Endogenous Activation of adenosine A1 receptors promotes post-ischemic electrocortical burst suppression

Following transient global cerebral ischemia (GCI), spontaneous electrocortical activity resumes from the isoelectric line through a sequence of “bursts” of activity alternating with periods of electrical “suppression,” commonly referred to as the post-ischemic burst suppression (BS) pattern. Several lines of evidence suggest that BS reflects an impairment of neocortical connectivity. Here we tested in vivo whether synaptic depression by adenosine A₁ receptor (A₁R) activation contributes to BS patterns following GCI. Male Wistar rats were subjected to 1, 5 or 10 min of GCI using a “four-vessel occlusion” model under chloral hydrate anesthesia. Quantification of BS recovery was carried out using BS ratio. During GCI full electrocortical suppression was attained (BS ratio reached 100%). During the following reperfusion the BS ratio returned to 0. The time course of the decay was exponential after 1 and 5-min GCI and bi-exponential after 10-min GCI. The BS recovery was progressively delayed with the duration of ischemia. Administration of the A₁R antagonist 1,3-dipropyl-8-cyclopentylxanthine (DPCPX, 1.25 mg/kg i.p.) accelerated the post-ischemic BS recovery for all GCI durations. Following the 10-min GCI the effect of DPCPX was only apparent on the initial fast decay of the BS ratio. These data suggest that endogenous adenosine release promotes BS patterns during reperfusion following transient cerebral ischemia. Furthermore, the endogenous A₁R activation may be the primary underlying cause of post-ischemic BS patterns following brief ischemic episodes. It is likely that synaptic depression by post-ischemic A₁R activation functionally disrupts the connectivity within the cortical networks to an extent that promotes BS patterns.     

Rapid tolerance to focal cerebral ischemia in rats is induced by preconditioning with electroacupuncture: window of protection and the role of adenosine

The aim of the present study was to investigate the first protective window of preconditioning with electroacupuncture (EA) against focal cerebral ischemia, and to explore whether adenosine is involved in the rapid tolerance phenomenon. Sixty-four male Sprague-Dawley rats were randomly assigned to eight groups (n=8 in each). Animals in the control group received no treatment, and animals in EA1-EA4 groups received EA at 0.5, 1, 2 and 3 h before induction of focal cerebral ischemia, respectively. Rats in DPCPX group were intraperitoneally injected with 1 mg kg-1 8-cyclopentyl-1,3-dipropylxanthine (DPCPX), 3 h before induction of focal cerebral ischemia. Animals in vehicle group and EA+DPCPX group were pretreated with 1 ml kg-1 dimethyl sulfoxide (DMSO, the solvent of DPCPX) and 1 mg kg-1 DPCPX 30 min before preconditioning with EA, respectively. All rats were anesthetized with 40 mg kg-1 pentobarbital sodium intraperitoneally. Animals that required EA preconditioning, received EA with intensity of 1 mA and frequency of 15 Hz at the Baihui acupoint (GV 20) for 30 min. The focal cerebral ischemia was produced by the right middle cerebral artery occlusion (MCAO) for 120 min. The neurologic deficit scores (NDS) and brain infarct volumes were evaluated at 24 h after reperfusion. All rats survived until 24 h after reperfusion. Preconditioning with EA at 2 h before induction of focal cerebral ischemia improved neurologic outcome (P<0.05 versus control) and reduced the infarct volume (P<0.01 versus control) at 24 h after reperfusion. These beneficial effects were reversed by pretreatment with 1 mg kg-1 DPCPX, whereas this agent itself did not affect the NDS and volume in drug-ischemic controls after ischemia. The results show that preconditioning with single EA session induces rapid tolerance to focal cerebral ischemia. The rapid ischemic tolerance appears at 2 h (but not at 0.5, 1, or 3 h) after preconditioning, and is possibly mediated through an adenosine A1 receptor-related mechanism.     

The effect of biodegradable gelatin microspheres on the neuroprotective effects of high mobility group box 1 A box in the postischemic brain

High mobility group box 1 (HMGB1) is a family of endogenous molecules that is released by necrotic cells and causes neuronal damages by triggering inflammatory processes. In the cerebral ischemic brain, sustained and regulated suppression of HMGB1 has been emerged as a therapeutic means to grant neuroprotection. HMGB1 consists of two HMG boxes (A and B) and an acidic C-terminal tail, and the A box peptide antagonistically competes with HMGB1 for its receptors. In the middle cerebral artery occlusion (MCAO) in rats, a murine model of transient cerebral ischemia, administration of HMGB1 A box intraparenchymally, after encapsulated in biodegradable gelatin microspheres (GMS), which enhances the stability of peptide inside and allows its sustained delivery, at 1 h, 3 h, or 6 h after MCAO, reduced mean infarct volumes by, respectively, 81.3%, 42.6% and 30.7% of the untreated MCAO-brain, along with remarkable improvement of neurological deficits. Furthermore, the administration of HMGB1 A box/GMS suppressed proinflammatory cytokine inductions more strongly than the injection of non-encapsulated HMGB1 A box. Given that insulted brains-like ischemia have enhanced gelatinase activity than the normal brain, our results suggest that GMS-mediated delivery of therapeutic peptides is a promising means to provide efficient neuroprotection in the postischemic brain.     

Infiltration and persistence of lymphocytes during late-stage cerebral ischemia in middle cerebral artery occlusion and photothrombotic stroke models

Background

Evidence suggests that brain infiltration of lymphocytes contributes to acute neural injury after cerebral ischemia. However, the spatio-temporal dynamics of brain-infiltrating lymphocytes during the late stage after cerebral ischemia remains unclear.

Methods

C57BL/6 (B6) mice were subjected to sham, photothrombosis, or 60-min transient middle cerebral artery occlusion (MCAO) procedures. Infarct volume, neurodeficits, production of reactive oxygen species (ROS) and inflammatory factors, brain-infiltrating lymphocytes, and their activation as well as pro-inflammatory cytokine IFN-γ production were assessed. Brain-infiltrating lymphocytes were also measured in tissue sections from post-mortem patients after ischemic stroke by immunostaining.

Results

In mice subjected to transient MCAO or photothrombotic stroke, we found that lymphocyte infiltration persists in the ischemic brain until at least day 14 after surgery, during which brain infarct volume significantly diminished. These brain-infiltrating lymphocytes express activation marker CD69 and produce proinflammatory cytokines such as IFN-γ, accompanied with a sustained increase of reactive oxygen species (ROS) and inflammatory cytokines release in the brain. In addition, brain-infiltrating lymphocytes were observed in post-mortem brain sections from patients during the late stage of ischemic stroke.

Conclusion

Our results demonstrate that brain-infiltration of lymphocytes persists after the acute stage of cerebral ischemia, facilitating future advanced studies to reveal the precise role of lymphocytes during late stage of stroke.

     

Preconditioning with +Gz acceleration (head-to-foot inertial load) produces neuroprotection against transient focal cerebral ischemia in rats

Ischemic preconditioning is considered to be the most robust endogenous neuroprotectant. However, the conventional ischemic preconditioning protocol is both invasive and impractical to apply. The aim of the present study was to evaluate whether preconditioning with +Gz centrifuge acceleration (head-to-foot inertial load) which could induce brief episodes of sublethal ischemia in brain had neuroprotection against focal cerebral ischemic injury. A total of 85 male Sprague-Dawley rats were randomly assigned to five groups (n = 17 in each). The 2 Gz, 4 Gz, 6 Gz and 8 Gz groups were subjected to 3 min exposures at +2 Gz, +4 Gz, +6 Gz and +8 Gz, respectively for consecutive three times in animal centrifuge, with a 30-min rest period between each centrifuge run. The control group had no exposure to +Gz acceleration. Twenty-four hours after the last pretreatment, 12 rats in each groups were subjected to focal cerebral ischemia for 120 min and the other five rats in each group were sacrificed to measure the expression of heat shock protein 70(HSP70) in hippocampus by Western blot analysis. The results indicated that the 6 Gz and 8 Gz groups showed smaller infarct volume and lower neurologic deficit scores than the control group. The expression of HSP70 was significantly increased in 6 Gz and 8 Gz groups than those in the control group. Therefore, preconditioning with +Gz acceleration produced delayed neuroprotection against focal cerebral ischemia and that the neuroprotection may be related to the induction of HSP70.     

Spatio-temporal expression of cysteinyl leukotriene receptor-2 mRNA in rat brain after focal cerebral ischemia

Cysteinyl leukotrienes (CysLTs) induce inflammatory responses mediated by activating CysLT(1) and CysLT(2) receptors. We have recently reported that CysLT(1) receptor expression is increased in rat brain after focal cerebral ischemia and the increased expression is spatio-temporally related to acute neuronal injury and late astrocyte proliferation. Here we report spatio-temporal expression of CysLT(2) receptor mRNA in rat brain after focal cerebral ischemia induced by 30min of middle cerebral artery occlusion. We found that the neuron density was gradually decreased or disappeared in the ischemic core and boundary zone during 14 days after reperfusion, and the astrocyte population in the boundary zone was increased 3-14 days after reperfusion. In the ischemic core, the expression of CysLT(2) receptor mRNA was increased at 6, 12 and 24h and then recovered at 3, 7 and 14 days after reperfusion. In the boundary zone, the expression was significantly increased 3, 7 and 14 days after reperfusion. The results suggest that CysLT(2) receptor may be related to the acute neuronal injury and late astrocyte proliferation in the ischemic brain.     

The selective A2 adenosine receptor agonist CGS 21680 enhances excitatory transmitter amino acid release from the ischemic rat cerebral cortex.

The effects of the selective adenosine A2 receptor agonist 2-p-(2-carboxyethyl)phenethylamino-5'-N-ethylcarboxamido adenosine hydrochloride (CGS 21680) on aspartate and glutamate release from the ischemic rat cerebral cortex were studied with the cortical cup technique. Cerebral ischemia (20 min) was elicited by four vessel occlusion. Pretreatment with CGS 21680 failed to alter basal excitatory amino acid levels, however, CGS 21680 at 10(-6) M significantly enhanced the ischemia-evoked release. Thus, aspartate and glutamate release during ischemia can be stimulated via the activation of A2 receptors, in addition to the suppression of excitatory amino acid release mediated by selective A1 receptor agonists.     

Bone Morphogenic Protein-7 Contributes to Cerebral Ischemic Preconditioning Induced-Ischemic Tolerance by Activating p38 Mitogen-Activated Protein Kinase Signaling Pathway

Cerebral ischemic preconditioning (IPC), which refers to a transient and noninjurious ischemia is able to induce tolerance against the subsequent lethal ischemia, including ischemic stroke. We have previously reported that bone morphogenic protein-7 (BMP-7) contributes to the neuroprotective effects of IPC-induced ischemic tolerance, and thus ameliorates the following ischemia/reperfusion (I/R) injury in rats. Consequently, in the present study, we continued to explore the underlying regulatory mechanisms involved in BMP-7-mediated cerebral IPC in the rat model of ischemic tolerance. Male Wistar rats were preconditioned by 15-min middle cerebral artery occlusion (MCAO). After 2-day reperfusion, these animals were subjected to prolonged MCAO for 2 h. Our results showed that the phosphorylated p38 mitogen-activated protein kinase (MAPK) paralleling to BMP-7 was up-regulated by IPC in rat brain. Inactivation of p38 MAPK by pretreatment of SB203580, a p38 MAPK-specific suppressor, weakened the protective effect of IPC on CA1 neurons. Moreover, the enhanced phosphorylation of p38 MAPK induced by IPC was attenuated when the endogenous BMP-7 was inhibited by BMP-7 antagonist noggin. Besides, blockade of p38 MAPK signal transduction pathway via SB203580 abrogated the protective effects of exogenous BMP-7 against cerebral infraction. These present findings suggest that BMP-7 contributes to cerebral IPC-induced ischemic tolerance via activating p38 MAPK signaling pathway.     

缺血再灌注大鼠脑内促红细胞生成素表达的变化

探讨缺血再灌注大鼠脑内促红细胞生成素（erythropoietin,Epo）表达的变化,揭示短暂缺血时神经系统发生内源性脑保护的机制。采用线栓法制作大鼠局灶性缺血再灌注模型,以免疫组织化学和逆转录聚合酶链式反应（PT-PCR）技术,检测缺血再灌注不同时间脑内Epo的表达变化。脑缺血再灌注不同时间Epo蛋白在脑内表达广泛,主要颁布在缺血侧基底节区、海马和部分皮层。在缺血再灌注1h和6h,基底节区Epo蛋白和mRNA表达较高,再灌注12h,基底节区Epo表达减少,额顶皮质Epo表达增加,再灌注24h,额顶皮质Epo表达达到高峰,48h开始下调。因此,促红细胞生成素在缺血再灌注大鼠脑内表达的变化,可能是机体发生内源性脑保护的机制之一。     

Increase of adenosine A1 receptor gene expression in cerebral ischemia of Wistar rats

In an attempt to know the role of adenosine A1 receptor in cerebral ischemia, the present study employed the ligation of bilateral carotid arteries to induce ischemia in Wistar rats. Changes of gene expression of adenosine A1 receptor in cerebral cortex of ischemic rats were compared with normal sham control and reperfusion group that received regular blood flow after a transient ischemia. The mRNA level of adenosine A1 receptor in cerebral cortex was markedly raised by this artificial ischemia. Also, reperfusion reversed this elevation to a level near the control. This change was also observed at the protein level using Western blot analysis of adenosine A1 receptor. The raised protein level of adenosine A1 receptor by ischemia was reversed to normal level after reperfusion. These data suggest that the gene expression of adenosine A1 receptor was increased by ischemia probably due to the compensative response of brain. The raised adenosine A1 receptor may play a protective role in these damaged tissues.     

Excitatory synaptic transmission in the spinal substantia gelatinosa is under an inhibitory tone of endogenous adenosine

Exogenous adenosine produces potent synaptic inhibition in spinal substantia gelatinosa (SG), a region involved in nociceptive and thermoreceptive mechanisms. To examine the possibility that endogenous adenosine tonically modulates excitatory synaptic transmission in spinal SG, whole-cell, voltage-clamp recordings were made from SG neurons in adult rat spinal cord slices. In all SG neurons sensitive to exogenous adenosine, the adenosine uptake inhibitor, NBTI, mimics adenosine's inhibitory actions on dorsal root evoked EPSCs (eEPSCs) and miniature spontaneous EPSCs (mEPSCs). These inhibitory effects were antagonized by A1 adenosine receptor antagonist, DPCPX. DPCPX also potentates eEPSCs in those SG neurons in which adenosine or adenosine A1 receptor agonists (CHA, CCPA) suppressed eEPSCs. DPCPX often increases mEPSC frequency without altering mEPSC amplitude, suggesting presynaptic action on adenosine A1 receptors. Selective A2 (DMPX) and A2a (ZM 241385) adenosine receptor antagonists had no or minimal effects upon either eEPSCs or mEPSCs. The adenosine degrading enzyme, adenosine deaminase, mimicked the effects of DPCPX on the mEPSC frequency. We conclude that the excitatory synaptic transmission in the spinal SG is under an inhibitory tone of endogenous adenosine through the activation of A1 receptors. The present results suggested that the background activity of A1 receptors in the spinal SG might be contributed to setting the physiological “noceceptive thresholds”.     

Regulatory effect of Dimethyl Sulfoxide (DMSO) on astrocytic reactivity in a murine model of cerebral infarction by arterial embolization

Introduction:

The pathophysiology of cerebral ischemia is essential for early diagnosis, neurologic recovery, the early onset of drug treatment and the prognosis of ischemic events. Experimental models of cerebral ischemia can be used to evaluate the cellular response phenomena and possible neurological protection by drugs.

Objective:

To characterize the cellular changes in the neuronal population and astrocytic response by the effect of Dimethyl Sulfoxide (DMSO) on a model of ischemia caused by cerebral embolism.

Methods:

Twenty Wistar rats were divided into four groups (n= 5). The infarct was induced with α-bovine thrombin (40 NIH/Unit.). The treated group received 90 mg (100 μL) of DMSO in saline (1:1 v/v) intraperitoneally for 5 days; ischemic controls received only NaCl (placebo) and two non-ischemic groups (simulated) received NaCl and DMSO respectively. We evaluated the neuronal (anti-NeuN) and astrocytic immune-reactivity (anti-GFAP). The results were analyzed by densitometry (NIH Image J-Fiji 1.45 software) and analysis of variance (ANOVA) with the Graph pad software (Prism 5).

Results:

Cerebral embolism induced reproducible and reliable lesions in the cortex and hippocampus (CA1)., similar to those of focal models. DMSO did not reverse the loss of post-ischemia neuronal immune-reactivity, but prevented the morphological damage of neurons, and significantly reduced astrocytic hyperactivity in the somato-sensory cortex and CA1 (p <0.001).

Conclusions:

The regulatory effect of DMSO on astrocyte hyperreactivity and neuronal-astroglial cytoarchitecture , gives it potential neuroprotective properties for the treatment of thromboembolic cerebral ischemia in the acute phase.     

Angiotensin type 1 receptor blockage improves ischemic injury following transient focal cerebral ischemia

Following cerebral ischemia, i.v. infusion of angiotensin II increases cerebral edema and mortality. Angiotensin type 1 receptor blockage should therefore improve acute cerebral ischemia. Left middle cerebral artery occlusion (120 min) followed by reperfusion was performed with the thread method under halothane anesthesia in Sprague-Dawley rats. Olmesartan (angiotensin type 1 receptor blocker; 0.01 or 0.1mumol/kg/h) was infused i.p. for 7 days following middle cerebral artery occlusion followed by reperfusion. Stroke index score, infarct volume, specific gravity, and brain angiotensin II and matrix metalloproteinases were quantified in the ischemic and non-ischemic hemispheres. Olmesartan treatment improved stroke index score, infarct volume, and cerebral edema in our cerebral ischemia model. In particular, stroke index score, infarct volume, and cerebral edema were reduced even with a low dose of olmesartan that did not decrease blood pressure. Paralleling these effects on cerebral ischemia, olmesartan treatment also reduced the reactive upregulation in brain angiotensin II, matrix metalloproteinase-2, matrix metalloproteinase-9, and membrane type 1-matrix metalloproteinase in the ischemic area. Angiotensin type 1 receptor stimulation may be one of the important factors that cause cerebral edema following cerebral ischemia, and that its inhibition may be of therapeutic advantage in cerebral ischemia.     

Activation of liver X receptor reduces global ischemic brain injury by reduction of nuclear factor-κB

Recent studies have found that liver X receptors (LXRs) agonists decrease brain inflammation and exert neuroprotective effect. The aim of this study was to examine the mechanisms of action of liver X receptor agonist GW3965 against brain injury following global cerebral ischemia in the rat. The 48 male SD (Sprague–Dawley) rats were randomly partitioned into three groups: sham, global ischemia (4-vessel occlusion for 15 min; 4VO) treated with vehicle and global ischemia treated with GW3965 (20 mg/kg, via i.p. injection at 10 min after reperfusion). The functional outcome was determined by neurological evaluation at 24 h post ischemia and by testing rats in T maze at 3 and 7 days after reperfusion. The rats' daily body weight, incidence of seizures and 72 h mortality were also determined. After Nissl staining and TUNEL in coronal brain sections, the numbers of intact and damaged cells were counted in the CA1 sector of the hippocampus. The expression of phosphorylated inhibitor of κB (p-IκBα), nuclear factor-κB (NF-κB) subunit p65, and cyclo-oxygenase-2 (COX-2) were analyzed with Western blot at 12 h after reperfusion. GW3965 tended to reduce 72 h mortality and the incidence of post-ischemic seizures. GW3965-treated rats showed an improved neuronal survivability in CA1 and a significant increase in the percentage of spontaneous alternations detected in T-maze on day 7 after ischemia. GW3965-induced neuroprotection was associated with a significant reduction in nuclear translocation of NF-kB p65 subunit and a decrease in the hippocampal expression of NF-kB target gene, COX-2. LXR receptor agonist protects against neuronal damage following global cerebral ischemia. The mechanism of neuroprotection may include blockade of NF-κB activation and the subsequent suppression of COX-2 in the post ischemic brain.     

Preconditioning of the Small Intestine to Ischemia in Rats

Measures reflecting the state of the small intestine were studied in rats after ischemia lasting 90 min produced by clamping the superior mesenteric artery and reperfusion for 30 min. Preconditioning of the intestine to ischemia was induced by producing intestinal ischemia for 10 min followed by 10 min of reperfusion (ischemic preconditioning), 30-min limb ischemia with 15-min reperfusion (distant ischemic preconditioning), and i.v. L-arginine. The smallest amount of damage to the intestine after 90 min of ischemia and 30 min of reperfusion was seen in the group of rats subjected to ischemic preconditioning. The protective effect of ischemic preconditioning was partially blocked by administration of N--nitro-L-arginine (a blocker of NO synthesis). Doses of L-arginine also had protective effects, though these were smaller than those of ischemic preconditioning. Preliminary ischemia-reperfusion of the limb had no effect on the state of the intestine. Thus: 1) ischemic preconditioning of the intestine is partially associated with activation of nitric oxide synthesis, and 2) distant ischemic preconditioning did not protect the intestine.     

Nimodipine improves the disruption of spatial cognition induced by cerebral ischemia

The direct neuroprotective effect of nimodipine, a central Ca antagonist, was investigated in in vitro experiments. Also, in in vivo experiments, the effects of nimodipine and amlodipine, a noncentral Ca antagonist, on rat cerebral ischemia models developing by different mechanisms were compared. In an in vitro ischemic model using acidotic and hypoglycemic rat cerebellar granule cells, nimodipine directly protects against brain neuronal cell damage. In in vivo models of single (one 10-min, four-vessel occlusion) and repeated rat cerebral ischemia (two 10-min, four-vessel occlusions; a 50-min interval), the impairment observed 24 h after the single ischemic procedure was likely to be prevented by nimodipine (0. 1-5mg/kg, i.p.). At 7 days after the repeated cerebral ischemia, the disruption of spatial cognition was significantly prevented by nimodipine (5 mg/kg, i.p.) but not amlodipine (5 mg/kg, i.p.), which was given after each ischemia. These results indicated that nimodipine may protect neuronal cells by a more persistent mode of action, that is, nimodipine may enter into the cell and control the intracellular Ca ion cascade by inhibiting excessive Ca(2)+ influx into the mitochondria.     

人参皂甙Rd在脑缺血亚急性期的脑保护效应

目的:探讨人参皂甙Rd(Ginsenoside Rd,GSRd)在脑缺血亚急性期的脑保护效应。方法:84只雄性SD大鼠随机分为四组,其中假手术+丙二醇(propylene glycol,PG)组和假手术+GSRd组各18只,脑缺血+PG组和脑缺血+GSRd组各24只。线栓法建立局灶性短暂性大脑中动脉栓塞模型(focal transient middle cerebral ar-tery occlusion,MCAO),栓塞后1 d经腹腔注射GSRd和PG。在用药前、用药后3 d和7 d分别进行神经功能评分(neurological behavior scores,NBS),每次评分后取脑进行2,3,5-氯化三苯基四氮唑(TTC)染色,用ImageJ图像分析软件计算出梗死体积百分比,并且对缺血区或半暗带进行Iba 1染色,观察小胶质细胞的变化情况以确定GSRd对脑缺血后炎性反应的影响作用。结果:脑缺血后1 d开始给予GSRd能改善MCAO模型大鼠的神经功能评分,减少脑梗死体积减弱小胶质细胞的活化,与PG组相比具有统计学差异(P<0.05)。结论:在脑缺血亚急性期给予GSRd能够起到脑保护作用。     

NAD(+) administration decreases ischemic brain damage partially by blocking autophagy in a mouse model of brain ischemia

Nicotinamide adenine dinuleotide (NAD(+)) plays critical roles in multiple biological functions. Previous studies have indicated that NAD(+) treatment decreases oxidative stress-induced death of primary neurons and astrocytes. Intranasal administration of NAD(+) also reduces brain damage in a rat model of transient focal brain ischemia. However, the mechanisms underlying this protective effect remain unknown. In this study, we used a mouse model of brain ischemia to test our hypothesis that NAD(+) decreases ischemic brain damage partially by preventing autophagy. Adult male mice were subjected to transient middle cerebral artery occlusion (tMCAO) for 90min, and NAD(+) was administered intraperitoneally (i.p.) immediately after reperfusion started. We found that administration with 50mg/kg NAD(+) led to significant decreases in infarct size, edema formation, and neurological deficits at 48h after ischemia. NAD(+) administration also significantly decreased brain ischemia-induced autophagy in the cortex and hippocampus. We further found that prevention of autophagy by 3-methyladenine (3-MA), a selective autophagy inhibitor, significantly reduced ischemic brain damage, suggesting an important role of autophagy in the ischemic brain injury in our animal model. Collectively, our findings have suggested that NAD(+) administration decreases ischemic brain damage at least partially by blocking autophagy. This is the first suggested mechanism regarding the protective effects of NAD(+) in cerebral ischemia, which further highlights the promise of NAD(+) for treating brain ischemia.     

缓激肽选择性增加局部脑缺血大鼠血脑屏障的通透性

目的　研究颈动脉灌注小剂量缓激肽对缺血后血脑屏障通透性的影响及机制。方法　免疫组化分析正常脑组织的缓激肽B2受体所在。大鼠大脑中动脉结扎 1h或 2h再灌流 1h。用放射自显影方法检测缓激肽对血脑屏障通透性的变化。WesternBlot方法检测bNOS ,iNOS和B2受体。NOS检测盒检测NOS的活性。结果　正常脑组织毛细血管内皮未见B2受体表达 ,在神经细胞上发现B2受体的表达。缺血 2h再灌流1h缓激肽灌注缺血区血脑屏障通透性显著增加。WesternBlot结果提示 ,在缓激肽灌注组和对照组间 ,缺血皮质区bNOS和B2受体没有明显变化 ,各组中均未检测出iNOS。缓激肽灌注组的NOS活性显著高于对照组。结论　正常脑组织毛细血管内皮未表达B2受体 ,神经细胞上可见B2受体的表达。灌注小剂量缓激肽能选择性增加局部脑缺血大鼠血脑屏障的通透性