Temperature modulation of ischemic neuronal death and inhibition of calcium/calmodulin-dependent protein kinase II in gerbils

We used brief bilateral carotid artery occlusion in gerbils to examine the effects of temperature on ischemia-induced inhibition of calcium/calmodulin-dependent protein kinase II activity and neuronal death. In normothermic (36 degrees C) gerbils, ischemia induced a severe loss of hippocampal CA1 pyramidal neurons measured 7 days after ischemia (28.4 neurons/mm, n = 10; control density in 10 naive gerbils 262.1 neurons/mm) and a significant decrease in forebrain calcium/calmodulin-dependent protein kinase II autophosphorylation measured 2 hours after ischemia (12.9 fmol/min, n = 6; control phosphorylation in six naive gerbils 23.5 fmol/min). The effect of temperature on these indicators of ischemic damage was examined by adjusting intracerebral temperature before and during the ischemic insult. Hyperthermic (39 degrees C) gerbils showed almost complete loss of neurons in the CA1 region (3.0 neurons/mm, n = 11) and extension of neuronal death into the CA2, CA3, and CA4 regions. In addition, hyperthermia exacerbated ischemia-induced inhibition of calcium/calmodulin-dependent protein kinase II activity (4.2 fmol/min, n = 6). Hypothermia (32 degrees C) protected against ischemia-induced CA1 pyramidal cell damage (257.0 neurons/mm, n = 20) and inhibition of calcium/calmodulin-dependent protein kinase II activity (26.0 fmol/min, n = 6). Our results are consistent with the hypothesis that loss of calcium/calmodulin-dependent protein kinase II activity may be a critical event in the development of ischemia-induced cell death.     

Keratinocyte growth factor prevents ischemia-induced delayed neuronal death in the hippocampal CA1 field of the gerbil brain

Fibroblast growth factors (FGFs) are polypeptides with various biological activities in vivo and in vitro, and their receptors are expressed in the widespread and specific neuronal populations of the brain. In this study, we asked whether keratinocyte growth factor (KGF), one of the FGF superfamily, would express in the brain, and have neuroprotective against ischemic brain injury. In situ hybridization analysis revealed that intense silver grains for KGF mRNA are observed in the neuronal cells of the cerebral cortex, hippocampus and amygdala in gerbil brain. Continuous cerebroventricular infusion of KGF (20 microg) for a 7 day period to gerbils starting 2 days before temporary right carotid artery occlusion (20 min) resulted in a higher survival rate than seen in vehicle-treated ischemic animals. Subsequent histological examinations showed that KGF effectively prevented delayed neuronal death of the hippocampal CA1 region. In situ detection of DNA fragmentation (TUNEL staining) revealed that ischemic animals infused with KGF contained fewer TUNEL-positive neurons in the hippocampal CA1 field than those infused with vehicle alone at the forth and seventh day after ischemia. KGF-treated brain showed over-expression of KGF mRNA in the neuronal cells of the cerebral cortex, hippocampus only in the right hemisphere, which was the side of carotid artery occlusion, 8-10 h after ischemia. These findings suggest that KGF has a protective effect against ischemic hippocampal neuronal damage in vivo, which may provide a new therapeutic strategy in the survival and reconstruction of neurons in response to cerebral injury.     

Preservation of GABAergic perikarya and boutons after transient ischemia in the gerbil hippocampal CA1 field

Using an antibody directed against the γ-aminobutyric acid (GABA)-synthetizing enzyme glutamate decarboxylase (GAD) the fate of the GABAergic innervation was investigated in the hippocampal field CA1 of gerbils up to 14 days after a bilateral transient 5-min occlusion of carotid arteries. As described previously, the CA1 pyramidal cells were subject to the ischemia-induced delayed neuronal death, the first signs of which were detectable after 2 days and which was fully developed after 4 days. Local GAD-immunoreactive neurons and boutons, however, exhibited no changes in their distribution and morphology over the whole 14-day period investigated, as studied both at the light and electron microscopic level. Thus, it can be assumed that the increased excitation observed during the development of delayed neuronal death, is not due to a loss of GABAergic neuronal profiles. The resistance of the GABAergic neurons to the ischemic insult is discussed in relation to the presence of Ca²⁺-binding proteins in this class of neurons, and the long persistance of innervation in an area nearly devoid of postsynaptic targets is considered.     

Neuroprotective effects of bilobalide, a component of Ginkgo biloba extract (EGb 761) in global brain ischemia and in excitotoxicity-induced neuronal death

In this study, we compared the protective effect of bilobalide, a purified terpene lactone component of ginkgo biloba extract EGb 761, (definition see editorial) and EGb 761 against ischemic injury and against glutamate-induced excitotoxic neuronal death. In ischemic injury, we measured neuronal loss and the levels of mitochondrial DNA (mtDNA)-encoded cytochrome oxidase (COX) subunit III mRNA in vulnerable hippocampal regions of gerbils. At 7 days of reperfusion after 5 min of transient global ischemia, a significant increase in neuronal death and a significant decrease in COX III mRNA were observed in the hippocampal CA1 neurons. Oral administration of EGb 761 at 25, 50 and 100 mg/kg/day and bilobalide at 3 and 6 mg/kg/day for 7 days before ischemia progressively protected CA1 neurons from death and from ischemia-induced reductions in COX III mRNA. In rat cerebellar neuronal cultures, addition of bilobalide or EGb 761 protected in a dose-dependent manner against glutamate-induced excitotoxic neuronal death (effective concentration [EC (50)] = 5 microg/ml (12 microM) for bilobalide and 100 microg/ml for EGb 761. These results suggest that both EGb 761 and bilobalide are protective against ischemia-induced neuronal death in vivo and glutamate-induced neuronal death in vitro by synergistic mechanisms involving anti-excitotoxicity, inhibition of free radical generation, scavenging of reactive oxygen species, and regulation of mitochondrial gene expression.     

Behavioral and histological changes after repeated brief cerebral ischemia by carotid artery occlusion in gerbils

The effects of repeated brief episodes of cerebral ischemia on passive avoidance learning and hippocampal neuronal degeneration were investigated in gerbils. Latency of step-through was shortened for the entire 7-day period when gerbils were trained at day 3 after 3 episodes of carotid artery occlusion for 2 min each at 60-min intervals. In addition, latency of passive avoidance was shortened for 63 days when gerbils were retrained at 14 days after occlusion. Severe neuronal degeneration in the CA1 regions of the hippocampus was observed 4 and 17 days after occlusion. A close correlation was seen between latency in the passive avoidance response and neurological degeneration of the hippocampal CA1 region. This fact was strongly supported by the results of another experiment that 1 2-min occlusion with almost no neuronal degeneration did not affect learning behavior while 3 1-min occlusions showed neuronal death and impairment of learning behavior of a moderate degree 4 days after occlusion. These results suggest that this gerbil model with carotid artery occlusion is useful for the quantitative measurement of functional changes in the chronic phase of repeated cerebral ischemia.     

Lithium ion does not protect brain against transient ischemia in gerbils.

It has been proposed that lithium ion desensitizes neuronal receptors that function via the inositol phospholipid signaling mechanism. We examined the effects of lithium chloride on the morphologic outcome after 5 minutes of cerebral ischemia induced in gerbils by occluding both common carotid arteries under brief halothane anesthesia. In three treated groups of 10 gerbils each, 5 meq/kg i.p. lithium chloride was given 2 days, 1 day, and 2 hours before ischemia; 2 hours before ischemia; or immediately after the end of ischemia. Corresponding control groups of nine or 10 gerbils each received equivalent volumes of saline injected at comparable times. All gerbils were perfusion-fixed 1 week later, and neuronal density of the hippocampal CA1 pyramidal cells was determined. Lithium induced very mild intraischemic systemic hypothermia, but postischemic hyperthermia developed in both treated and control groups. Neuronal densities were equal in corresponding groups. The results indicate that our regimen of lithium administration provides no benefit in survival of hippocampal neurons, and intraischemic hypothermia of less than 0.8 degrees C is not protective. Other strategies to inactivate the signal transduction system that is specific for excitatory neurotransmission should be evaluated.     

Comparative neuroprotective properties of the benzodiazepine receptor full agonist diazepam and the partial agonist PNU-101017 in the gerbil forebrain ischemia model

Recent studies have demonstrated the neuroprotective properties of the novel imidazoquinoline benzodiazepine receptor partial agonist, PNU-101017, in the gerbil forebrain ischemia model. The compound effectively reduces delayed post-ischemic (5 min bilateral carotid occlusion) hippocampal CA₁ neuronal degeneration even when its administration is witheld until 4 h after reperfusion and the effect is unrelated to hypothermia. The purpose of the present study was to determine the comparative abilities of PNU-101017 versus the full agonist diazepam to attenuate post-ischemic CA₁ damage. Male gerbils were treated either 30 min before ischemia induction or immediately after reperfusion with an initial dose of PNU-101017 (30 mg/kg i.p.) or diazepam (10 mg/kg i.p.) with a second dose being given at 2 h after reperfusion. Possible hypothermic effects of either compound were prevented by external heating. In vehicle (0.05 N HCl)-treated gerbils, the loss of hippocampal CA₁ neurons at 5 days was 85%. PNU-101017 pretreatment reduced the loss to 50% (p<0.05 vs. vehicle) whereas pretreatment with diazepam attenuated damage to only 17% (p<0.001 vs. vehicle). Delaying treatment with PNU-101017 until just after reperfusion still resulted in a reduction in CA₁ degeneration statistically that was indistinguishable from that seen with pretreatment. In contrast, diazepam post-treatment did not significantly decrease CA₁ neuronal loss. These results suggest that a benzodiazepine receptor partial agonist may have greater neuroprotective practicality than a full agonist for the treatment of global cerebral ischemia. The mechanistic basis for this difference may relate to the partially pro-excitatory neuronal response to endogenous GABA before and after neuronal insult.     

Single oral dose of geranylgeranylacetone for protection against delayed neuronal death induced by transient ischemia

The present study evaluated the potential effect of geranylgeranylacetone (GGA), which is known as an antiulcer agent, against the delayed neuronal death (DND) of hippocampal neurons an otherwise lethal ischemic insult. Pretreatment with single oral GGA (800 mg/kg, 2 days before ischemia) significantly attenuated DND in gerbil hippocampus. These effects of GGA were prevented by the coinjection of MK801, a noncompetitive N-methyl-d-aspartate glutamate receptor antagonist, which indicates that the protection was indeed glutamate receptor activation mediated.     

Erythropoietin protects neurons against chemical hypoxia and cerebral ischemic injury by up-regulating Bcl-xL expression

Erythropoietin (EPO) promotes neuronal survival after cerebral ischemia in vivo and after hypoxia in vitro. However, the mechanisms underlying the protective effects of EPO on ischemic/hypoxic neurons are not fully understood. The present in vitro experiments showed that EPO attenuated neuronal damage caused by chemical hypoxia at lower extracellular concentrations (10(- 4)-10(-2) U/ml) than were previously considered. Moreover, EPO at a concentration of 10(-3) U/ml up-regulated Bcl-xL mRNA and protein expressions in cultured neurons. Subsequent in vivo study focused on whether EPO rescued hippocampal CA1 neurons from lethal ischemic damage and up-regulated the expressions of Bcl-xL mRNA and protein in the hippocampal CA1 field of ischemic gerbils. EPO was infused into the cerebroventricles of gerbils immediately after 3 min of ischemia for 28 days. Infusion of EPO at a dose of 5 U/day prevented the occurrence of ischemia-induced learning disability. Subsequent light microscopic examinations showed that pyramidal neurons in the hippocampal CA1 field were significantly more numerous in ischemic gerbils infused with EPO (5 U/day) than in those receiving vehicle infusion. The same dose of EPO infusion caused significantly more intense expressions of Bcl-xL mRNA and protein in the hippocampal CA1 field of ischemic gerbils than did vehicle infusion. These findings suggest that EPO prevents delayed neuronal death in the hippocampal CA1 field, possibly through up-regulation of Bcl-xL, which is known to facilitate neuron survival.     

Neuroprotective effects of ischemic preconditioning on hippocampal CA1 pyramidal neurons through maintaining calbindin D28k immunoreactivity following subsequent transient cerebral ischemia

Ischemic preconditioning elicited by a non-fatal brief occlusion of blood flow has been applied for an experimental therapeutic strategy against a subsequent fatal ischemic insult. In this study, we investi-gated the neuroprotective effects of ischemic preconditioning (2-minute transient cerebral ischemia) on calbindin D28k immunoreactivity in the gerbil hippocampal CA1 area following a subsequent fatal tran-sient ischemic insult (5-minute transient cerebral ischemia). A large number of pyramidal neurons in the hippocampal CA1 area died 4 days after 5-minute transient cerebral ischemia. Ischemic preconditioning reduced the death of pyramidal neurons in the hippocampal CA1 area. Calbindin D28k immunoreactivity was greatly attenuated at 2 days after 5-minute transient cerebral ischemia and it was hardly detected at 5 days post-ischemia. Ischemic preconditioning maintained calbindin D28k immunoreactivity after transient cerebral ischemia. These findings suggest that ischemic preconditioning can attenuate transient cerebral ischemia-caused damage to the pyramidal neurons in the hippocampal CA1 area through maintaining cal-bindin D28k immunoreactivity.     

Effect of cyclooxygenase and lipoxygenase inhibitors on delayed neuronal death in the gerbil hippocampus

The purpose of our study was to examine whether cyclooxygenase and lipoxygenase inhibitors ameliorate delayed neuronal death in the hippocampal CA1 sector in Mongolian gerbils after 5 minutes of forebrain ischemia. Gerbils were injected intraperitoneally with cyclooxygenase inhibitors piroxicam and flurbiprofen or with lipoxygenase inhibitors AA-861 and BW-755C. Seven days after ischemic insult, the animals were perfusion-fixed, and the neuronal density in the hippocampal CA1 sector was estimated. The average neuronal density in unoperated normal gerbils was 247 +/- 9/mm (mean +/- SEM). In ischemic gerbils with vehicle administration, the average neuronal densities were 13 +/- 2, 14 +/- 2, 13 +/- 2, and 13 +/- 1 for piroxicam, flurbiprofen, AA-861, and BW-755C, respectively. The average neuronal densities in ischemic gerbils treated with 1.5 and 10 mg/kg piroxicam and 1.5 and 10 mg/kg flurbiprofen were 13 +/- 2, 194 +/- 9, 19 +/- 5, and 143 +/- 12, respectively. In ischemic gerbils treated with 15 and 100 mg/kg AA-861 and 30 mg/kg BW-755C, the average neuronal densities were 12 +/- 1, 13 +/- 1, and 14 +/- 2, respectively. At their higher doses, both piroxicam and flurbiprofen significantly (p less than 0.01) ameliorated delayed neuronal death in the hippocampal CA1 sector. Our results suggest that cyclooxygenase products play an important role in the development of delayed neuronal injury after cerebral ischemia.     

Systemic administration of MK-801 protects against ischemia-induced hippocampal neurodegeneration in the gerbil

The neuroprotective effects of MK-801, a noncompetitive antagonist of N-methyl-D-aspartate (NMDA) receptors, were evaluated in models of cerebral ischemia using Mongolian gerbils. Bilateral occlusion of the carotid arteries for a period of 5 min resulted in a consistent pattern of degeneration of hippocampal CA1 and CA2 pyramidal neurons, which was quantified using an image analyzer. Systemic administration of MK-801 (0.01-10 mg/kg, i.p.) 1 hr prior to the occlusion caused a dose-dependent protection of the CA1 and CA2 neurons. The ED50 value for neuroprotection by MK-801 was calculated to be 0.3 mg/kg, and at doses greater than or equal to 3 mg/kg the majority of animals were completely protected against the ischemic insult. Systemic administration of MK-801 (1 or 10 mg/kg, i.p.) 1 hr prior to unilateral occlusion of the right carotid artery resulted in significant protection against hippocampal neurodegeneration following 10 min of occlusion, and increased the survival rate after 30 min of occlusion. The potent neuroprotective effects of MK-801 in these cerebral ischemia models add further weight to the evidence that NMDA receptors are involved in the mechanism of ischemia-induced neuronal degeneration.     

Prevention of delayed neuronal death in gerbil hippocampus by ion channel blockers

We used a gerbil model of cerebral ischemia to study the effects of ion channel blockers on neuronal death resulting from enhanced glutamate release and calcium ion influx. The common carotid arteries of gerbils were occluded for 5 minutes and injected intraperitoneally immediately after ischemia with an alkylene iminopropylene derivative (glutamate blocker) or a piperazinyl ethanol derivative (calcium blocker) given at high or low doses. Two vehicle groups received saline or 0.2% methyl cellulose solution. Seven days later, the gerbils were perfusion-fixed and their brains were processed for histologic study. The number of neurons per millimeter (neuronal density) of the CA1 region was calculated, and the neuronal density in each group was statistically compared using the Mann-Whitney U test. Compared with a control group not subjected to carotid ligation, neurons of the two vehicle groups and the low-dose calcium blocker group were almost nonexistent in the CA1 region. Neuronal densities of the glutamate blocker group and the high-dose calcium blocker group were similar and were found to be within normal limits by statistical analysis. Our study shows that detrimental membrane phenomena and the incidence of delayed neuronal death may be counteracted by the systemic administration of these ion channel blockers after ischemic insult.     

Delayed hippocampal neuronal death in young gerbil following transient global cerebral ischemia is related to higher and longer-term expression of p63 in the ischemic hippocampus

The tumor suppressor p63 is one of p53 family members and plays a vital role as a regulator of neuronal apoptosis in the development of the nervous system. However, the role of p63 in mature neuronal death has not been addressed yet. In this study, we first compared ischemia-induced effects on p63 expression in the hippocampal regions (CA1–3) between the young and adult gerbils subjected to 5 minutes of transient global cerebral ischemia. Neuronal death in the hippocampal CA1 region of young gerbils was significantly slow compared with that in the adult gerbils after transient global cerebral ischemia. p63 immunoreactivity in the hippocampal CA1 pyramidal neurons in the sham-operated young group was significantly low compared with that in the sham-operated adult group. p63 immunoreactivity was apparently changed in ischemic hippocampal CA1 pyramidal neurons in both ischemia-operated young and adult groups. In the ischemia-operated adult groups, p63 immunoreactivity in the hippocampal CA1 pyramidal neurons was significantly decreased at 4 days post-ischemia; however, p63 immunoreactivity in the ischemia-operated young group was significantly higher than that in the ischemia-operated adult group. At 7 days post-ischemia, p63 immunoreactivity was decreased in the hippocampal CA1 pyramidal neurons in both ischemia-operated young and adult groups. Change patterns of p63 level in the hippocampal CA1 region of adult and young gerbils after ischemic damage were similar to those observed in the immunohistochemical results. These findings indicate that higher and longer-term expression of p63 in the hippocampal CA1 region of the young gerbils after ischemia/reperfusion may be related to more delayed neuronal death compared to that in the adults.     

Neuroprotective effects of bilobalide, a component of the Ginkgo biloba extract (EGb 761), in gerbil global brain ischemia.

The neuroprotective effect of Ginkgo biloba extract (EGb 761) against ischemic injury has been demonstrated in animal models. In this study, we compared the protective effect of bilobalide, a purified terpene lactone from EGb 761, and EGb 761 against ischemic injury. We measured neuronal loss and the levels of mitochondrial DNA (mtDNA)-encoded cytochrome oxidase (COX) subunit III mRNA in vulnerable hippocampal regions of gerbils. At 7 days of reperfusion after 5 min of transient global forebrain ischemia, a significant increase in neuronal death and a significant decrease in COX III mRNA were observed in the hippocampal CA1 neurons. Oral administration of EGb 761 at 25, 50 and 100 mg/kg/day and bilobalide at 3 and 6 mg/kg/day for 7 days before ischemia progressively protected CA1 neurons from death and from ischemia-induced reductions in COX III mRNA. In addition, both bilobalide and EGb 761 protected against ischemia-induced reductions in COX III mRNA in CA1 neurons prior to their death, at 1 day of reperfusion. These results suggest that oral administration of bilobalide and EGb 761 protect against ischemia-induced neuron death and reductions in mitochondrial gene expression.     

Wide therapeutic time window for fasudil neuroprotection against ischemia-induced delayed neuronal death in gerbils

The neuroprotective potential and therapeutic time window for fasudil, a Rho-kinase inhibitor (RKI), were evaluated for delayed neuronal death in gerbils. A preliminary screening was done on fasudil, ozagrel, and edaravone using a single administration in a delayed neuronal death study. Intraperitoneal (i.p.) administration of edaravone, a free radical scavenger (3, 10 mg/kg) immediately after re-circulation did not reduce neuronal degeneration. We previously reported that ozagrel, a thromboxane A(2) synthetase inhibitor (30 mg/kg) also did not reduce neuronal degeneration, while fasudil (3, 30 mg/kg) significantly protected against the ischemia-induced neuronal loss. To clarify the therapeutic time window of fasudil, which showed a positive effect in a preliminary screening, animals received their first i.p. administration of fasudil (10 mg/kg) 24 or 48 h after ischemia. Administration of fasudil twice daily was continued until day 6. Fasudil significantly protected against the ischemia-induced delayed neuronal death when the treatment was started 24 h after ischemia. In gerbils, hydroxyfasudil, an active metabolite of fasudil, was found following an i.p. administration of fasudil (10 mg/kg), and the value of the area under the plasma level curve of hydroxyfasudil was 7 times higher than that of fasudil. Hydroxyfasudil may contribute to the potency of fasudil. The present findings indicate that the RKI fasudil reduces ischemic neuronal damage with a wide therapeutic time window in gerbil, and may be useful in the treatment of acute ischemic stroke in humans.     

A reversible type of neuronal injury following ischemia in the gerbil hippocampus

The Mongolian gerbil is known to develop delayed neuronal death in the hippocampus following brief forebrain ischemia (Brain Res 239: 57-69, 1982). The effect of pentobarbital on this slow process of neuronal damage was examined. Immediately following 5 min of bilateral carotid occlusion, pentobarbital (10, 20, or 40 mg/kg) was injected. The control animals received saline injection. Seven days following ischemic insult, animals were perfusion-fixed and the neuronal density in the hippocampal CA1 subfield was counted. Most of the neurons in the CA1 sector survived ischemic insult when pentobarbital was given, whereas most of control group neurons were lost without the treatment. The average neuronal density of 20 mg/kg group was 168.2 +/- 12.3 (SEM) per 1 mm linear length of the CA1 subfield. The density in 40 mg/kg group was 181.1 +/- 14.9. The neuronal density in the whole control group was 34.3 +/- 5.1. The density of unoperated normal gerbils was 212.3 +/- 3.9. This result indicates that the neuronal damage of "delayed neuronal death" is reversible. On the other hand, when pentobarbital was injected 1 hr following ischemia, it showed no effect. The cell change in the CA1 sector, reversible at the initial stage, seems to rapidly become irreversible, while neurons still remain intact morphologically.     

异丙酚对全脑缺血大鼠海马CA_1区锥体神经元的保护作用

目的 观察不同剂量、给药时间点及给药方式的异丙酚对全脑缺血大鼠海马CA_1区锥体细胞的保护作用.方法 80只雄性Wistar大鼠按照不同的给药剂量、给药时间点及方式随机分组.采用4血管闭塞法制造大鼠全脑缺血模型,硫堇染色下对锥体细胞的死亡进行评估.结果 异丙酚可剂量依赖性地保护锥体细胞,给药时间点越接近脑缺血,异丙酚的神经保护作用越明显.持续静脉输注异丙酚的神经保护作用较好.结论 在脑缺血前后的"有效时间窗"内,静脉持续输注麻醉剂量的异丙酚可有效地保护神经元.     

Calpain inhibitor entrapped in liposome rescues ischemic neuronal damage

Transient forebrain ischemia induces activation of calpain and proteolysis of a neuronal cytoskeleton, fodrin, in gerbil hippocampus. This phenomenon precedes delayed neuronal death in hippocampal CA1 neurons. We examined effects of a calpain inhibitor on delayed neuronal death after transient forebrain ischemia. In gerbils, a selective calpain inhibitor entrapped in liposome was given transvenously and 30 min later, 5-min forebrain ischemia was produced by occlusion of both common carotid arteries. On day 7, CA1 neuronal damage was examined in the hippocampal slices stained with cresyl violet. Calpain-induced proteolysis of fodrin was also examined by immunohistochemistry and immunoblot. Additionally, to assure entrapment of the inhibitor by CA1 neurons, the inhibitor-liposome complex was labeled with FITC and given to gerbils. Fluorescence in the hippocampal slices was examined by confocal laser scanning microscope. Selective CA1 neuronal damage induced by forebrain ischemia was prevented by administration of the inhibitor in a dose-dependent manner. Calpain-induced proteolysis of fodrin was also extinguished by the calpain inhibitor in a dose-dependent manner. Bright fluorescence of the FITC-labeled inhibitor was observed in the CA1 neurons. The data show an important role of calpain in the development of the ischemic delayed neuronal death. Calpain seems to produce neuronal damage by degrading neuronal cytoskeleton. Our data also show a palliative effect of the calpain inhibitor on the neurotoxic damage, which offers a new and potent treatment of transient forebrain cerebral ischemia.     

Indomethacin ameliorates ischemic neuronal damage in the gerbil hippocampal CA1 sector

The purpose of our experiment was to examine whether the cyclooxygenase inhibitor indomethacin ameliorates neuronal injury in the gerbil hippocampal CA1 sector following 5 minutes of forebrain ischemia. Thirty minutes before bilateral carotid artery occlusion, Mongolian gerbils were injected intraperitoneally with 1 (n = 10), 2 (n = 10), 5 (n = 12), or 10 (n = 7) mg/kg of indomethacin. Seven days after occlusion, the gerbils were perfusion-fixed and neuronal density in the hippocampal CA1 sector was assessed. The mean +/- SEM neuronal density in nine unoperated normal gerbils was 307 +/- 9/mm, in 10 untreated ischemic gerbils 55 +/- 21/mm, and in seven vehicle-treated ischemic gerbils 15 +/- 9/mm. The mean +/- SEM neuronal density in ischemic gerbils treated with 1, 2, 5, or 10 mg/kg indomethacin was 132 +/- 28/mm, 154 +/- 29/mm, 176 +/- 30/mm, and 136 +/- 39/mm, respectively. Indomethacin at any dose significantly ameliorated ischemic neuronal damage in the gerbil hippocampal CA1 sector.