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Abstract
In vitro studies have well established the neuroprotective action of the noble gas argon. However, only limited data from in vivo models are available, and particularly whether postexcitotoxic or postischemic argon can provide neuroprotection in vivo still remains to be demonstrated. Here, we investigated the possible neuroprotective effect of postexcitotoxic-postischemic argon both ex vivo in acute brain slices subjected to ischemia in the form of oxygen and glucose deprivation (OGD), and in vivo in rats subjected to an intrastriatal injection of N-methyl-D-aspartate (NMDA) or to the occlusion of middle-cerebral artery (MCAO). We show that postexcitotoxic-postischemic argon reduces OGD-induced cell injury in brain slices, and further reduces NMDA-induced brain damage and MCAO-induced cortical brain damage in rats. Contrasting with its beneficial effect at the cortical level, we show that postischemic argon increases MCAO-induced subcortical brain damage and provides no improvement of neurologic outcome as compared to control animals. These results extend previous data on the neuroprotective action of argon. Particularly, taken together with previous in vivo data that have shown that intraischemic argon has neuroprotective action at both the cortical and subcortical level, our findings on postischemic argon suggest that this noble gas could be administered during but not after ischemia, i.e. before but not after reperfusion has occurred, in order to provide cortical neuroprotection and to avoid increasing subcortical brain damage. Also, the effects of argon are discussed as regards to the oxygen-like chemical, pharmacological, and physical properties of argon.
Citation: David HN, Haelewyn B, Degoulet M, Colomb DG Jr, Risso J-J, Abraini JH (2012) Ex Vivo and In Vivo Neuroprotection Induced by Argon When Given after an Excitotoxic or Ischemic Insult. PLoS ONE 7(2): e30934. https://doi.org/10.1371/journal.pone.0030934
Editor: Ken Arai, Massachusetts General Hospital/Harvard Medical School, United States of America
Received: November 11, 2011; Accepted: December 24, 2011; Published: February 22, 2012
This is an open-access article, free of all copyright, and may be freely reproduced, distributed, transmitted, modified, built upon, or otherwise used by anyone for any lawful purpose. The work is made available under the Creative Commons CC0 public domain dedication.
Funding: This work was supported by the Direction Générale de l'Armement of the French Ministry of Defence and NNOXe Pharmaceuticals (Quebec City, QC, Canada). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. No additional external funding was received for this study.
Competing interests: The authors have read the journal's policy and have the following conflicts: This study was partly funded by NNOXe Pharmaceuticals. There are no patents, products in development or marketed products to declare. Lt. Colomb is a military service member and this work was prepared as part of his official duties. The views expressed in this article are those of the author and do not reflect the official policy or position of the Department of the Navy, Department of Defense, nor the U.S. Government. This does not alter the authors' adherence to all the PLoS ONE policies on sharing data and materials.
Introduction
Acute brain ischemia is primarily caused by a disruption of cerebral blood flow through thromboembolism that leads to an oxygen and glucose deprivation, excessive glutamate release and subsequent postsynaptic overstimulation of glutamate receptors, a process known to be critical in ischemia-induced neuronal death [1]–[3]. Over the past 10 years, a series of in vitro and in vivo studies in models of hypoxic/ischemic insults has demonstrated the neuroprotective potential of some inert gases, among which xenon has been identified as the most promising agent [4]–[13]. However, the major obstacle to the widespread clinical use of xenon is its scarceness and excessive cost of production. Interestingly argon, which is a cost-efficient and easily available gas with no narcotic and anesthetic action at normal atmospheric pressure, has been also shown to provide organoprotection and neuroprotection against hypoxic-ischemic insults [14]–[18] (but see reference [19] for discrepant results). Recent in vivo studies have further shown that intraischemic argon at 50 vol% (with the remainder being 50 vol% oxygen) provides both cortical and subcortical neuroprotection in rats subjected to transient middle cerebral artery occlusion (MCAO) [18]. However, despite this latter study, our knowledge on the ability of argon to provide neuroprotection in acute ischemic stroke still remains limited and should be augmented to evaluate the actual neuroprotective potential of this gas. Particularly, whether argon could provide neuroprotection when given after ischemia still remained unknown. This latter point is not trivial since previous data have shown the critical importance of the time at which inert gases are administered, during or after ischemia [7], [20], to obtain neuroprotection.
Therefore, in the present report, in order to assess thoroughly the neuroprotective potential of argon and to determine whether argon could be a cost-efficient alternative to xenon, we studied the neuroprotective effect of postischemic argon in vitro in brain slices exposed to ischemia in the form of oxygen and glucose deprivation (OGD) and in vivo in rats subjected to transient MCAO-induced ischemia or NMDA-induced excitotoxic insult.
Results
Effect of argon on OGD-induced cell injury in vitro
First, we investigated the effect of postischemic argon on neuronal injury induced by OGD in acute brain slices. Cell injury was assessed by the release of OGD-induced lactate dehydrogenase (LDH). Exposure to OGD led to a sustained increase of LDH release compared to sham slices (P<0.0001; Fig. 1A,B). Argon given after OGD, i.e. during the postischemic “reperfusion” period, led to significant changes in OGD-induced LDH release (P<0.0001), so that LDH levels with argon of 37.5 vol% to 75 vol% were lower than those recorded in OGD control slices exposed to nitrogen (P<0.01−0.0001; Fig. 1A). Maximal neuroprotection occurred with argon at 50 vol%, and increased with time (Fig. 1B).
(A) Concentration-response effects of argon (Ar) on OGD-induced LDH release (n = 23–27 per group). Exposure to OGD led to an increase in LDH release. Argon of 37.5 vol% to 75 vol% significantly reduced the increase in LDH release induced by OGD. (B) Time-course effects of OGD-induced cell injury and argon-induced neuroprotection. Cell injury induced by OGD as well as neuroprotection by argon increased as a function of time during the 3-h “reperfusion” period. Data are expressed as mean ± SEM. * P<0.05 vs OGD control slices; + P<0.05 vs sham slices.
Effect of argon on NMDA-induced neuronal death in vivo
Next, because postsynaptic overactivation of NMDA receptors plays a critical role in ischemic brain damage, we investigated the concentration-response effects of argon on NMDA-induced neuronal death when given 1 h after injection. Intrastriatal injection of NMDA led to significant brain damage compared to sham rats injected with saline (P<0.001). Argon of 15 vol% to 75 vol% reduced NMDA-induced brain damage (P<0.05), so that rats treated with argon at 37.5 vol% or 50 vol% had reduced brain damage compared to control animals treated with medical air (P<0.02–0.05; Fig. 2).
Injection of NMDA led to significant neuronal death. Sham rats had no brain damage data not shown). Rats treated with argon at 37.5 vol% and 50 vol% had a significant reduction of neuronal death as compared to NMDA control animals treated with medical air. Data are expressed as the median value and 25–75th percentiles * P<0.05 vs NMDA control rats treated with medical air.
Effects of argon on MCAO-induced brain damage and behavioral motor alterations
Next, based on the ex vivo and in vivo findings above that indicate that maximal neuroprotection by postinsult argon occurs at 50 vol%, we studied the effects of postischemic argon at 50 vol% on MCAO-induced brain damage and neurologic deficits. When given 1 h after reperfusion, i.e. 2 h after MCAO induction, argon reduced cortical volumes of brain damage by ∼35% (P<0.02; Fig. 3A) but increased subcortical brain damage by ∼35% (P<0.02; Fig. 3A) compared to control rats treated with air. At the behavioral motor level, MCAO rats treated with argon still exhibited a decreased score of neurologic outcome on day 1 (P<0.0001) and day 2 (P<0.0001) postischemia as compared to sham rats (Fig. 3B) due to marked deficits in motor coordination and rearing activity similar to those of control rats treated with medical air (Fig. 3C; P<0.0001). Also, argon did increase the rats' body temperature by 1.1°C (P<0.0001), but had no effect on other physiological parameters (Table 1).
(A) Control rats subjected to MCAO (treated with medical air) exhibited significant cortical and subcortical brain damage. Sham rats had no brain damage (data not shown). Argon at 50 vol% produces cortical neuroprotection, but did increase subcortical brain damage. (B) MCAO-induced global score of neurologic outcome. Control rats (line with black boxes), but not sham rats (line with white boxes), exhibited neurological deficits. Rats treated with argon at 50 vol% (line with gray boxes) showed no improvement of neurologic deficits as compared to control rats treated with medical air. (C) Scores of motor coordination and rearing activity in sham rats, control animals treated with medical air, and rats treated with argon at 50 vol%. Data are expressed as mean ± SEM. * P<0.05 vs MCAO control rats treated with medical air; + P<0.05 vs sham rats.
Discussion
The present study shows that argon given after an excitotoxic or ischemic insult reduces cell injury induced by OGD ex vivo, subcortical (mainly striatal) neuronal death induced by NMDA injection, and cortical brain damage induced by MCAO. In contrast with these beneficial effects, postinsult argon did increase subcortical brain damage and failed to reduce neurologic deficits induced by MCAO.
Argon possesses oxygen-like properties that could explain at least partly its neuroprotective action. As reported by Semenov, the Nobel Prize Laureate of chemistry in 1956 with Sir Cyril Norman Hinshelwood for their work about branched chain reactions, argon that is incapable of chemical reactions behaves however as a sort of catalyst for some of them by producing a kind of oxygen synergy [21]. In that way, physiological studies have clearly demonstrated that argon at 50 vol% allowed rats exposed to hypoxic conditions incompatible with life to adapt and maintain their oxygen demand [22] Also, argon of 25 to 80 vol% increased survival in rats exposed to normocapnic or hypercapnic hypoxia, and conversely potentiated oxygen-induced convulsions [22]. In addition, in rats exposed to non-lethal hypoxia (7 vol% oxygen for 1 hour), argon restored by ∼35% the mitochondrial respiratory enzyme activity [22], whose impairment following excitotoxic insults triggers neuronal death [23], [24]. Such a kind of argon-oxygen synergism could explain the neuroprotective effects of argon against hypoxia [14] and ischemia as reported in the present and previous studies [15], [17], [18]. Also, since previous data have shown redox modulation of the NMDA receptor with reduction inducing a potentiation and oxidation an inhibition of the NMDA receptor activity and glutamate-induced neuronal death [25], [26], the synergistic effect of argon on oxygen could explain its ability at reducing NMDA-induced neuronal death (a normoxic excitotoxic model). Indeed, in line with these data and the ability of argon at reducing NMDA-induced neuronal death, intrainsult oxygen has been shown to have antiexcitotoxic effects and to reduce NMDA-induced neuronal death and intraneuronal calcium influx [27], a critical event known to play a major role in excitotoxic and ischemic brain damage [1]–[3].
In contrast with its beneficial effect at the cortical level, we found that postischemic argon did increase MCAO-induced subcortical brain damage and further failed to reduce MCAO-induced neurologic deficits. This latter effect is in line with previous data that have demonstrated that neuroprotection requires preserving 80–90% of both the cortex and the subcortical areas to provide significant neurologic recovery and further predicted that 30% of subcortical brain damage might be sufficient to suppress any voluntary motor behavior [28]. Physiologically, the striatum that is the main part of the subcortical brain areas that suffer MCAO-induced brain damage is well recognized to be difficult to protect due to its lack of collateral vasculature and dramatic reduction in cerebral blood flow and oxygen supply as compared to the cortex. In these conditions, the occurrence of argon-induced oxygen synergy would not be possible as it occurs in brain slices exposed to OGD, where oxygen diffusion exists at the air-saline solution interface, in the cortex of rats subjected to MCAO-induced ischemia, where residual oxygen exists due to collateral vasculature, and in the striatum of rats subjected to NMDA injection that is a normoxic model of excitotoxic insult. Also, in contrast with the hypothermia-mediated neuroprotective effects of helium shown to result from the specific heat of helium that is higher than that of air [29], we found that argon as it could be expected from its specific heat that is half that of air produces mild hyperthermia, a condition well known to worsen ischemic brain damage that could have affected particularly the yet dramatically suffering subcortical brain areas as compared to the cortex. This latter possibility is in line with previous investigations that have shown that, in contrast with the cortex, neuroprotection of the striatum is tenuous and can be no longer obtained when time-to-treatment with xenon increases from 1 to 2 h after induction of ischemia or when blood/gas solubility (that is inversely proportional to diffusion) increases from 0.12 to 0.46 as a function of whether xenon or nitrous oxide is used to provide neuroprotection [30]. Finally, if one consider that argon would act by modulating membrane proteins [31] and particularly by potentiating the γ-amino-butyric acid type A (GABAA) receptor-mediated inhibitory neurotransmission [32], the present finding that postischemic argon provides no improvement of neurologic outcome while reducing cortical but not subcortical brain damage is in good agreement with a recent clinical study that has reported – in contrast with histological studies in animal models [33], [34] – that administration after reperfusion of GABAA agonists, but not of anti-cholinergic drugs, disrupts the neuronal plasticity that supports functional recovery in stroke patients and thereby reinduces clinical deficits after stroke [35]. In line with this latter study, it has to be noted that xenon that provides dramatic postischemic neuroprotection possesses efficient antagonistic properties at the cholinergic neurotransmission [36].
It is noteworthy that the lack of neuroprotective effect of argon on ischemia-induced subcortical brain damage reported herein opposes recent data that have demonstrated argon-induced subcortical (and cortical) neuroprotection [18]. Rather than an actual discrepancy (different results obtained from similar experimental conditions), it is likely with little doubt that this can be attributed to major differences in the protocol used. Particularly, administration of argon during ischemia [18] or after ischemia (in the present report) could have played a major role, since previous studies have shown the critical importance of the time inert gases are given, during or after ischemia, to obtain neuroprotection [7], [20]. Also, in contrast with the work by Ryang and colleagues [18] where intraischemic argon was given for 1 h through a face mask in anesthetized rats, in the present study postischemic argon was given for 3 h in freely-moving animals not controlled for temperature in an anesthesia box, an environmental condition shown to allow the effect of inert gases on body temperature to occur much more rapidly than using face mask [37]. Finally, whether neuroprotection by intraischemic argon together with 50 vol% oxygen (vs 25 vol% oxygen+25 vol% nitrogen in the present study) can be fully attributed to argon remains actually and highly questionable since intraischemic oxygen has been shown to reduce ischemic brain damage both through anti-excitotoxic and thrombolytic processes (David et al., unpublished data).
The present study has possible weaknesses that must be discussed. First, although there is no real alternative to use an anesthetic such as isoflurane to induce transient brain ischemia, the impact of isoflurane administration in combination with 100 vol% oxygen on the findings of this study must be examined. Indeed, since isoflurane is known as a neuroprotectant preconditioning agent in the adult but not in the newborn rat [38]–[43], it is possible that residual isoflurane could have contributed at least partly to provide neuroprotection against MCAO-induced cortical brain damage. Conversely, it is possible that residual isoflurane could have contributed to increase MCAO-induced subcortical brain damage by producing apoptosis, an adverse effect of isoflurane yet demonstrated when given in combination with nitrous oxide but not alone [38], [41]. Likewise, since oxygen has been shown either to reduce or to increase excitotoxic and/or ischemic brain damage [27], [44], [45], it is possible that the use of 100 vol% oxygen to induce isoflurane anesthesia could have contributed also to reduce cortical brain damage and/or to increase subcortical brain damage. However, these possibilities are unlikely to be true since in the present study (i) the total duration of surgery was no more than 25 minutes, thereby allowing rapid isoflurane and oxygen desaturation from the rat's body and brain and thereby the virtual absence of residual isoflurane and oxygen during the period of treatment with argon [46]; (ii) isoflurane was never given in combination with argon. Support for this are previous data with the same protocol that have shown that xenon, in contrast with what found in the present study with argon, provides neuroprotection both at the cortical and subcortical levels [6]. Second, in this study, cerebral blood flow was not measured throughout MCAO since we wanted to shorten anesthesia to limit isoflurane effects. However, this should not have biased our results since postischemic argon obviously induces neuroprotective and adverse effects at the cortical and subcortical level respectively. Indeed, it is unlikely that such a dichotomic response could be attributed to a technical problem of occlusion/reperfusion, which if such should have biased data in the same way at the cortical and subcortical level.
In conclusion, our results do not support that postischemic argon could be an efficient alternative to xenon, shown to provide dramatic postischemic neuroprotection. However, given its antiexcitotoxic effect, it is possible that argon could be useful for treating other brain insults, such as brain trauma.
Materials and Methods
Animals
All animal-use procedures were examined by a local ethic committee (Cyceron, Caen, France) in accordance with the European Communities Council Directive for the use of animals in biomedical experimentation (Declaration of Helsinki), and approved with permit number 14–27. Adult male Sprague-Dawley rats (Janvier, Le Genest Saint-Isle, France) weighing 250 to 280 g were housed with food and water ad libitum by group of six (before surgery) or individually (after surgery) at a room temperature of 21±0.5°C with lights on from 8:00 pm to 8:00 am.
Ex vivo OGD studies
OGD studies were performed as detailed previously [6]. Coronal brain slices of 400 µM thickness including the striatum were cut and allowed to recover at room temperature for 45 min in freshly prepared oxygenated artificial cerebrospinal fluid (aCSF). Cell injury was assessed by performing LDH activity assay. Brain slices were incubated individually at 36±0.5°C in oxygenated aCSF. After stabilisation of LDH release, the incubation aCSF was renewed and the slices were incubated for 1 h to allow recording basal LDH levels. Sham slices were incubated for an additional 20-min period in the same conditions. Brain slices subjected to OGD were incubated in a glucose-free solution continuously saturated with 100 vol% nitrogen. Then, to mimic reperfusion and treatment, the medium was replaced in all groups with aCSF continuously saturated with: (i) medical air; or (ii) argon of 25 vol% to 75 vol% (with the remainder being oxygen at 25 vol% completed with nitrogen when necessary). During the 3-h “reperfusion” period, aCSF was renewed every 1 h to obtain samples and to assess the kinetics of LDH release as a percentage change from LDH control value before OGD. The number of animals and slices per group was N = 4 and n = 23–27, respectively.
In vivo NMDA studies
NMDA studies were performed aas detailed previously [6]. Briefly, rats were anesthetised with 1.5% halothane in oxygen alone, mounted on a stereotaxic apparatus, allowed breathing spontaneously, and maintained normothermic at 37±0.5°C. Rats were given 70 nmol NMDA in 1 µL PBS (pH 7.4) into the right striatum. Total duration of surgery was no more than 10 min, thereby allowing rapid halothane and oxygen desaturation from the rat's body, and the virtual absence of residual halothane and oxygen during post-NMDA treatment with medical air or argon [46]. The animals waked up in their home cage after about 10 min, with free access to food and water. Sixty minutes after injection of NMDA, the rats were treated as described below with either (i) medical air (controls); or (ii) argon of 25 vol% to 75 vol% (with the remainder being 25 vol% oxygen completed with nitrogen when necessary). The number of rats per group was as follows: 75 vol% argon, n = 7; 50 vol% argon, n = 8; 37.5 vol% argon, n = 8; 25 vol% argon, n = 8; 15 vol% argon, n = 8; NMDA control group, n = 11. Sham rats were given an intrastriatal saline solution and medical air.
In vivo MCAO studies
MCAO studies were performed as described in details previously [6]. Briefly, rats were subjected to transient cerebral ischemia for 60 min by the intraluminal MCAO method using a nylon thread. Rats were anesthetised with 1.5% isoflurane in oxygen alone, allowed breathing spontaneously, and maintained normothermic at 37.5°C±0.5°C. Total duration of surgery was no more than 25 min, thereby allowing rapid isoflurane and oxygen desaturation from the rat's body, and little residual isoflurane and oxygen during the MCAO period [46]. After surgery, rats were allowed moving freely in their home wages with free access to food and water. Sixty minutes later, the nylon was removed under a short isoflurane anesthesia of less than 10 min duration, thereby allowing rapid isoflurane and oxygen desaturation from the rat's body, and the virtual absence of residual isoflurane and oxygen during the rats' reperfusion and 3-h post-MCAO treatment with either medical air (MCAO controls, n = 14) or argon at 50 vol% (n = 14) [46]. Treatment with medical air or argon at 50 vol% was performed according to a blinded procedure in a closed chamber at a flow rate of 1 chamber volume per minute, a condition that allowed maintaining carbon dioxide less than 0.03%. No analgesic treatment was given during recovery. Physiological parameters were recorded immediately before and immediately after treatment with medical air or argon at 50 vol%. Sham-treated rats (n = 6) were subjected to the same surgical protocol than controls and argon-treated rats, but not to ischemia, and exposed to medical air for 3 h.
The rats' neurologic outcome was assessed by recording the animals' motor coordination and rearing activity as detailed previously [6]. Motor coordination was quantified in individual non-motorized activity wheels equipped with 1/8 rotation sensors; rearing activity was quantified in individual activity cages equipped with infrared beams (Imetronic, Pessac, France). Beam interruptions and rotation signals were detected and recorded on a computer. Rats were familiarized with the activity cages and wheels. Then, on day 2 before MCAO (−48 h), and on day 1 (24 h) and day 2 (48 h) after MCAO, the rats were placed again and recorded for 15 min in the activity cages and the activity wheels. Post-MCAO scores of motor coordination and rearing activity were expressed as a percentage of pre-MCAO scores on day 2 before MCAO using each rat as its own control, and averaged to obtain a global score of neurologic outcome expressed as a percentage of that recorded in sham rats.
Histologic analysis
Fifty hours after MCAO or NMDA injection, coronal brain sections (20 µm) were cryostat-cut, stained with thionin, digitized on a computer and analysed using an image analyzer (ImageJ® software, Scion corp., USA). The infarction volume was calculated by integration over the whole brain of the infarcted surfaces delineated by the pallor of staining compared to the surrounding healthy tissue, corrected for tissue oedema when needed (MCAO-induced ischemia) by calculating and dividing volume by the ipsilateral/contralateral brain hemispheres ratio, and expressed in mm3.
Gas treatment
Oxygen, nitrogen and argon of medical grade were bought from Messer and Air Liquide. Gas mixtures containing 75 vol% nitrogen and 25 vol% oxygen (medical air), or argon of 15 vol% to 75 vol% (with the remainder being oxygen at 25 vol% completed with nitrogen when necessary) were obtained using calibrated flowmeters and gas analysers.
Statistical analysis
Data on OGD-induced LDH release (n>20) are expressed as the mean ± the standard error to the mean (SEM) and were analysed using one-way parametric ANOVA and post-hoc unpaired Student t-test. Data on MCAO-induced brain damage (n<20) are expressed as the median value ± the 25–75th percentiles and were analysed using the Kruskall-Wallis nonparametric ANOVA and/or post-hoc Mann-Whitney unpaired U-test. The number of animals per group for the in vivo studies was assessed through power analysis through online software (www.dssresearch.com/toolkit/sscalc/size_a2.asp). Because one objective of the study was to assess whether or not argon could be a cost-efficient alternative to xenon, the number of rats was calculated with the following variables from a previous study from our laboratory [6] with xenon at 50 vol%: mean ± standard deviation (SD) for ischemic brain damage volume in control group = 218±60 mm3; mean ± SD for ischemic brain damage volume in xenon-treated group = 59±28 mm3; α error level (confidence level) = 0.05; β error level (statistical power) = 0.01.
Author Contributions
Conceived and designed the experiments: HND BH DC JJR JHA. Performed the experiments: HND BH MD. Analyzed the data: HND BH MD. Wrote the paper: HND DC JJR JHA.
References
- 1. Dirnagl U, Iadecola C, Moskowitz MA (1999) Pathobiology of ischemic stroke: An integrated view. Trends Neurosci 22: 391–397.
- 2. Lee JM, Zipfel GJ, Choi DW (1999) The changing landscape of ischemic brain injury mechanisms. Nature 399: A7–14.
- 3. Lo EH, Moskowitz MA, Jacobs TP (1999) Exciting, radical, suicidal: How brain cells die after stroke. Stroke 36: 189–192.
- 4. Petzelt C, Blom P, Schmehl W, Muller J, Kox WJ (2003) Prevention of neurotoxicity in hypoxic cortical neurons by the noble gas xenon. Life Sci 72: 1909–1918.
- 5. David HN, Leveillé F, Chazalviel L, MacKenzie ET, Buisson A, et al. (2003) Reduction of ischemic brain damade by nitrous oxide and xenon. J Cereb Blood Flow Metab 23: 1168–1173.
- 6. David HN, Haelewyn B, Rouillon C, Lecocq M, Chazalviel L, et al. (2008) Neuroprotective effects of xenon: a therapeutic window of opportunity in rats subjected to transient cerebral ischemia. FASEB J 22: 1275–1286.
- 7. David HN, Haelewyn B, Risso JJ, Colloc'h N, Abraini JH (2010) Xenon is an inhibitor of tissue-plasminogen activator: adverse and beneficial effects in a rat model of thromboembolic stroke. J Cereb Blood Flow Metab 30: 718–728.
- 8. Homi HM, Yokoo N, Warner DS, Franks NP, Maze M, et al. (2003) The neuroprotective effect of xenon administration during transient middle cerebral artery occlusion in mice. Anesthesiology 99: 876–881.
- 9. Ma D, Yang H, Lynch J, Franks NP, Maze M, et al. (2003) Xenon attenuates cardiopulmonary bypass-induced neurologic and neurocognitive dysfunction in the rat. Anesthesiology 98: 690–698.
- 10. Ma D, Hossain M, Chow A, Arshad M, Battson RM, et al. (2005) Xenon and hypothermia combine to provide neuroprotection from neonatal asphyxia. Ann Neurol 58: 182–193.
- 11. Abraini JH, Lemaire M, David HN (2005) Potentially neuroprotective and therapeutic properties of nitrous oxide and xenon. Ann N Y Acad Sci 1053: 289–300.
- 12. Dingley J, Tooley J, Porter H, Thoresen M (2006) Xenon provides short-term neuroprotection in neonatal rats when administered after hypoxia-ischemia. Stroke 37: 501–506.
- 13. Natale G, Cattano D, Abramo A, Forfori F, Fulceri F, et al. (2006) Morphological evidence that xenon neuroprotects against N-methyl-DL-aspartic acid-induced damage in the rat arcuate nucleus: a time-dependent study. Ann N Y Acad Sci 1074: 650–658.
- 14. Yarin YM, Amarjargal N, Fuchs J, Haupt H, Mazurel B, et al. (2005) Argon protects hypoxia-, cisplatin- and gentamycin-exposed hair cells in the newborn rat's organ of Corti. Hear Res 201: 1–9.
- 15. Loetscher PD, Rossaint J, Rossaint R, Weis J, Fries M, et al. (2009) Argon: neuroprotection in in vitro models of cerebral ischemia and traumatic brain injury. Critical Care 13: R206.
- 16. Pagel PS, Krolikowski JG, Shim YH, Venkatapuram S, Kersten JR, et al. (2007) Noble gases without anesthetic properties protect myocardium against infarction by activating prosurvival signalling kinases and inhibiting mitochondrial permeability transition in vivo. Anesth Analg 105: 562–569.
- 17. Jawad N, Rizvi M, Gu J, Adeyi O, Tao G, et al. (2009) Neuroprotection (and lack of neuroprotection) afforded by a series of noble gases in an in vitro model of neuronal injury. Neurosci Lett 460: 232–236.
- 18. Ryang YM, Fahlenkamp AV, Rossaint R, Loetscher PD, Beyer C, et al. (2011) Neuroprotective effect of argon in an in vivo model of transient middle cerebral artery occlusion in rats. Crit Care Med 39: 1448–1453.
- 19. Rizvi M, Jawad N, Li Y, Vizcaychipi MP, Maze M, et al. (2010) Effect of noble gases on oxygen and glucose deprived injury in human tubular kidney cells. Exp Biol Med 235: 886–891.
- 20. Haelewyn B, David HN, Colloc'h N, Colomb DG Jr, Risso JJ, et al. (2011) Interactions between nitrous oxide and tissue plasminogen activator in a rat mdel of thromboembolic stroke. Anesthesiology 115: 1044–1053.
- 21.
Semenov N (1989) How I can go to idea. Short moment of triumph. Nauka: Moscow. pp. 6–19.
- 22.
Pavlov BN, Grigoreiv AI, Smolin VV, Komordin IP, Sokolv GM, et al. (1997) Investigations of different hyperoxic, hypoxic and normoxic oxygen-argon gaseous mixtures under different barometric pressure and respiration period. In: Bennett PB, Demchenko I, Marquis RE, editors. High Pressure Biology and Medicine. University of Rochester Press, NY. pp. 133–142.
- 23. Massieu L, Del Rio P, Montiel T (2001) Neurotoxicity of glutamate uptake inhibition in vivo: correlation with succinate dehydrogenase activity and prevention by energy substrates. Neuroscience 106: 669–677.
- 24. Wu KL, Hsu C, Chan JY (2007) Impairment of the mitochondrial respiratory enzyme activity triggers sequential activation of apoptosis-inducing factor-dependent and caspase-dependent signalling pathways to induced apoptosis after spinal cord injury. J Neurochem 101: 1552–1566.
- 25. Levy DI, Sucher NJ, Lipton SA (1990) Redox modulation of NMDA receptor-mediated toxicity in mammalian central neurons. Neurosci Lett 110: 291–296.
- 26. Sucher NJ, Wong LA, Lipton SA (1990) Redox modulation of NMDA receptor-mediated Ca2+ flux in mammalian central neurons. Neuroreport 1: 29–32.
- 27. Haelewyn B, Chalzaviel L, Lecocq M, Risso JJ, Abraini JH (2011) Moderately delayed post-insult treatment with normobaric hyperoxia reduces excitotoxin-induced neuronal degeneration but increases ischemia-induced brain damage. Med Gas Res 1: 2.
- 28. Haelewyn B, Rouillon C, Risso JJ, Abraini JH (2008) Functional (neurologic) recovery following transient focal cerebral ischemia in the rat requires at least 80% of ipsilateral cortical and subcortical integrity. Exp Neurol 213: 238–240.
- 29. David HN, Haelewyn B, Chazalviel L, Lecocq M, Degoulet M, et al. (2009) Post-ischemic helium provides neuroprotection in rats subjected to middle cerebral artery occlusion-induced ischemia by producing hypothermia. J Cereb Blood Flow Metab 29: 1159–1165.
- 30. Goto T, Suwa K, Uezono S, Ichinose F, Uchiyama M, et al. (1998) The blood-gas partition coefficient of xenon may be lower than generally accepted. Br J Anesth 80: 255–256.
- 31. Abraini JH, Rostain JC, Kriem B (1998) Sigmoidal compression rate-dependence of inert gas narcotic potency in rats: implication for lipid vs. protein theories of inert gas action in the central nervous system. Brain Res 808: 300–304.
- 32. Abraini JH, Kriem B, Balon N, Rostain JC, Risso JJ (2003) Gamma-aminobutyric neuropharmacological investigations on narcosis produced by nitrogen, argon, or nitrous oxide. Anesth Analg 96: 746–749.
- 33. Schwartz-Bloom RD, Miller KA, Evenson DA, Crain BJ, Nadler JV (2000) Benzodiazepines protect hippocampal neurons from degeneration after transient cerebral ischemia: an ultrastructural study. Neuroscience 98: 471–484.
- 34. Zhang F, Li C, Wang R, Han D, Zhang QG, et al. (2007) Activation of GABA receptors attenuates neuronal apoptosis through inhibiting the tyrosine phosphorylation of NR2A by Src after cerebral ischemia and reperfusion. Neuroscience 150: 938–949.
- 35. Lazar RM, Berman MF, Festa JR, Geller AE, Matejovsky TG, et al. (2010) GABAergic but not anti-cholinergic agents re-induce clinical deficits after stroke. J Neurol Sci 292: 72–76.
- 36. Yamakura T, Harris RA (2000) Effects of gaseous anesthetics nitrous oxide and xenon on ligand-gated ion channels. Comparison with isoflurane and ethanol. Anesthesiology 93: 1095–1101.
- 37. Tapper D, Arensman R, Johnson C, Folkman J (1974) The effect of helium-oxygen mixtures on body temperature. J Pediatric Surg 9: 597–603.
- 38. Sugaya T, Kitani Y (1997) Nitrous oxide attenuates the protective effect of isoflurane on microtubule-asssociated protein2 degradation during forbrain ischemia in the rat. Brain Res Bull 44: 307–309.
- 39. Olney JW, Farber NB, Wozniak DF, Jevtovic-Todorovic V, Ikonomidou C (2000) Environmental agents that have the potential to trigger massive apoptotic neurodegeneration in the developing brain. Environ Health Perpect 108: suppl 3383–388.
- 40. Abraini JH, David HN, MacKenzie ET, Lemaire M (2005) Postischemic nitrous oxide alone versus intraischemic nitrous oxide in the presence of isoflurane: What it may change for neuroprotection against cerebral stroke in the rat. Anesth Analg 101: 614.
- 41. Sanders RD, Ma D, Maze M (2005) Anesthesia-induced neuroprotection. Best Pract Res Clin Anesthesiol 19: 461–474.
- 42. Nasu I, Yokoo N, Takaoka S, Takata K, Hoshikawa T, et al. (2006) The dose-dependent effects of isoflurane on outcome from severe forebrain ischemia in the rat. Anesth Analg 103: 413–418.
- 43. Istaphanous GK, Howard J, Nan X, Hughes EA, McCaan JC, et al. (2011) Comparison of the neuroapoptotic properties of equipotent anesthetic concentrations of desflurane, isoflurane, or sevoflurane in neonatal mice. Anesthesiology 114: 578–587.
- 44. Singhal AB, Dijhuizen RM, Rosen BR, Lo EH (2002) Normobaric hyperoxia reduces MRI diffusion abnormalities and infarct size in experimental stroke. Neurology 58: 945–952.
- 45. Mickel HS, Vaishnav YN, Kempski O (1987) Breathing 100% oxygen after global brain ischemia in Mongolian gerbils results in increased lipid peroxydation and increased mortality. Stroke 18: 426–430.
- 46. White PF, Johnston RR, Eger EI 2nd (1974) Determination of anesthetic requirement in rats. Anesthesiology 40: 52–57.