Neurol Sci DOI 10.1007/s10072-014-1726-4
ORIGINAL ARTICLE
Sevoflurane postconditioning provides neuroprotection against brain hypoxia–ischemia in neonatal rats Xiaoyan Ren • Zhi Wang • Hong Ma Zhiyi Zuo
•
Received: 25 February 2014 / Accepted: 15 March 2014 Ó Springer-Verlag Italia 2014
Abstract Application of volatile anesthetics after brain ischemia provides neuroprotection in adult animals (anesthetic postconditioning). We tested whether postconditioning with sevoflurane, the most commonly used general anesthetic in pediatric anesthesia, reduced neonatal brain injury in rats. Seven-day-old Sprague–Dawley rats were subjected to brain hypoxia–ischemia (HI). They were postconditioned with sevoflurane in the presence or absence of 5-hydroxydecanoic acid, a mitochondrial KATP channel inhibitor. Sevoflurane postconditioning dosedependently reduced brain tissue loss observed 7 days after brain HI. This effect was induced by clinically relevant concentrations and abolished by 5-hydroxydecanoic acid. These results suggest that sevoflurane postconditioning protects neonatal brain against brain HI via mitochondrial KATP channels. Keywords Brain hypoxia–ischemia Neonates Postconditioning Sevoflurane
X. Ren H. Ma (&) Department of Anesthesiology, The First Hospital of China Medical University, Shenyang 110001, People’s Republic of China e-mail: [email protected] X. Ren Z. Wang Z. Zuo (&) Department of Anesthesiology, University of Virginia Health System, 1 Hospital Drive, PO Box 800710, Charlottesville, VA 22908-0710, USA e-mail: [email protected] Z. Wang Department of Anesthesiology, Sun Yat-Sen Memorial Hospital, Sun Yat-Sen University, Guangzhou 510120, China
Abbreviations 5-HD KATP channels HI
5-Hydroxydecanoic acid ATP sensitive potassium channels Hypoxia–ischemia
Introduction Neonatal brain injury occurs at about one in every 4,000 live births [1]. Most of them survive to adulthood and can have significant long-term neurological and cognitive impairment, such as cerebral palsy and epilepsy [2–4]. Thus, neonatal brain injury has a major impact on patients, their families and our society. However, there are no effective and practical interventions available for use in clinical practice to reduce neonatal brain injury, indicating an urgent need to identify these interventions. Neonatal brain injury is usually caused by brain hypoxia, ischemia or the combination of hypoxia and ischemia [1]. This situation is often modeled in rodents by inducing brain hypoxia–ischemia (HI) [5]. It has been shown that application of isoflurane, a volatile anesthetic used clinically, after focal brain ischemia provides neuroprotection in adult rodents [6]. This isoflurane postconditioning effect also protects brain against neonatal brain HI in rats [7]. However, isoflurane is now not commonly used in pediatric anesthesia in the USA. Sevoflurane is the most commonly used general anesthetic in pediatric anesthesia and also more commonly used in adults than isoflurane in current clinical practice in the USA and many other developed countries [8]. However, it is not known yet whether sevoflurane can induce a postconditioning effect against neonatal brain HI. Based on the information of isoflurane postconditioning, we hypothesize that sevoflurane postconditioning reduces
123
Neurol Sci
neonatal brain injury. To test this hypothesis, we applied sevoflurane after brain HI in neonatal rats. Because mitochondrial ATP-sensitive potassium (KATP) channels may be involved in neuroprotection and cardioprotection induced by volatile anesthetics [6, 9], we also determined whether mitochondrial KATP channels played a role in the sevoflurane postconditioning effects on neonatal rats using 5-hydroxydecanoic acid (5-HD), a specific inhibitor of mitochondrial KATP channels.
Methods All experimental protocols were approved by the Institutional Animal Care and Use Committee of the University of Virginia (Charlottesville, VA). All surgical and experimental procedures were carried out in accordance with the National Institutes of Health Guide for the Care and Use of Laboratory Animals (NIH publications number 80-23) revised in 2011. Efforts were made to minimize the number of animals used and their suffering. Neonatal brain hypoxia–ischemia modal Brain HI was performed in 7-day-old male and female Sprague–Dawley rats as described previously [10, 11]. In brief, neonates were anesthetized by isoflurane and their left common carotid arteries were permanently ligated with a double 7-0 surgical silk. The procedure lasted \5 min. After surgery, neonates were returned to the cages with their mothers for 3 h. The neonates were then placed in a chamber filled with humidified 8 % oxygen–92 % nitrogen for 2 h at 37 °C. The oxygen concentration and temperature in the chamber were continuously monitored.
Fig. 1 Neuroprotective effects of sevoflurane. Seven-day-old rats were subjected to or were not subjected to the brain HI and then postconditioned with various concentrations of sevoflurane. Brain was harvested at 7 days after the brain HI. The results are mean ± SD (n = 8–10). *P \ 0.05 compared with control rats. ^P \ 0.05 compared with rats that had brain HI only
Brain injury/tissue loss quantification After 7 days of the brain HI, rats were sacrificed under deep isoflurane anesthesia and then their brains were harvested as described previously [11, 13]. The hindbrain was removed from cerebral hemispheres and bilateral hemispheres were weighed separately. The weight ratio of left to right hemispheres was calculated. Statistical analysis The results are presented as mean ± SD (n C 6). Statistical analysis was performed by one-way analysis of variance followed by the Tukey’s test. A P B 0.05 was considered statistically significant.
Drug application The neonates were randomly divided into the following groups: (1) control, (2) brain HI, (3) brain HI and postconditioning with 1, 2 and 3 % sevoflurane, (4) brain HI and 5-HD treatment (10 mg/kg) and (5) brain HI, 5-HD treatment and postconditioning with 2 % sevoflurane. Sevoflurane postconditioning was performed by exposing neonates to various concentrations of sevoflurane in 30 % O2 for 1 h immediately after brain HI. Neonates of brain HI alone group were placed in a chamber flushed with 30 % O2 for 1 h. The mitochondrial KATP channel inhibitor 5-HD was dissolved in normal saline and administered intraperitoneally just before the start of brain HI. The dose of 5-HD was based on a previous study in which intraperitoneal injection of 10 mg/kg 5-HD blocked ischemic preconditioning-induced protection [12].
123
Results The control rats had similar right and left hemisphere weights. The left brain HI significantly reduced left cerebral hemisphere weight examined 7 days after brain HI (weight ratio of left to right cerebral hemispheres: 0.987 ± 0.012 of control group vs. 0.740 ± 0.049 of brain HI group, n = 8–10, P \ 0.001), suggesting brain tissue loss in this side of brain. This tissue loss was dosedependently reduced by sevoflurane postconditioning, an effect that was already significant even after being postconditioned with 1 % sevoflurane (Fig. 1) (weight ratio of left to right cerebral hemispheres: 0.740 ± 0.049 of brain HI group vs. 0.820 ± 0.051 of brain HI plus 1 % sevoflurane group, n = 8–10, P = 0.003). The sevoflurane
Neurol Sci
Fig. 2 Inhibition of sevoflurane postconditioning-induced neuroprotection by a mitochondrial KATP inhibitor. Seven-day-old rats were subjected to or were not subjected to the brain HI and then postconditioned with 2 % sevoflurane in the presence or absence of 10 mg/kg 5-HD. a Representative brain images. The inserted length marker = 2 mm; b Quantification results. The results are mean ± SD (n = 6–10). *P \ 0.05 compared with control rats, ^P \ 0.05 compared with rats that had brain HI only, #P \ 0.05 compared with rats that had brain HI and sevoflurane postconditioning. HD 5-HD, Sevo sevoflurane
postconditioning effects were abolished by 5-HD (weight ratio of left to right cerebral hemispheres: 0.884 ± 0.049 of brain HI plus 2 % sevoflurane group vs. 0.759 ± 0.044 of brain HI plus 2 % sevoflurane plus 5-HD group, n = 8–10, P \ 0.001), although 5-HD alone did not affect the brain tissue loss caused by brain HI (Fig. 2).
Discussion Our results clearly showed that postconditioning with sevoflurane-induced neuroprotection against neonatal brain HI. These results extend the previous finding of isoflurane postconditioning effects in neonatal rats [7] to sevoflurane,
the most commonly used general anesthetic in pediatric anesthesia. To test the effects of sevoflurane, we used the classical Rice–Vannucci model, a widely used and wellcharacterized animal model to study neonatal brain injury. This model incorporates both brain ischemia and hypoxia, factors that contribute to neonatal brain injury in humans [1, 5]. We monitored brain tissue loss that signifies brain structure damage as showed in our previous studies [11, 13]. Our results suggest that the sevoflurane postconditioning effects may be mediated by mitochondrial KATP channels because 5-HD, a mitochondrial KATP channel inhibitor, abolished sevoflurane effects and 5-HD alone did not alter brain tissue loss after brain HI. Consistent with this finding, our previous study has implicated the involvement of mitochondrial KATP channels in isoflurane postconditioning-induced neuroprotection in adult rats after focal brain ischemia [6]. These channels are inhibited by ATP and activated under energy-depleted conditions. They also can be activated by volatile anesthetics [14]. The opening of these channels produces an outward current. This current can maintain the mitochondrial membrane potential and reduce the opening of mitochondrial permeability transition pore to inhibit cell injury and death [15, 16]. Of note, previous studies have shown that application of sevoflurane before brain ischemia (sevoflurane preconditioning) also provides neuroprotection. This effect may be mediated by inhibition of brain inflammation and activation of glutamate transporters in the brain [17–19]. Although these mechanisms play a role in the sevoflurane postconditioning-induced neuroprotection and whether there is a relationship between these mechanisms and the mitochondrial KATP channel pathway require further investigation. We used cerebral weight ratio to quantify the brain tissue loss after brain HI. The method is used by many studies before and the brain injury quantified by this method is highly correlated with that measured by biochemical, electrophysiological and morphometric methods [20, 21]. In addition, the cerebral weight ratio method is simple to perform and very objective. Our finding may have clinical translational implications. Sevoflurane is the most commonly used general anesthetic in pediatric population [8]. It is often used in patients for Cesarean section if general anesthesia is used. One minimum alveolar concentration (the concentration in the lungs to prevent movement in 50 % subjects in response to surgical stimuli) of sevoflurane is *2 % in human [22] and 2.9 % in rats younger than 30 days [23]. Our results showed that sevoflurane at a concentration as low as 1 % was effective to induce neuroprotection, suggesting that a subclinical concentration for anesthesia can induce the postconditioning effect. In addition, we have demonstrated the postconditioning effect, which may be easy to apply
123
Neurol Sci
clinically because its application does not require the predication of ischemia occurrence. However, exposure of neonatal mice to 3 % sevoflurane for 6 h can lead to brain cell apoptosis and increase in brain tumor necrosis factor a [24]. Thus, it is judicious to avoid exposure to a high concentration of sevoflurane for a long time so that the potential sevoflurane-induced neurotoxicity will not occur. In summary, we have shown that sevoflurane at clinically relevant concentrations can induce a postconditioning effect against neonatal brain injury. This effect may be mediated by mitochondrial KATP channels. Acknowledgments This study was supported by Grants (R01 GM065211 and R01 GM098308 to Z. Zuo) from the National Institutes of Health, Bethesda, Maryland, by a Grant from the International Anesthesia Research Society (2007 Frontiers in Anesthesia Research Award to Z. Zuo), Cleveland, Ohio, by a Grant-in-Aid from the American Heart Association Mid-Atlantic Affiliate (10GRNT3900019 to Z. Zuo), Baltimore, Maryland, and the Robert M. Epstein Professorship Endowment, University of Virginia. Conflict of interest
None.
References 1. Ferriero DM (2004) Neonatal brain injury. N Engl J Med 351(19):1985–1995 2. Lynch JK, Nelson KB (2001) Epidemiology of perinatal stroke. Curr Opin Pediatr 13:499–505 3. Sran SK, Baumann RJ (1988) Outcome of neonatal strokes. Am J Dis Child 142:1086–1088 4. Sreenan C, Bhargave R, Robertson CM (2000) Cerebral infarction in the term new-born: clinical presentation and long-term outcome. J Pediatr 137:351–355 5. Silbereis JC, Huang EJ, Back SA, Rowitch DH (2010) Towards improved animal models of neonatal white matter injury associated with cerebral palsy. Dis Models Mech 3(11–12):678–688 6. Lee JJ, Li L, Jung H–H, Zuo Z (2008) Postconditioning with isoflurane reduced ischemia-induced brain injury in rats. Anesthesiology 108:1055–1062 7. Zhou Y, Lekic T, Fathali N, Ostrowski RP, Martin RD, Tang J et al (2010) Isoflurane posttreatment reduces neonatal hypoxicischemic brain injury in rats by the sphingosine-1-phosphate/ phosphatidylinositol-3-kinase/Akt pathway. Stroke 41(7): 1521–1527 8. Fox AJ, Rowbotham DJ (1999) Anaesthesia. BMJ 319(7209): 557–560 9. Obal D, Dettwiler S, Favoccia C, Scharbatke H, Preckel B, Schlack W (2005) The influence of mitochondrial KATP-channels in the cardioprotection of preconditioning and postconditioning by sevoflurane in the rat in vivo. Anesth Analg 101(5):1252–1260
123
10. Zhao P, Peng L, Li L, Xu X, Zuo Z (2007) Isoflurane preconditioning improves long-term neurologic outcome after hypoxicischemic brain injury in neonatal rats. Anesthesiology 107(6):963–970 11. Wang Z, Zhao H, Peng S, Zuo Z (2013) Intranasal pyrrolidine dithiocarbamate decreases brain inflammatory mediators and provides neuroprotection after brain hypoxia-ischemia in neonatal rats. Exp Neurol 249:74–82 12. Gozen A, Demiryurek S, Taskin A, Ciralik H, Bilinc H, Kara S et al (2013) Protective activity of ischemic preconditioning on rat testicular ischemia: effects of Y-27632 and 5-hydroxydecanoic acid. J Pediatr Surg 48(7):1565–1572 13. Zhao P, Zuo Z (2004) Isoflurane preconditioning induces neuroprotection that is inducible nitric oxide synthase-dependent in the neonatal rats. Anesthesiology 101:695–702 14. Jiang MT, Nakae Y, Ljubkovic M, Kwok WM, Stowe DF, Bosnjak ZJ (2007) Isoflurane activates human cardiac mitochondrial adenosine triphosphate-sensitive K? channels reconstituted in lipid bilayers. Anesth Analg 105(4):926–932 table of contents 15. Kowaltowski AJ, Seetharaman S, Paucek P, Garlid KD (2001) Bioenergetic consequences of opening the ATP-sensitive K(?) channel of heart mitochondria. Am J Physiol Heart Circ Physiol 280(2):H649–H657 16. Gateau-Roesch O, Argaud L, Ovize M (2006) Mitochondrial permeability transition pore and postconditioning. Cardiovasc Res 70(2):264–273 17. Wang H, Lu S, Yu Q, Liang W, Gao H, Li P et al (2011) Sevoflurane preconditioning confers neuroprotection via antiinflammatory effects. Front Biosci 3(1):604–615 18. Sanders RD, Manning HJ, Robertson NJ, Ma D, Edwards AD, Hagberg H et al (2010) Preconditioning and postinsult therapies for perinatal hypoxic-ischemic injury at term. Anesthesiology 113(1):233–249 19. Wang C, Lee J, Jung H, Zuo Z (2007) Pretreatment with volatile anesthetics, but not with the nonimmobilizer 1,2-dichlorohexafluorocyclobutane, reduced cell injury in rat cerebellar slices after an in vitro simulated ischemia. Brain Res 1152:201–208 20. Andine P, Thordstein M, Kjellmer I, Nordborg C, Thiringer K, Wennberg E et al (1990) Evaluation of brain damage in a rat model of neonatal hypoxic-ischemia. J Neurosci Methods 35(3):253–260 21. McDonald JW, Roeser NF, Silverstein FS, Johnston MV (1989) Quantitative assessment of neuroprotection against NMDAinduced brain injury. Exp Neurol 106(3):289–296 22. Behne M, Wilke HJ, Harder S (1999) Clinical pharmacokinetics of sevoflurane. Clin Pharmacokinet 36(1):13–26 23. Orliaguet G, Vivien B, Langeron O, Bouhemad B, Coriat P, Riou B (2001) Minimum alveolar concentration of volatile anesthetics in rats during postnatal maturation. Anesthesiology 95(3): 734–739 24. Lu Y, Wu X, Dong Y, Xu Z, Zhang Y, Xie Z (2010) Anesthetic sevoflurane causes neurotoxicity differently in neonatal naive and Alzheimer disease transgenic mice. Anesthesiology 112(6): 1404–1416
ORIGINAL ARTICLE
Sevoflurane postconditioning provides neuroprotection against brain hypoxia–ischemia in neonatal rats Xiaoyan Ren • Zhi Wang • Hong Ma Zhiyi Zuo
•
Received: 25 February 2014 / Accepted: 15 March 2014 Ó Springer-Verlag Italia 2014
Abstract Application of volatile anesthetics after brain ischemia provides neuroprotection in adult animals (anesthetic postconditioning). We tested whether postconditioning with sevoflurane, the most commonly used general anesthetic in pediatric anesthesia, reduced neonatal brain injury in rats. Seven-day-old Sprague–Dawley rats were subjected to brain hypoxia–ischemia (HI). They were postconditioned with sevoflurane in the presence or absence of 5-hydroxydecanoic acid, a mitochondrial KATP channel inhibitor. Sevoflurane postconditioning dosedependently reduced brain tissue loss observed 7 days after brain HI. This effect was induced by clinically relevant concentrations and abolished by 5-hydroxydecanoic acid. These results suggest that sevoflurane postconditioning protects neonatal brain against brain HI via mitochondrial KATP channels. Keywords Brain hypoxia–ischemia Neonates Postconditioning Sevoflurane
X. Ren H. Ma (&) Department of Anesthesiology, The First Hospital of China Medical University, Shenyang 110001, People’s Republic of China e-mail: [email protected] X. Ren Z. Wang Z. Zuo (&) Department of Anesthesiology, University of Virginia Health System, 1 Hospital Drive, PO Box 800710, Charlottesville, VA 22908-0710, USA e-mail: [email protected] Z. Wang Department of Anesthesiology, Sun Yat-Sen Memorial Hospital, Sun Yat-Sen University, Guangzhou 510120, China
Abbreviations 5-HD KATP channels HI
5-Hydroxydecanoic acid ATP sensitive potassium channels Hypoxia–ischemia
Introduction Neonatal brain injury occurs at about one in every 4,000 live births [1]. Most of them survive to adulthood and can have significant long-term neurological and cognitive impairment, such as cerebral palsy and epilepsy [2–4]. Thus, neonatal brain injury has a major impact on patients, their families and our society. However, there are no effective and practical interventions available for use in clinical practice to reduce neonatal brain injury, indicating an urgent need to identify these interventions. Neonatal brain injury is usually caused by brain hypoxia, ischemia or the combination of hypoxia and ischemia [1]. This situation is often modeled in rodents by inducing brain hypoxia–ischemia (HI) [5]. It has been shown that application of isoflurane, a volatile anesthetic used clinically, after focal brain ischemia provides neuroprotection in adult rodents [6]. This isoflurane postconditioning effect also protects brain against neonatal brain HI in rats [7]. However, isoflurane is now not commonly used in pediatric anesthesia in the USA. Sevoflurane is the most commonly used general anesthetic in pediatric anesthesia and also more commonly used in adults than isoflurane in current clinical practice in the USA and many other developed countries [8]. However, it is not known yet whether sevoflurane can induce a postconditioning effect against neonatal brain HI. Based on the information of isoflurane postconditioning, we hypothesize that sevoflurane postconditioning reduces
123
Neurol Sci
neonatal brain injury. To test this hypothesis, we applied sevoflurane after brain HI in neonatal rats. Because mitochondrial ATP-sensitive potassium (KATP) channels may be involved in neuroprotection and cardioprotection induced by volatile anesthetics [6, 9], we also determined whether mitochondrial KATP channels played a role in the sevoflurane postconditioning effects on neonatal rats using 5-hydroxydecanoic acid (5-HD), a specific inhibitor of mitochondrial KATP channels.
Methods All experimental protocols were approved by the Institutional Animal Care and Use Committee of the University of Virginia (Charlottesville, VA). All surgical and experimental procedures were carried out in accordance with the National Institutes of Health Guide for the Care and Use of Laboratory Animals (NIH publications number 80-23) revised in 2011. Efforts were made to minimize the number of animals used and their suffering. Neonatal brain hypoxia–ischemia modal Brain HI was performed in 7-day-old male and female Sprague–Dawley rats as described previously [10, 11]. In brief, neonates were anesthetized by isoflurane and their left common carotid arteries were permanently ligated with a double 7-0 surgical silk. The procedure lasted \5 min. After surgery, neonates were returned to the cages with their mothers for 3 h. The neonates were then placed in a chamber filled with humidified 8 % oxygen–92 % nitrogen for 2 h at 37 °C. The oxygen concentration and temperature in the chamber were continuously monitored.
Fig. 1 Neuroprotective effects of sevoflurane. Seven-day-old rats were subjected to or were not subjected to the brain HI and then postconditioned with various concentrations of sevoflurane. Brain was harvested at 7 days after the brain HI. The results are mean ± SD (n = 8–10). *P \ 0.05 compared with control rats. ^P \ 0.05 compared with rats that had brain HI only
Brain injury/tissue loss quantification After 7 days of the brain HI, rats were sacrificed under deep isoflurane anesthesia and then their brains were harvested as described previously [11, 13]. The hindbrain was removed from cerebral hemispheres and bilateral hemispheres were weighed separately. The weight ratio of left to right hemispheres was calculated. Statistical analysis The results are presented as mean ± SD (n C 6). Statistical analysis was performed by one-way analysis of variance followed by the Tukey’s test. A P B 0.05 was considered statistically significant.
Drug application The neonates were randomly divided into the following groups: (1) control, (2) brain HI, (3) brain HI and postconditioning with 1, 2 and 3 % sevoflurane, (4) brain HI and 5-HD treatment (10 mg/kg) and (5) brain HI, 5-HD treatment and postconditioning with 2 % sevoflurane. Sevoflurane postconditioning was performed by exposing neonates to various concentrations of sevoflurane in 30 % O2 for 1 h immediately after brain HI. Neonates of brain HI alone group were placed in a chamber flushed with 30 % O2 for 1 h. The mitochondrial KATP channel inhibitor 5-HD was dissolved in normal saline and administered intraperitoneally just before the start of brain HI. The dose of 5-HD was based on a previous study in which intraperitoneal injection of 10 mg/kg 5-HD blocked ischemic preconditioning-induced protection [12].
123
Results The control rats had similar right and left hemisphere weights. The left brain HI significantly reduced left cerebral hemisphere weight examined 7 days after brain HI (weight ratio of left to right cerebral hemispheres: 0.987 ± 0.012 of control group vs. 0.740 ± 0.049 of brain HI group, n = 8–10, P \ 0.001), suggesting brain tissue loss in this side of brain. This tissue loss was dosedependently reduced by sevoflurane postconditioning, an effect that was already significant even after being postconditioned with 1 % sevoflurane (Fig. 1) (weight ratio of left to right cerebral hemispheres: 0.740 ± 0.049 of brain HI group vs. 0.820 ± 0.051 of brain HI plus 1 % sevoflurane group, n = 8–10, P = 0.003). The sevoflurane
Neurol Sci
Fig. 2 Inhibition of sevoflurane postconditioning-induced neuroprotection by a mitochondrial KATP inhibitor. Seven-day-old rats were subjected to or were not subjected to the brain HI and then postconditioned with 2 % sevoflurane in the presence or absence of 10 mg/kg 5-HD. a Representative brain images. The inserted length marker = 2 mm; b Quantification results. The results are mean ± SD (n = 6–10). *P \ 0.05 compared with control rats, ^P \ 0.05 compared with rats that had brain HI only, #P \ 0.05 compared with rats that had brain HI and sevoflurane postconditioning. HD 5-HD, Sevo sevoflurane
postconditioning effects were abolished by 5-HD (weight ratio of left to right cerebral hemispheres: 0.884 ± 0.049 of brain HI plus 2 % sevoflurane group vs. 0.759 ± 0.044 of brain HI plus 2 % sevoflurane plus 5-HD group, n = 8–10, P \ 0.001), although 5-HD alone did not affect the brain tissue loss caused by brain HI (Fig. 2).
Discussion Our results clearly showed that postconditioning with sevoflurane-induced neuroprotection against neonatal brain HI. These results extend the previous finding of isoflurane postconditioning effects in neonatal rats [7] to sevoflurane,
the most commonly used general anesthetic in pediatric anesthesia. To test the effects of sevoflurane, we used the classical Rice–Vannucci model, a widely used and wellcharacterized animal model to study neonatal brain injury. This model incorporates both brain ischemia and hypoxia, factors that contribute to neonatal brain injury in humans [1, 5]. We monitored brain tissue loss that signifies brain structure damage as showed in our previous studies [11, 13]. Our results suggest that the sevoflurane postconditioning effects may be mediated by mitochondrial KATP channels because 5-HD, a mitochondrial KATP channel inhibitor, abolished sevoflurane effects and 5-HD alone did not alter brain tissue loss after brain HI. Consistent with this finding, our previous study has implicated the involvement of mitochondrial KATP channels in isoflurane postconditioning-induced neuroprotection in adult rats after focal brain ischemia [6]. These channels are inhibited by ATP and activated under energy-depleted conditions. They also can be activated by volatile anesthetics [14]. The opening of these channels produces an outward current. This current can maintain the mitochondrial membrane potential and reduce the opening of mitochondrial permeability transition pore to inhibit cell injury and death [15, 16]. Of note, previous studies have shown that application of sevoflurane before brain ischemia (sevoflurane preconditioning) also provides neuroprotection. This effect may be mediated by inhibition of brain inflammation and activation of glutamate transporters in the brain [17–19]. Although these mechanisms play a role in the sevoflurane postconditioning-induced neuroprotection and whether there is a relationship between these mechanisms and the mitochondrial KATP channel pathway require further investigation. We used cerebral weight ratio to quantify the brain tissue loss after brain HI. The method is used by many studies before and the brain injury quantified by this method is highly correlated with that measured by biochemical, electrophysiological and morphometric methods [20, 21]. In addition, the cerebral weight ratio method is simple to perform and very objective. Our finding may have clinical translational implications. Sevoflurane is the most commonly used general anesthetic in pediatric population [8]. It is often used in patients for Cesarean section if general anesthesia is used. One minimum alveolar concentration (the concentration in the lungs to prevent movement in 50 % subjects in response to surgical stimuli) of sevoflurane is *2 % in human [22] and 2.9 % in rats younger than 30 days [23]. Our results showed that sevoflurane at a concentration as low as 1 % was effective to induce neuroprotection, suggesting that a subclinical concentration for anesthesia can induce the postconditioning effect. In addition, we have demonstrated the postconditioning effect, which may be easy to apply
123
Neurol Sci
clinically because its application does not require the predication of ischemia occurrence. However, exposure of neonatal mice to 3 % sevoflurane for 6 h can lead to brain cell apoptosis and increase in brain tumor necrosis factor a [24]. Thus, it is judicious to avoid exposure to a high concentration of sevoflurane for a long time so that the potential sevoflurane-induced neurotoxicity will not occur. In summary, we have shown that sevoflurane at clinically relevant concentrations can induce a postconditioning effect against neonatal brain injury. This effect may be mediated by mitochondrial KATP channels. Acknowledgments This study was supported by Grants (R01 GM065211 and R01 GM098308 to Z. Zuo) from the National Institutes of Health, Bethesda, Maryland, by a Grant from the International Anesthesia Research Society (2007 Frontiers in Anesthesia Research Award to Z. Zuo), Cleveland, Ohio, by a Grant-in-Aid from the American Heart Association Mid-Atlantic Affiliate (10GRNT3900019 to Z. Zuo), Baltimore, Maryland, and the Robert M. Epstein Professorship Endowment, University of Virginia. Conflict of interest
None.
References 1. Ferriero DM (2004) Neonatal brain injury. N Engl J Med 351(19):1985–1995 2. Lynch JK, Nelson KB (2001) Epidemiology of perinatal stroke. Curr Opin Pediatr 13:499–505 3. Sran SK, Baumann RJ (1988) Outcome of neonatal strokes. Am J Dis Child 142:1086–1088 4. Sreenan C, Bhargave R, Robertson CM (2000) Cerebral infarction in the term new-born: clinical presentation and long-term outcome. J Pediatr 137:351–355 5. Silbereis JC, Huang EJ, Back SA, Rowitch DH (2010) Towards improved animal models of neonatal white matter injury associated with cerebral palsy. Dis Models Mech 3(11–12):678–688 6. Lee JJ, Li L, Jung H–H, Zuo Z (2008) Postconditioning with isoflurane reduced ischemia-induced brain injury in rats. Anesthesiology 108:1055–1062 7. Zhou Y, Lekic T, Fathali N, Ostrowski RP, Martin RD, Tang J et al (2010) Isoflurane posttreatment reduces neonatal hypoxicischemic brain injury in rats by the sphingosine-1-phosphate/ phosphatidylinositol-3-kinase/Akt pathway. Stroke 41(7): 1521–1527 8. Fox AJ, Rowbotham DJ (1999) Anaesthesia. BMJ 319(7209): 557–560 9. Obal D, Dettwiler S, Favoccia C, Scharbatke H, Preckel B, Schlack W (2005) The influence of mitochondrial KATP-channels in the cardioprotection of preconditioning and postconditioning by sevoflurane in the rat in vivo. Anesth Analg 101(5):1252–1260
123
10. Zhao P, Peng L, Li L, Xu X, Zuo Z (2007) Isoflurane preconditioning improves long-term neurologic outcome after hypoxicischemic brain injury in neonatal rats. Anesthesiology 107(6):963–970 11. Wang Z, Zhao H, Peng S, Zuo Z (2013) Intranasal pyrrolidine dithiocarbamate decreases brain inflammatory mediators and provides neuroprotection after brain hypoxia-ischemia in neonatal rats. Exp Neurol 249:74–82 12. Gozen A, Demiryurek S, Taskin A, Ciralik H, Bilinc H, Kara S et al (2013) Protective activity of ischemic preconditioning on rat testicular ischemia: effects of Y-27632 and 5-hydroxydecanoic acid. J Pediatr Surg 48(7):1565–1572 13. Zhao P, Zuo Z (2004) Isoflurane preconditioning induces neuroprotection that is inducible nitric oxide synthase-dependent in the neonatal rats. Anesthesiology 101:695–702 14. Jiang MT, Nakae Y, Ljubkovic M, Kwok WM, Stowe DF, Bosnjak ZJ (2007) Isoflurane activates human cardiac mitochondrial adenosine triphosphate-sensitive K? channels reconstituted in lipid bilayers. Anesth Analg 105(4):926–932 table of contents 15. Kowaltowski AJ, Seetharaman S, Paucek P, Garlid KD (2001) Bioenergetic consequences of opening the ATP-sensitive K(?) channel of heart mitochondria. Am J Physiol Heart Circ Physiol 280(2):H649–H657 16. Gateau-Roesch O, Argaud L, Ovize M (2006) Mitochondrial permeability transition pore and postconditioning. Cardiovasc Res 70(2):264–273 17. Wang H, Lu S, Yu Q, Liang W, Gao H, Li P et al (2011) Sevoflurane preconditioning confers neuroprotection via antiinflammatory effects. Front Biosci 3(1):604–615 18. Sanders RD, Manning HJ, Robertson NJ, Ma D, Edwards AD, Hagberg H et al (2010) Preconditioning and postinsult therapies for perinatal hypoxic-ischemic injury at term. Anesthesiology 113(1):233–249 19. Wang C, Lee J, Jung H, Zuo Z (2007) Pretreatment with volatile anesthetics, but not with the nonimmobilizer 1,2-dichlorohexafluorocyclobutane, reduced cell injury in rat cerebellar slices after an in vitro simulated ischemia. Brain Res 1152:201–208 20. Andine P, Thordstein M, Kjellmer I, Nordborg C, Thiringer K, Wennberg E et al (1990) Evaluation of brain damage in a rat model of neonatal hypoxic-ischemia. J Neurosci Methods 35(3):253–260 21. McDonald JW, Roeser NF, Silverstein FS, Johnston MV (1989) Quantitative assessment of neuroprotection against NMDAinduced brain injury. Exp Neurol 106(3):289–296 22. Behne M, Wilke HJ, Harder S (1999) Clinical pharmacokinetics of sevoflurane. Clin Pharmacokinet 36(1):13–26 23. Orliaguet G, Vivien B, Langeron O, Bouhemad B, Coriat P, Riou B (2001) Minimum alveolar concentration of volatile anesthetics in rats during postnatal maturation. Anesthesiology 95(3): 734–739 24. Lu Y, Wu X, Dong Y, Xu Z, Zhang Y, Xie Z (2010) Anesthetic sevoflurane causes neurotoxicity differently in neonatal naive and Alzheimer disease transgenic mice. Anesthesiology 112(6): 1404–1416
