糖皮质激素加剧缺血诱导神经细胞损害的研究

目的观察糖皮质激素对神经细胞缺血性损害的影响。方法21只雄性沙士鼠随机分为3组，未缺血组(A组，n=7)，缺血组(B组，n=7)和实验组(C组，n=7)。A和B组脑室内给予生理盐水10μl，而C组给与地塞米松3μg。A组未作缺血处理，B组和C组1h后阻断双侧颈总动脉，2.5min后恢复血流。实验过程中控制脑温度在37.5℃±0.2℃。1周后将动物脑组织取出，做5μm厚的脑组织切片，经苏木素-伊红染色后计数海马CA1区1mm长度内残存的锥体细胞数。结果(1)与A组相比，B组和C组残存细胞数明显减少(P＜0.01);(2)C组的残存细胞数明显少于B组(P＜0.01)。结论(1)前脑缺血2.5min可致海马CA1区的锥体神经细胞的明显损害；(2)地塞米松明显加剧缺血时的脑细胞死亡。     

Systemic administration of procaine suppresses the somato-sympathetic reflex discharges in anesthetized cats

Somato-sympathetic reflex discharges (SSRDs) which were induced from the lumbar sympathetic truck elicited by train pulse stimulation of femoral nerves in anesthetized cats were examined during and after intravenous infusion of 1mg·kg^–1·min^–1 of procaine. The infusion demonstrated that amplitudes were suppressed with prolonged latencies and/or their tendencies in A- and C-reflex potentials of SSRDs. Heart rate and blood pressure decreased slightly, and the decreases correlated with time courses of suppressed SSRDs. These results suggest that the systemic procaine infusion may be a reliable method to alleviate somato-sympathetic reflexes under such conditions as extreme stimulation of somatic afferent nerves observed in surgical procedures.(Xue FW, Ogawa S, Nakamura T, et al.: Systemic administration of procaine suppresses the somato-sympathetic reflex discharges in anesthetized cats. J Anesth 6: 461–466, 1992)     

Effect of CNTF on ischaemic cell damage in rat hippocampus

Summary The neuroprotective effect of neurotrophic factors has been demonstrated in experimental cerebral ischaemia recently. These include nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF), insulin-like growth factor-1 (IGF-1), and basic fibroblast growth factor (basic FGF). The neuroprotective effect of ciliary neurotrophic factor (CNTF), however, has not been studied so far.We have examined the neuroprotective effect of recombinant rat CNTF in a rat forebrain ischaemia model. A continuous infusion of CNTF was started 1 week before the induction of ischaemia and continued until 1 week after the ischaemia. Reversible forebrain ischaemia was induced by 7 minutes of bilateral carotid occlusion with hypotension. Neuronal cell death in the hippocampal CA1 sector was evaluated 1 week after the ischaemia. For the control group artificial CSF (cerebrospinal fluid) was infused instead of CNTF.Per cent neuronal cell death was 83.4 ± 5.9% (mean ± SEM, n=5) in the control group, and 71.1 ± 10.0% (mean ± SEM, n=5) in the CNTF group. Although percentage of neuronal cell death was lower in the CNTF group, the difference was not statistically significant.This result suggests that the protective effect of CNTF in the rat forebrain ischaemia model may be limited compared with other neurotrophic factors. It is considered that the number of neurons protected by CNTF may be small.     

Sevoflurane reduces synaptic glutamate release in human synaptosomes

Volatile anesthetics reduce excitatory synaptic transmission in the mammalian brain. In the present study, the effect of sevoflurane on synaptic glutamate release, free cytosolic Ca2+ ([Ca2+]i), and glutamate uptake was investigated using isolated presynaptic terminals prepared from human cerebral cortex. The tissue was obtained from standard temporal lobe specimens removed because of epilepsy. The glutamate release and [Ca2+]i was measured as the fluorescence of nicotinamide adenine dinucleotide phosphate (NADPH) and fura-2, respectively. The uptake of radiolabeled glutamate was measured in a beta-scintillation counter. Membrane depolarization with 4-aminopyridine for three minutes evoked a Ca2+-dependent glutamate release of 3.4 +/- 0.5 nmol/mg. Sevoflurane 2.5 and 4.0% attenuated the evoked release by 45 and 55%, respectively. The evoked increase in [Ca2+]i was not significantly altered by the anesthetic agent. The uptake studies were performed in the high-affinity area, and Km was calculated to 19.3 +/- 5.7 x 10(-6) M and Vmax to 5.7 +/- 1.0 micromol g(-1) min(-1). The Km and Vmax values were not significantly altered by sevoflurane 2.5%. These results demonstrate that sevoflurane in the human brain reduces Ca2+-dependent glutamate release. The exact mode of action is still to be resolved.     

Adenosine reduces glutamate release in rat spinal synaptosomes

BACKGROUND: A1 adenosine receptor activation reduces hypersensitivity in animal models of chronic pain, but intrathecal adenosine does not produce analgesia to acute noxious stimuli. Here, the authors test whether increased inhibition by adenosine of glutamate release from afferents after injury accounts for this difference. METHODS: Synaptosomes were prepared from the dorsal half of the lumbar spinal cord of normal rats or those with spinal nerve ligation. Glutamate release evoked by the TRPV-1 receptor agonist, capsaicin, was measured. Adenosine with or without adenosine A1 and A2 receptor antagonists was applied to determine the efficacy and mechanism of adenosine to reduce capsaicin-evoked glutamate release. RESULTS: Capsaicin produced a concentration-dependent glutamate release similarly in normal and nerve-injured rats. Capsaicin-evoked glutamate release was inhibited by adenosine or R-PIA (R-N6-(2- phenylisopropyl)-adenosine) in a concentration-dependent manner, with a threshold of 10 nm in both normal and nerve-ligated synaptosomes. Blockade of capsaicin-evoked glutamate release by adenosine was reversed similarly in synaptosomes from normal and spinal nerve-ligated animals by an A1 adenosine receptor antagonist DPCPX (8-cyclopentyl-1,3-dipropylxanthine) but not by an A2 adenosine receptor antagonist DMPX (3'7-dimethyl-1-proparaglyxanthine). Capsaicin-evoked glutamate release, as well as its inhibition by adenosine, did not differ between synaptosomes prepared from tissue ipsilateral and contralateral to spinal nerve ligation. CONCLUSION: These observations confirm previous neurophysiologic studies that presynaptic adenosine A1 receptor activation inhibits glutamate release from primary afferents. This effect is unaltered after peripheral nerve injury and thereby is unlikely to account for the enhanced analgesic efficacy of intrathecal adenosine in this setting.     

Differential effects of ketamine enantiomers on anesthetic levels and glutamate release in the hippocampus

The anesthetic effects of ketamine enantiomers (40 and 100 mg.kg-1, i.p.) were evaluated in the rat by behavioral parameters such as the orienting response, the tail-pinch test and the spontaneous motor behavior including stereotyped behavior (circling and nodding). The intercellular glutamate concentration in the dorsal hippocampus was measured by brain microdialysis method. The analgesic and sedative effects of the S(+)-ketamine were more potent than those of the R(-)-ketamine at each dose. The incidence of stereotyped behaviors in R(-)-ketamine groups was higher than that of S(+)-ketamine groups. The basal release of glutamate under perfusion with normal artificial cerebrospinal fluid (ACSF) was unchanged after the administration of these enantiomers. Repeat perfusion with high-potassium ACSF (100 mM K+) for 20 min increased the glutamate content in vehicle-treated rats. This depolarization-related glutamate release was suppressed by ketamine, except in the 100 mg R(-)-ketamine group in which the stimulated glutamate release tended to increase after the treatment. The inhibitory effect on this stimulated glutamate release was marked in S(+)-ketamine groups. These results suggest that the anesthetic effects of ketamine enantiomers may be mediated at least in part, via regulation of depolarization-released glutamate release.     

脂肪乳剂减轻布比卡因抑制原代培养海马神经元活力

目的研究脂肪乳剂对布比卡因致原代培养海马神经元神经毒性的影响。方法选择原代培养第8天的海马神经元,采用四甲基偶氮唑蓝(MTT)法计算布比卡因对海马神经元的半抑制浓度(IC50),用接近IC50的3个浓度分别作用12h、24h和36h,MTT法检测神经元活力,观察布比卡因对海马神经元的神经毒性作用,得出最佳布比卡因浓度和作用时间进行后续实验。将原代海马神经元细胞分为四组(n=3):空白对照组(C组)不做任何处理;布比卡因组(B组)加入布比卡因;脂肪乳剂组(L组)加入1%脂肪乳剂;布比卡因+脂肪乳剂组(BL组)加入布比卡因和1%脂肪乳剂。各组均处理一定时间后光学显微镜观察海马神经元生长状态,检测各组神经元活力,免疫荧光染色法检测裂解半胱天冬酶-3(cleaved caspase-3)的表达并统计cleaved caspase-3阳性神经元占神经元总数的比例。结果布比卡因的IC50为0.033 62%,摩尔浓度为980.4μM。选择布比卡因浓度为1mmol/L和作用时间为24h进行后续实验。与C组比较,B组海马神经元细胞活力明显减弱(P0.01)。与B组比较,L组和BL组海马神经元细胞活力明显增强(P0.01)。与C组比较,B组和BL组cleaved caspase-3阳性率明显升高(P0.01),与B组比较,BL组神经元cleaved caspase-3阳性率明显降低(P0.01),C组和L组cleaved caspase-3阳性率差异无统计学意义。结论布比卡因诱发海马神经元神经毒性呈浓度剂量和时间依赖性,脂肪乳剂减轻布比卡因的神经毒性可能与增加海马神经元的细胞活力有关。     

Effect of fentanyl on ischemic depolarization and ischemic neuronal damage of hippocampal CA1 in the gerbil

Purpose  

Temporary brain ischemia occurring during surgery under general anesthesia may induce the death of neuronal cells and cause severe neurological deficits. On the other hand, it is not clear whether μ-opioid receptor agonists promote ischemic brain injury. It is known that duration of ischemic depolarization affects the degree of neuronal damage. However, the effects of fentanyl during brain ischemia on ischemic depolarization have not been investigated. Therefore, in the current study, the effects of fentanyl on ischemic neuronal damage and ischemic depolarization were quantitatively evaluated.     

Contribution of Ih to neuronal damage in the hippocampus after traumatic brain injury in rats

Traumatic brain injury (TBI) causes selective neuronal damage in the hippocampus; however, the underlying mechanisms are still unclear. Post-traumatic alterations of ion channel activity, which actively regulate neuronal excitability and thus impact on excitotoxicity, may be involved in TBI-induced neuronal injury. Here we report that hyperpolarization-activated cation current (I(h)) contributes to the distinct vulnerability of hippocampal neurons in TBI. In a rat model of controlled cortical injury, moderate TBI produced neuronal death of both hippocampal CA3 neurons and mossy cells in the hilus, but not CA1 pyramidal cells. Treatment with lamotrigine, which enhances dendritic I(h), ameliorated TBI-induced neuronal damage to CA3 neurons and mossy cells. In contrast, intraventricular administration of I(h) channel blocker caused cell death in the CA1 region after TBI. Whole-cell recordings revealed that, differently from CA3 neurons, CA1 pyramidal cells expressed larger I(h) and exhibited a post-traumatic increase of I(h) amplitude. Moreover, blocking I(h) led to an increase of neuronal excitability, with greater effects seen in post-traumatic CA1 pyramidal cells than in CA3 neurons. In addition, the I(h) in mossy cells was dramatically inhibited early after TBI. Our findings indicate that differential changes of I(h) in hippocampal neurons may be one of the mechanisms of selective cell death, and that an enhancement of functional I(h) may protect hippocampal neurons against TBI.     

Dizocilpine but not ketamine reduces the volume of ischaemic damage after acute subdural haematoma in the rat

BACKGROUND: AND OBJECTIVE: Increased glutamate concentration in the cerebrospinal fluid has been reported in severely head-injured patients, suggesting that an excessive release of glutamate may be involved in the process of neuronal damage. Ischaemic damage after subdural haematoma has been reported to be reduced by glutamate (N-methyl-D-aspartate: NMDA) receptor antagonists such as dizocilpine and CGS 19755; even though these drugs were given 20-30 min after insult. Excessive release of excitatory amino acids may produce the neural damage after subdural haematoma and NMDA receptor antagonists may become valuable therapeutic drugs. This study compared the effects of ketamine and dizocilpine, on intracranial pressure and histopathological changes after acute subdural haematoma produced by an injection of autologous blood (150 microL) in rats. METHODS: The control (n = 9), ketamine (n = 9) and dizocilpine (n = 9) groups, respectively, received saline, ketamine (total dose: 210 mg kg-1) or dizocilpine (total dose: 1.0 mg kg-1) from 0.5 to 8 h after acute subdural haematoma. A silicone group (n = 9) had the same volume of silicone injected subdurally. RESULTS: The volume of ischaemic damage in the silicone group (1.3 +/- 1.2 mm3) was significantly smaller than in the control group (11.9 +/- 3.8 mm3). Ketamine and dizocilpine did not increase intracranial pressure. Dizocilpine significantly decreased the volume of ischaemic damage (6.1 +/- 3.8 mm3). Ketamine failed to significantly decrease damage (7.8 +/- 5.0 mm3). CONCLUSIONS: These results suggest that the factors elicited by the clotted blood contribute to the ischaemic damage after subdural haematoma, and that the glutamate receptor antagonist dizocilpine reduces the damage, while ketamine shows only a trend reduction of the damage.     

Aminoacid incorporation into polyribosomes of ischaemic and reperfused gerbil brain

Summary Aminoacid incorporation into polyribosomes from the gerbil brain in a cell-free system is strongly affected by experimental ischaemia. Progression of the impairment of the protein synthesis in vitro occurs even following early reperfusion.     

Ischemic neuronal damage after acute subdural hematoma in the rat: effects of pretreatment with a glutamate antagonist

The ability of a competitive N-methyl-D-aspartate (NMDA) receptor antagonist (D-CPP-ene) to reduce irreversible brain damage has been examined in a rodent model of acute subdural hematoma. Acute subdural hematoma was produced by the slow injection of 400 microliters homologous blood into the subdural space overlying the parietal cortex in halothane-anesthetized rats. Brain damage was assessed histologically in sections at multiple coronal planes in animals sacrificed 4 hours after induction of the subdural hematoma. Pretreatment with D-CPP-ene (15 mg/kg) significantly reduced the volume of ischemic brain damage produced by the subdural hematoma from 62 +/- 8 cu mm (mean +/- standard error of the mean) in vehicle-treated control rats to 29 +/- 7 cu mm in drug-treated animals. These data demonstrate the anti-ischemic efficacy of NMDA antagonists in an animal model of intracranial hemorrhage in which intracranial pressure is elevated, and suggest that excitotoxic mechanisms (which are susceptible to antagonism by D-CPP-ene) may play a role in the ischemic brain damage which is observed in patients who die after acute subdural hematoma.     

异丙酚对异相睡眠剥夺大鼠海马谷氨酸和γ-氨基丁酸释放的影响

目的 探讨异丙酚对24 h异相睡眠剥夺大鼠海马谷氨酸(Glu)和γ-氨基丁酸(γ-GABA)释放的影响.方法 健康雄性SD大鼠16只,成功完成24 h异相睡眠剥夺实验后,随机分为2组(n=8),自然睡眠组(N组)腹腔注射5%脂肪乳剂3 ml,异丙酚组(P组)腹腔注射异丙酚100mg/kg.于异相睡眠剥夺前(基础状态)、异相睡眠剥夺24 h时(T1)、异相睡眠剥夺结束后1 h(T2)、3 h(T3)和6 h(T4)收集2组海马微透析液,检测Glu和γ-GABA的浓度.结果 与基础值比较,N组和P组T1-3时海马微透析液内Glu、γ-GABA浓度升高,N组T4时上述指标仍较高;与T1时比较,N组和P组T3,4时上述指标降低(P＜0.05或0.01).两组间Glu、γ-GABA浓度差异无统计学意义(P＞0.05).结论 异丙酚麻醉对大鼠异相睡眠剥夺时海马内紊乱的Glu和γ-GABA递质有恢复作用,其作用与自然睡眠相似.     

NAAG peptidase inhibitor reduces acute neuronal degeneration and astrocyte damage following lateral fluid percussion TBI in rats

Traumatic brain injury (TBI) produces a rapid and excessive elevation in extracellular glutamate associated with excitotoxicity and secondary brain pathology. The peptide neurotransmitter Nacetylaspartylglutamate (NAAG) suppresses glutamate transmission through selective activation of presynaptic Group II metabotropic glutamate receptor subtype 3 (mGluR3). Thus, inhibition of NAAG peptidase activity and the prolong presence of synaptic NAAG were hypothesized to have significant potential for cellular protection following TBI. In the present study, a novel NAAG peptidase inhibitor, ZJ-43, was used in four different doses (0, 50, 100, or 150 mg/kg). Each dose was repeatedly administered i.p. (n=5/group) by multiple injections at three times (0 time, 8 h, 16 h) after moderate lateral fluid percussion TBI in the rat. An additional group was co-administered ZJ-43 (150 mg/kg) and the Group II mGluR antagonist, LY341495 (1 mg/kg), which was predicted to abolish any protective effects of ZJ-43. Rats were euthanized at 24 h after TBI, and brains were processed with a selective marker for degenerating neurons (Fluoro-Jade B) and a marker for astrocytes (GFAP). Ipsilateral neuronal degeneration and bilateral astrocyte loss in the CA2/3 regions of the hippocampus were quantified using stereological techniques. Compared with vehicle, ZJ-43 significantly reduced the number of the ipsilateral degenerating neurons (p<0.01) with the greatest neuroprotection at the 50 mg/kg dose. Moreover, LY341495 successfully abolished the protective effects of ZJ-43. 50 mg/kg of ZJ-43 also significantly reduced the ipsilateral astrocyte loss (p<0.05). We conclude that the NAAG peptidase inhibitor ZJ-43 is a potential novel strategy to reduce both neuronal and astrocyte damage associated with the glutamate excitotoxicity after TBI.     

Post-traumatic hypoxia exacerbates neuronal cell death in the hippocampus

Hypoxia frequently occurs in patients with traumatic brain injury (TBI) and is associated with increased morbidity and mortality. This study examined the effects of immediate or delayed post-traumatic hypoxia (fraction of inspired oxygen [FiO(2)] 11%) on acute neuronal degeneration and long-term neuronal survival in hippocampal fields after moderate fluid percussion injury in rats. In Experiment 1, hypoxia was induced for 15 or 30?min alone or immediately following TBI. In Experiments 2 and 3, 30?min of hypoxia was induced immediately after TBI or delayed until 60?min after TBI. In Experiment 1, acute neurodegeneration was evaluated in the hippocampal fields 24?h after insults using Fluoro-Jade staining and stereological quantification. During hypoxia alone, or in combination with TBI, mean arterial blood pressure was significantly reduced by approximately 30%, followed by a rapid return to normal values upon return to pre-injury FiO(2). Hypoxia alone failed to cause hippocampal neuronal degeneration when measured at 24?h after insult. TBI alone resulted in neuronal degeneration in each ipsilateral hippocampal field, predominantly in CA2-CA3 and the dentate gyrus. Compared to TBI alone, TBI plus immediate hypoxia for either 15 or 30?min significantly increased neuronal loss in most ipsilateral hippocampal fields and in the contralateral hilus and dentate gyrus. In Experiment 2, TBI plus hypoxia delayed 30?min significantly increased degeneration only in ipsilateral CA2-CA3. In Experiment 3, 30?min of immediate hypoxia significantly reduced the numbers of surviving neurons in the CA3 at 14 days after TBI. The greatly increased vulnerability in all hippocampal fields by immediate 30?min post-traumatic hypoxia provides a relevant model of TBI complicated with hypoxia/hypotension. These data underscore the significance of the secondary insult, the necessity to better characterize the range of injuries experienced by the TBI patient, and the importance of strictly avoiding hypoxia in the early management of TBI patients.     

Tissue pH in the assessment of ischaemic muscle damage.

Measurement in vivo of the pH of superficial muscles of dogs was found suitable for the assessment of the severity of ischaemic muscle damage and for the detection of irreversible lesions.     

静脉注射普鲁卡因对脊髓反射的影响

目的 研究静注射普鲁卡因对脊髓反射的影响,旨在探讨普鲁卡因注射产生镇痛是否发生在脊髓水平。方法 利用猫处周伤害性刺激能够引起的屈肌反射产生易化的模型,观察静脉注射普鲁卡因５和１０ｍｇ／ｋｇ对屈肌反射及屈肌反射及屈肌反射易化的影响。结果 ５ｍｇ／ｋｇ组屈肌反射及屈肌反射易化的抑制率分别为２０．５％（Ｐ〈０．０１,ｎ＝１２）和５３．３％（Ｐ〈０．０１,ｎ＝１２）,１０ｍｇ／ｋｇ组抑制率分别为５０．６％