperoxia (HBO2) result in at least two distinct but overlapping patterns of pulmonary injury. Our new findings are that the role of nitric oxide in the pulmonary pathophysiology of hyperoxia depends both on the specific NOS isozyme that is its source and on the level of hyperoxia. Thus, iNOS predominates in the etiology of lung injury in NBO2, and SOD3 provides an important defense. But in HBO2, nNOS is a major contributor to pulmonary injury, whereas eNOS is protective. In addition, we demonstrated that nitric oxide derived from nNOS is involved in a neurogenic mechanism of HBO2-induced lung injury that is linked to central nervous system oxygen toxicity through adrenergic/cholinergic pathways.

A rat model to study decompression sickness after a trimix dive

depth limitations for diving with air as the breathing gas are well established. During the dive, increased ambient pressure generates an increase in both oxygen and nitrogen partial pressures. Central nervous system oxygen toxicity is well documented ([8][1]), and to prevent the risk of hyperoxic-

Response to CO2 in novice closed-circuit apparatus divers and after 1 year of active oxygen diving at shallow depths

Elevated arterial Pco(2) (hypercapnia) has a major effect on central nervous system oxygen toxicity in diving with a closed-circuit breathing apparatus. The purpose of the present study was to follow up the ability of divers to detect CO(2) and to determine the CO(2) retention trait after 1 year of active oxygen diving with closed-circuit apparatus. Ventilatory and perceptual responses to variations in inspired CO(2) (range: 0-5.6 kPa, 0-42 Torr) during moderate exercise were assessed in Israeli Navy combat divers on active duty. Tests were carried out on 40 divers during the novice oxygen diving phase (ND) and the experienced oxygen diving phase. No significant changes were found between the two phases for the minimal mean inspired Pco(2) that could be detected. The mean (with SD in parentheses) end-tidal Pco(2) during exposure to an inspired Pco(2) of 5.6 kPa (42 Torr) was significantly higher in the novice diving phase than in the experienced diving phase [8.1 kPa (SD 0.7), 62 Torr (SD 5) and 7.8 kPa (SD 0.6), 59 Torr (SD 4), respectively; P < or = 0.001]. One year of shallow oxygen diving activity with a closed-circuit apparatus does not affect the ability to detect CO(2) nor does it lead to increased CO(2) retention; rather, it may even bring about a decrease in this trait. This finding suggests that acquiring experience in oxygen diving with a closed-circuit apparatus at shallow depths does not place the diver at a greater risk of central nervous system oxygen toxicity due to CO(2) retention.

Effects of nitrogen and helium on CNS oxygen toxicity in the rat.

The contribution of inert gases to the risk of central nervous system (CNS) oxygen toxicity is a matter of controversy. Therefore, diving regulations apply strict rules regarding permissible oxygen pressures (Po(2)). We studied the effects of nitrogen and helium (0, 15, 25, 40, 50, and 60%) and different levels of Po(2) (507, 557, 608, and 658 kPa) on the latency to the first electrical discharge (FED) in the EEG in rats, with repeated measurements in each animal. Latency as a function of the nitrogen pressure was not homogeneous for each rat. The prolongation of latency observed in some rats at certain nitrogen pressures, mostly in the range 100 to 500 kPa, was superimposed on the general trend for a reduction in latency as nitrogen pressure increased. This pattern was an individual trait. In contrast with nitrogen, no prolongation of latency to CNS oxygen toxicity was observed with helium, where an increase in helium pressure caused a reduction in latency. This bimodal response and the variation in the response between rats, together with a possible effect of ambient temperature on metabolic rate, may explain the conflicting findings reported in the literature. The difference between the two inert gases may be related to the difference in the narcotic effect of nitrogen. Proof through further research of a correlation between individual sensitivity to nitrogen narcosis and protection by N(2) against CNS oxygen toxicity in rat may lead to a personal O(2) limit in mixed-gas diving based on the diver sensitivity to N(2) narcosis.

Heat acclimation prolongs the time to central nervous system oxygen toxicity in the rat: Possible involvement of HSP72

Oxygen toxicity of the central nervous system (CNS-OT) can occur during diving with oxygen-enriched gas mixtures, or during hyperbaric medical treatment. CNS-OT is characterised by convulsions and sudden loss of consciousness, which may be fatal in diving. Heat acclimation is known to provide cross-tolerance to various forms of stress in different organs, including the brain. We hypothesised that heat acclimation may delay the onset of CNS-OT in the rat. Male Sprague–Dawley rats were acclimated to an ambient temperature of 32 °C for 4 weeks. Rats in the control group were kept at 24 °C. Both groups were exposed to oxygen at 608 kPa. EEG was recorded continuously until the appearance of the first electrical discharge preceding clinical convulsions. CO 2 production was measured simultaneously with the EEG. Latency to CNS-OT was measured and brain samples were taken for evaluation of heat shock protein 72 (HSP72) levels by Western blot analysis at the end of the acclimation period and during 4 weeks of deacclimation. Latency to CNS-OT was twice as long in the heat-acclimated rat, with insignificant changes in CO 2 production. This prolongation continued for 2 weeks during deacclimation. There was a significant increase in the level of HSP72 following heat acclimation, with a subsequent decrease during deacclimation. We conclude that heat acclimation prolongs latency to CNS-OT in a way that does not involve changes in metabolic rate. During deacclimation there was a linear relationship between latency to CNS oxygen toxicity and the level of HSP72. A possible beneficial effect of HSP72 is discussed.

Modeling pulmonary and CNS O(2) toxicity and estimation of parameters for humans.

The power expression for cumulative oxygen toxicity and the exponential recovery were successfully applied to various features of oxygen toxicity. From the basic equation, we derived expressions for a protocol in which PO(2) changes with time. The parameters of the power equation were solved by using nonlinear regression for the reduction in vital capacity (DeltaVC) in humans: %DeltaVC = 0.0082 x t(2)(PO(2)/101.3)(4.57), where t is the time in hours and PO(2) is expressed in kPa. The recovery of lung volume is DeltaVC(t) = DeltaVC(e) x e(-(-0.42 + 0.00379PO(2))t), where DeltaVC(t) is the value at time t of the recovery, DeltaVC(e) is the value at the end of the hyperoxic exposure, and PO(2) is the prerecovery oxygen pressure. Data from different experiments on central nervous system (CNS) oxygen toxicity in humans in the hyperbaric chamber (n = 661) were analyzed along with data from actual closed-circuit oxygen diving (n = 2,039) by using a maximum likelihood method. The parameters of the model were solved for the combined data, yielding the power equation for active diving: K = t(2) (PO(2)/101.3)(6.8), where t is in minutes. It is suggested that the risk of CNS oxygen toxicity in diving can be derived from the calculated parameter of the normal distribution: Z = [ln(t) - 9.63 +3.38 x ln(PO(2)/101.3)]/2.02. The recovery time constant for CNS oxygen toxicity was calculated from the value obtained for the rat, taking into account the effect of body mass, and yielded the recovery equation: K(t) = K(e) x e(-0.079t), where K(t) and K(e) are the values of K at time t of the recovery process and at the end of the hyperbaric oxygen exposure, respectively, and t is in minutes.

PCO(2) threshold for CNS oxygen toxicity in rats in the low range of hyperbaric PO(2).

Central nervous system (CNS) oxygen toxicity, as manifested by the first electrical discharge (FED) in the electroencephalogram, can occur as convulsions and loss of consciousness. CO(2) potentiates this risk by vasodilation and pH reduction. We suggest that CO(2) can produce CNS oxygen toxicity at a PO(2) that does not on its own ultimately cause FED. We searched for the CO(2) threshold that will result in the appearance of FED at a PO(2) between 507 and 253 kPa. Rats were exposed to a PO(2) and an inspired PCO(2) in 1-kPa steps to define the threshold for FED. The results confirmed our assumption that each rat has its own PCO(2) threshold, any PCO(2) above which will cause FED but below which no FED will occur. As PO(2) decreased from 507 to 456, 405, and 355 kPa, the percentage of rats that exhibited FED without the addition of CO(2) (F(0)) dropped from 91 to 62, to 8 and 0%, respectively. The percentage of rats (F) having FED as a function of PCO(2) was sigmoid in shape and displaced toward high PCO(2) with the reduction in PO(2). The following formula is suggested to express risk as a function of PCO(2) and PO(2) [abstract: see text] where P(50) is the PCO(2)for the half response and N is power. A small increase in PCO(2) at a PO(2) that does not cause CNS oxygen toxicity may shift an entire population into the risk zone. Closed-circuit divers who are CO(2)retainers or divers who have elevated inspired CO(2)are at increased risk of CNS oxygen toxicity.

Nitric oxide production is enhanced in rat brain before oxygen-induced convulsions

Central nervous system oxygen toxicity (CNS O 2 toxicity) is preceded by release of hyperoxic vasoconstriction, which increases regional cerebral blood flow (rCBF). These increases in rCBF precede the onset of O 2-induced convulsions. We have tested the hypothesis that hyperbaric oxygen (HBO 2) stimulates NO production in the brain that leads to hyperemia and anticipates electrical signs of neurotoxicity. We measured rCBF and EEG responses in rats exposed at 4 to 6 atmospheres (ATA) of HBO 2 and correlated them with brain interstitial NO metabolites (NO x ) as an index of NO production. During exposures to hyperbaric oxygen rCBF decreased at 4 ATA, decreased for the initial 30 min at 5 ATA then gradually increased, and increased within 30 min at 6 ATA. Changes in rCBF correlated positively with NO x production; increases in rCBF during HBO 2 exposure were associated with large increases in NO x at 5 and 6 ATA and always preceded EEG discharges as a sign of CNS O 2 toxicity. In rats pretreated with l-NAME, rCBF remained maximally decreased throughout 75 min of HBO 2 at 4, 5 and 6 ATA. These data provide the first direct evidence that increased NO production during prolonged HBO 2 exposure is responsible for escape from hyperoxic vasoconstriction. The finding suggests that NO overproduction initiates CNS O 2 toxicity by increasing rCBF, which allows excessive O 2 to be delivered to the brain.

Humidity does not affect central nervous system oxygen toxicity.

Central nervous system (CNS) oxygen toxicity can occur as convulsions and loss of consciousness when hyperbaric oxygen is breathed in diving and hyperbaric medical therapy. Lin and Jamieson (J Appl Physiol 75: 1980-1983, 1993) reported that humidity in the inspired gas enhances CNS oxygen toxicity. Because alveolar gas is fully saturated with water vapor, we could not see a cause and effect and surmised that other factors, such as metabolic rate, might be involved. Rats were exposed to 507- and 608-kPa O(2) in dry (31 or 14%) or humid (99%) atmosphere until the appearance of the first electrical discharge preceding the clinical convulsions. Each rat served as its own control. A thermoneutral temperature (28 +/- 0.4 degrees C) yielded resting CO(2) production of 0.81 +/- 0.06 ml x g(-1) x h(-1). Latency to the first electrical discharge was not affected by humidity. At 507-kPa O(2), latency was 23 +/- 0.4 and 22 +/- 0.7 min in dry and humid conditions, respectively, and, at 608-kPa O(2), latency was 15 +/- 4 and 14 +/- 3 min in dry and humid conditions, respectively. When no effects of CO(2) and metabolic rate are present, humidity does not affect CNS oxygen toxicity. Relevance of the findings to diving and hyperbaric therapy is discussed.

Prolonged exposure to hyperbaric oxygen induces neuronal damage in primary rat cortical cultures

While seizure attack is one of the serious complications during the hyperbaric oxygen (HBO) therapy, there is still no direct evidence showing that HBO can induce neuronal damage in the brain. The objective of this study was first to investigate whether HBO would lead to neurotoxicity in the primary rat cortical culture. Second, since alterations in neurotransmitters have been suggested in the pathophysiology of central nervous system (CNS) oxygen toxicity, the protective effects of the N-methyl- d-aspartate (NMDA) receptor antagonism and nitric oxide (NO) synthase inhibition on the HBO-induced neuronal damage were examined. The results showed that HBO exposure to 6 atmosphere absolute pressure (ATA) for 30, 60, and 90 min increased the lactate dehydrogenase (LDH) activity in the culture medium in a time-dependent manner. Accordingly, the cell survival, measured by the 3,(4,5-dimethyl-2-thiazolyl)2,5-diphenyl-tetrazolium bromide (MTT) assay, was decreased after HBO exposure. Pretreatment with the NMDA antagonist MK-801 protected the cells against the HBO-induced damage. The protective effect was also noted in the cells pretreated with l- N G-nitro-arginine methyl ester, an NO synthase inhibitor. Thus, our results suggest that activation of NMDA receptors and production of NO play a role in the neurotoxicity produced by hyperbaric oxygen exposure.

Latency to CNS oxygen toxicity in rats as a function of PCO(2) and PO(2).

Central nervous system (CNS) oxygen toxicity can occur as convulsions and loss of consciousness, without any premonitory symptoms. We have made a quantitative study of the effect of inspired carbon dioxide on sensitivity to oxygen toxicity in the rat. Rats were exposed to four oxygen pressures (PO(2); 456, 507, 608 and 709 kPa) and an inspired partial pressure of carbon dioxide (PCO(2)) in the range 0-12 kPa until the appearance of the electroencephalograph first electrical discharge (FED) that precedes the clinical convulsions. Exposures were conducted at a thermoneutral temperature of 27 degrees C. Latency to the FED decreased linearly with the increase in PCO(2) at all four PO(2) values studied. This decrease, which is probably related to the cerebral vasodilatory effect of carbon dioxide, reached a minimal value that remained constant on further elevation of PCO(2). The slopes (absolute value) and intercepts of latency to the FED as a function of carbon dioxide decreased with the increase in PO(2). This log-linear relationship made possible the derivation of equations that describe latency to the FED as a function of both PO(2) and PCO(2) in the PCO(2) - dependent range: Latency (min) = e((5.19-0.0040)(P)(O(2)))-e((2.77-0.0034)(P)(O(2))) x PCO(2) (kPa), and in the PCO(2)-independent range: Latency(min) = e((2.44-0. 0009)(P)(O(2))). A PCO(2) as low as 1 kPa significantly reduced the latency to the FED. It is suggested that in closed-circuit oxygen diving, any accumulation of carbon dioxide should be avoided in order to minimize the risk of CNS oxygen toxicity.

Latency of oxygen toxicity of the central nervous system in rats as a function of carbon dioxide production and partial pressure of oxygen.

Oxygen toxicity of the central nervous system (CNS) can occur as convulsions and loss of consciousness, with no warning symptoms. A quantitative study of the effect of metabolic rate on sensitivity to oxygen toxicity was made in the rat. A group of 19 rats were exposed (126 exposures) to 12 combinations of four pressures (456, 507, 608 and 709 kPa) and three ambient temperatures (15, 23 and 29 degrees C) until the appearance of the first electrical discharge (FED) preceding clinical convulsions. Carbon dioxide production (VCO2) was also measured. A thermoneutral zone (mean VCO2 0.87 ml x g(-1) x h(-1)) existed between the temperatures of 24 and 29 degrees C; at temperatures lower than this, the metabolic rate increased by 1.2 to 4 times the resting level. Latency of FED decreased linearly with the increase in VCO2 at all four oxygen pressures. The slopes (absolute value) and intercepts decreased with the increase in oxygen pressure. This linear relationship made possible the derivation of an equation which described latency of the FED as a function of both oxygen pressure and metabolic rate. Various environmental and other physiological factors that have been said to influence sensitivity to CNS oxygen toxicity, enhancing the effect of the partial pressure of oxygen, can be explained by their effect on metabolic rate. It is suggested that in situations where there is a risk of oxygen toxicity of the CNS, that risk would be reduced by a lower metabolic rate.

Recovery time constant in central nervous system O2 toxicity in the rat.

The development of oxygen toxicity can be delayed by intermittent periods of normoxia. However, there is no accepted procedure for quantifing the recovery during normoxia. A cumulative oxygen toxicity index - K, when K reaches a critical value (Kc) and the toxic effect is manifested, can be calculated using the equation K = t(2)e x PO(2)c where t(e) is hyperoxic exposure time and PO2 is oxygen pressure and c is a power parameter. Recovery during normoxia (reducing K) is calculated by the equation K2 = K1 x e(-rt(r)) where t(r) is recovery time, r being the recovery time constant. A combination of accumulation of oxygen toxicity and its recovery can be used to calculate central nervous system oxygen toxicity. In protocol A (n = 25), r was calculated for rats exposed either continuously to 608 kPa oxygen or to PO2 = 608 kPa followed by a period of normoxia (3.5% O2), with a subsequent return to PO2 = 608 kPa until appearance of the first electrical discharge (FED) in the electroencephalogram which precedes clinical convulsions. In protocol B (n = 22), predicted latency to the FED was compared to measured latency for seven different exposures to hyperbaric oxygen (HBO), followed by a period of normoxia and further HBO exposure. Recovery followed an exponential path, with r = 0.31 (SD 0.12) min(-1). The predicted latency to FED in protocol B correlated with the measured latencies. Calculation of the recovery of the CNS oxygen toxicity agreed with the previously suggested exponential recovery of the hypoxic ventilatory response and was probably a general recovery process. We concluded that recovery can be applied to the design of various hyperoxic exposures.

Susceptibility of the injured rat brain to CNS oxygen toxicity.

The possibility of an altered susceptibility of the injured brain to central nervous system (CNS) oxygen toxicity was examined in awake rats. Moderate to severe closed head injury with diffuse axonal damage was produced in anesthetized rats by the fluid percussion method (2-2.5 atm), after which chronic EEG electrodes were implanted. Twenty-four hours later, the rats were exposed to 5 atm abs (506.5 kPa) oxygen and the time to appearance of paroxysmal EEG patterns was noted. The difference between the 19 minute median latency of this group and 16 minute of a control group which underwent a sham operation did not reach statistical significance. Some injured animals convulsed with minimal or no EEG changes. The clinical implication could be that brain injured patients are not at higher risk of CNS oxygen toxicity but the EEG alterations that could potentially be used to forecast incipient convulsions, or be the indication of actual convulsions in the intact brain of a paralyzed patient, may not always be present.

A Model for Predicting Central Nervous System Oxygen Toxicity from Hyperbaric Oxygen Exposures in Humans

Under certain circumstances, Navy divers breathe 100% O2 when working underwater. Serious symptoms of central nervous system (CNS) O2 toxicity can develop from hyperbaric O2 exposure; immersion and exercise are also known to exacerbate toxicity. We developed risk models for quantitative prediction of the probability of developing symptoms using a large set of human data in which occupational exposure conditions were simulated. Exposures were 5 to 265 min at PO2 levels from 20 to 50 feet of sea water (fsw) (1 fsw = 3.06 kPa). Approximately half of the exposures were to a single PO2, while the remainder were more complicated consisting of exposures to multiple levels of hyperbaric O2. In 688 trials, there were 42 exposure-stopping symptoms. We used maximum likelihood to estimate parameters, likelihood ratios to compare model fits, and chi 2 tests to judge goodness-of-fit of model predictions to observations. The modeling shows that risk has a steep PO2 dependence. A model with autocatalytic features fits the data as well as a simpler model: when PO2 is elevated beyond 34 fsw, risk accumulates rapidly without bound while accumulating toward an asymptote at lower PO2 levels. This autocatalytic feature of risk accumulation implies a testable hypothesis that substantial protection from human CNS O2 toxicity can be obtained from intermittent exposure (periodic exposure to lower PO2). The models predict that the probability of O2 toxicity is less than 7% with current Navy limits while breathing 95% O2. Probability of symptoms is < 1% if FIO2 is maintained at the United States Navy recommended level of 75%.

Beta-carotene and CNS oxygen toxicity in rats.

Beta-carotenes are reported to be potent free radical quenchers, singlet oxygen scavengers, and lipid antioxidants. Oxygen free radicals that are produced in excess during exposure to oxygen at high pressures and overwhelm the body's normal antioxidant defense systems seem to mediate the hyperoxic insult. We decided to test the possible protective effect against central nervous system oxygen toxicity of a natural beta-carotene composed of equal amounts of the all-trans and 9-cis isomers obtained from the unicellular halotolerant alga Dunaliella bardawil. Rats implanted with chronic cortical electrodes for continuous electroencephalogram monitoring were fed on ground commercial food enriched with natural beta-carotene (1 g/kg diet). On completion of 1 wk of the diet, the rats were exposed to 0.5 MPa oxygen and then their livers were removed for beta-carotene and vitamin A analysis. A significant increase was noted in the latent period preceding oxygen seizures in the group of rats in which the diet was supplemented by natural beta-carotene compared with rats given a normal diet (38.5 +/- 3.4 vs. 16.8 +/- 1.8 min; P < 0.05). Further experiments are required to evaluate the potential benefit of supplementing the diet of divers and patients exposed to high pressures of oxygen with the beta-carotene-rich D. bardawil.

The effect of flunarizine on central nervous system oxygen toxicity in rats

The toxicity of hyperbaric oxygen in the central nervous system is expressed by generalized tonic-clonic seizures. In the search for drugs effective against these seizures, we tested flunarizine, a calcium antagonist known to have antiepileptic properties and only minimal cardiovascular side effects. 49 rats with chronic cortical electrodes were injected i.p. with six different doses of flunarizine (10–300 mg/kg) or vehicle, before exposure to 0.5 MPa oxygen two doses of flunarizine and vehicle were given to rats exposed to oxygen with 5% CO 2 at an absolute pressure of 0.5 MPa. EEG and spectral analysis of background EEG activity were monitored. The duration of the latent period before the appearance of electrical discharges in the EEG was used as an index of oxygen toxicity. There was no statistical difference between the duration of the latent periods for the seven groups treated by flunarizine or by vehicle on exposure to 0.5 MPa pure oxygen (P=0.9 in ANOVA), but on exposure to oxygen with CO 2, there was significant prolongation of the latent periods in comparison with vehicle (P<0.001). Our results suggest that on exposure to hyperbaric oxygen, the antiepileptic effect of flunarizine might be masked, probably by its cerebral antivasoconstrictive effect.

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 occured ( 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 5 ATA oxygen for 30 min revealed both oxygen-centered and carbon-centered PBN spin adducts in three of the brains. One of the five rats in this group showed an ascorbyl signal in the ESR spectrum. In contrast, only one of the six rats with recurrent convulsions and one of five which were sacrified immediately after the first visible convulsions showed a low-intensity ESR signal due to PBN spin adducts. Unconvulsed rats exposed to 5 ATA oxygen for 15 min or 4 ATA oxygen for 60 min showed no PBN spin adducts but had an ascorbyl signal in two of four and three of five ESR spectra, respectively. Lipid extracts from control rats exposed to 5 ATA air showed no ascorbyl ESR signal. The preconvulsive appearance of PBN spin adducts and ascorbyl radical ESR signals is consistent with the free radical theory of the CNS oxygen toxicity. However, the reason for diminution or disappearance of PBN spin adducts after HBO-induced convulsions remains to be investigated.