Common Carotid Arteries Of Gerbils Research Articles

Effects of swimming rehabilitation training on learning and memory function and CaMK II in gerbils with cerebral ischemia reperfusion

Objective To explore the protective effect and mechanism of swimming rehabilitation training on learning and memory impairment of cerebral ischemia reperfusion gerbil. Methods Forty adult healthy male gerbils were randomly divided into sham group, sham+ swimming group (Sham+ S group), cerebral ischemia / reperfusion group (I/R group), cerebral ischemia/reperfusion+ swimming group (I/R+ S group), with 10 rats in each group. The gerbil models of cerebral ischemia/reperfusion in I/R group and I/R+ S group were established by blocking bilateral common carotid artery, while for gerbils in Sham group and Sham+ S group, only bilateral common carotid arteries of gerbils were exposed, but no arteries were clamped. Morris water maze was used to detect the changes of learning and memory function in rats.Oxidative stress injury in hippocampal neurons was detected by detection kit analysis.And the expression of Bax, Bcl-2 and CaMK Ⅱ protein in hippocampal tissue was detected by Western blot. Results Compared with Sham group, the gerbils in I/R group had longer positioning cruise time and less shuttle times (both P<0.01). Compared with I/R group, the positioning cruise time and shuttle times in I/R+ S group were significantly shortened and increased respectively (both P<0.01). Compared with sham group(SOD: (123.13±7.50)U/mg, GSH: (42.10±2.17)μg/g, GSH-Px: (61.37±2.51)μg/g, MDA: (2.91±0.23)nmol/mg), the activities of SOD, GSH, GSH-Px in I/R group decreased significantly, while the content of MDA increased significantly(SOD: (75.50±6.96)U/mg, GSH: (22.50±1.64)μg/g, GSH-Px: (33.15±2.04)μg/g, MDA: (5.96±0.32)nmol/mg; all P<0.01). Furthermore, compared with I/R group, the above indexes in I/R+ S group were significantly reversed(SOD: (110.30±5.90)U/mg, GSH: (34.31±1.73)μg/g, GSH-Px: (50.13±2.31)μg/g, MDA: (3.57±0.29)nmol/mg; all P<0.01). Compared with Sham group, the expression of Bax protein in hippocampus of gerbils in I/R group was increased, while the expression of Bcl-2 protein and p-CaMK Ⅱ protein was decreased (all P<0.05). Compared with I/R group, the expression of Bax protein in hippocampus of gerbils in I/R+ S group was decreased, while the expression of Bcl-2 protein and p-CaMK Ⅱ protein was increased (all P<0.05). Conclusion Swimming rehabilitation training can improve learning and memory impairment of gerbils after ischemia-reperfusion through anti-oxidative stress and anti-apoptosis, which may be related to CaMK Ⅱ signaling system. Key words: Cerebral ischemia/reperfusion; Rehabilitation training by swimming; Calcium/calmodulin-dependent protein kinase Ⅱ; Learning and memory function; Gerbil

Spatial and temporal patterns of oxidative stress in the brain of gerbils submitted to different duration of global cerebral ischemia

The present study was undertaken to examine spatial and temporal patterns of oxidative stress rate in the brain of Mongolian gerbils submitted to different duration of global ischemia/reperfusion. The common carotid arteries of gerbils were occluded for 5, 10, or 15 min. We followed the temporal ischemia-induced oxidative stress rate, the most important factor that exacerbates brain damage by reperfusion, starting from 24 h up to 28 days after reperfusion. The spatial ischemia-induced oxidative stress distribution was measured parallely in different brain regions: forebrain cortex, striatum, hippocampus and cerebellum. Post-ischemic effects were followed in vivo by monitoring the neurological status of whole animals and at the intracellular level by standard biochemical assays in different brain regions. We measured superoxide production, superoxide dismutase activity, nitric oxide production, index of lipid peroxidation, and reduced glutathione. Our results revealed a pattern of dynamic changes in each oxidative stress parameter that corresponded with ischemia duration in all tested brain structures. The highest levels were obtained in the first 24 h after the insult. After that, they slowly returned to nearly control values 28 days after reperfusion (with the exception of SOD activity that returned to control values at fourth day after reperfusion). The most sensitive oxidative stress parameter was index of lipid peroxidation. Our study confirmed spatial distribution of ischemia-induced oxidative stress. Tested brain structures showed different sensitivity to each oxidative stress parameter, although their basal levels were similar. These new findings could be valuable for creation and strategy of post-ischemic therapy.

Neuroprotective efficiency of NMDA receptor blockade in the striatum and CA3 hippocampus after various durations of cerebral ischemia in gerbils

The purpose of this study was to investigate neuroprotective efficiency of N-methyl D-aspartate (NMDA) receptor (NMDAR) blockade on the neuronal damage in the less studied and allegedly less affected CA3 hippocampus and striatum in the Mongolian gerbil model of global cerebral ischemia. The common carotid arteries of gerbils were occluded for 5, 10 or 15 minutes. Gerbils were given a low dose of non-competitive NMDA antagonist (MK-801, 3 mg/kg i.p.) or saline immediately after the occlusion in normothermic conditions. Neuronal damage was examined on 4th, 14th and 28th day after reperfusion. The effect of NMDAR blockade was followed in vivo by monitoring the neurological status of whole animals or at the cellular level by standard light- and confocal microscopy on brain slices. Increased duration of cerebral ischemia resulted in a progressive loss of striatal and CA3 hippocampal neurons. The most beneficial NMDAR blockade effect was observed when the neuronal damage was most severe - on the 28th day after 15-min ischemia. As judged by morphological and neurological data, the effect of ischemia is also apparent in the presumed less vulnerable regions (CA3 and striatum) which are functionally important in stroke plasticity. So, NMDAR blockade in normothermic conditions showed neuroprotective efficiency.

MK-801 effect on regional cerebral oxidative stress rate induced by different duration of global ischemia in gerbils

We investigated MK-801 effect on ischemia-induced oxidative stress-the most important factor that exacerbates brain damage by reperfusion. The common carotid arteries of gerbils were occluded for 5, 10, or 15 min. Immediately after the occlusion, MK-801 (3 mg/kg i.p.) or saline were given in normothermic conditions. The MK-801 effects were followed in vivo by monitoring the neurological status of animals and at the intracellular level by standard biochemical assays. We investigated nitric oxide levels, superoxide production, superoxide dismutase activity, index of lipid peroxidation (ILP), and reduced glutathione content in hippocampus, striatum, forebrain cortex, and cerebellum. The measurements took place at different times (1, 2, 4, 7, 14, and 28 days) after reperfusion. Increased duration of cerebral ischemia resulted in a progressive induction of oxidative stress. Our results revealed pattern of dynamic changes in each oxidative stress parameter level which corresponded with ischemia duration in all tested brain structures. Most sensitive oxidative stress parameters were ILP and superoxide production. Our study confirmed spatial distribution of ischemia-induced oxidative stress. Tested brain structures showed different sensitivity to each oxidative stress parameter. As judged by biochemical and neurological data, applied MK-801 showed neuroprotective efficiency by reduction of ischemia-induced oxidative stress in brain.

Functional recovery after delayed treatment of ischemic stroke with melanocortins is associated with overexpression of the activity-dependent gene Zif268

Melanocortin peptides afford strong neuroprotection and improve functional recovery in experimental ischemic stroke; they also have established neurotrophic actions. The expression of the immediate early gene Zif268 is dependent on synaptic activity and is involved in injury repair and memory formation. Here, we investigated the role of Zif268 in learning and memory recovery after delayed treatment of ischemic stroke with the melanocortin analog [Nle4, d-Phe7]α-MSH (NDP-α-MSH). A 10-min period of global cerebral ischemia was induced by occluding both common carotid arteries in gerbils. Treatment with a nanomolar dose of NDP-α-MSH (every 12h for 11days) was performed starting 3h or 9h after stroke induction; where indicated, gerbils were pretreated with the melanocortin MC4 receptor antagonist HS024. Animals were subjected to the Morris water-maze test (four sessions from 4 to 50days after the ischemic episode). Fifty days after stroke, histological damage and Zif268 expression were investigated in the hippocampus. Treatment with NDP-α-MSH significantly reduced hippocampal damage, including neuronal death, and improved learning and memory recovery. This protective effect was long-lasting (50days, at least) and associated with Zif268 overexpression, with both schedules of NDP-α-MSH treatment. Pharmacological blockade of MC4 receptors prevented these effects. Our data indicate that MC4 receptor-mediated actions of melanocortins could trigger repair mechanisms able to improve neuronal functionality and synaptic plasticity, and to promote long-lasting functional recovery from ischemic stroke with Zif268 gene involvement.

Adopted cognitive tests for gerbils: Validation by studying ageing and ischemia

Transient occlusion of common carotid arteries in gerbils is a simple and widely used model for assessing histological and functional consequences of transient forebrain ischemia and neuroprotective action of pharmaceuticals. In the present study we aimed to introduce additional behavioural tests as novel object recognition and food-motivated hole-board learning in order to measure attention and learning capacity in gerbils. For validating these cognitive tests the effects of ageing (4, 9 and 18 months) and those of transient forebrain ischemia induced by bilateral carotid occlusion at 9 months of age were investigated. Neuronal cell death was estimated in the hippocampus using TUNEL and caspase-3 double fluorescence labelling and confocal microscopy. Ageing within the selected range although influenced ambulatory activity, did not considerably change attention and memory functions of gerbils. As a result of transient ischemia a selective neuronal damage in CA1 and CA2 regions of the hippocampus has been observed and tested 4 days after the insult. Ischemic gerbils became hyperactive, but showed decreased attention and impaired spatial memory functions as compared to sham-operated controls. According to our results the novel object recognition paradigm and the hole-board spatial learning test could reliably be added to the battery of conventional behavioural tests applied previously in this species. The novel tests can be performed within a wide interval of adult age and provide useful additional methods for assessing ischemia-induced cognitive impairment in gerbils.

A novel conjugate of low-molecular-weight heparin and Cu,Zn-superoxide dismutase: Study on its mechanism in preventing brain reperfusion injury after ischemia in gerbils

Low-molecular-weight heparin (LMWH) and Cu,Zn-superoxide dismutase (SOD) were extensively investigated on preventing brain reperfusion injury after ischemia (BRII) in the past few years and both exhibited some advantages as well as limits in practice. To explore whether chemical modification for LMWH and SOD can lead to improved bioactivity, in our present study, we examined the efficacy of LMWH conjugated SOD (LMWH − SOD) in the model of BRII gerbils. Ischemia/reperfusion was performed for 5 min by clamping the bilateral common carotid arteries of gerbils. LMWH − SOD, SOD and the mixture of LMWH and SOD (LMWH + SOD) were administered intravenously to corresponding animals just before ischemia. 24 h after reperfusion, serum malondialdehyde (MDA) content and SOD activity were measured, the expression of intercellular adhesion molecule-1 (ICAM-1) was examined by immunohistochemistry, and the brain sections were processed for Nissl staining and terminal deoxynucleotidyl transferase (TdT)-mediated dUTP nick end-labeling. The results showed that LMWH − SOD significantly lowered MDA content ( P < 0.001, versus SOD and LMWH + SOD) and elevated SOD activity ( P < 0.05, versus SOD and LMWH + SOD) in the serum of BRII gerbils. Immunohistochemical results indicated ICAM-1 positive staining was lighter, pyramidal cells of hippocampal CA 1 region were more regular and the changes in cell edema were minor, and apoptosis of hippocampal cells was milder in LMWH − SOD treated animals than in SOD or LMWH + SOD treated animals, untreated BRII animals and sham-operated animals. The results suggest that the novel LMWH − SOD conjugate can inhibit upregulation of ICAM-1 and prevent neuronal cell apoptosis in BRII gerbils, and the LMWH − SOD conjugate has better anti-inflammatory and neuroprotective effects in BRII than native SOD and the mixture of LMWH and SOD.

Temporal patterns of motor behavioural improvements by MK-801 in Mongolian gerbils submitted to different duration of global cerebral ischemia

The purpose of this study was to investigate the temporal pattern of NMDA receptors antagonist—MK-801 on motor behaviour parameters in gerbils submitted to different duration of global cerebral ischemia. The common carotid arteries of gerbils were occluded for 5, 10 or 15 min. Gerbils were given MK-801 (3 mg/kg i.p.) or saline immediately after the occlusion in normothermic conditions prior to testing. Motor activity was registered 1, 2, 4, 7, 14, 21 and 28 days after reperfusion during 60 min by open field test. At the same time, the effect of NMDA receptor blockade was followed in vivo by monitoring the neurological status of whole animals or at the cellular level by standard light and confocal microscopy on brain slices. Post-ischemic gerbils quickly developed hypermotor response with the most intensity in animals submitted to 15 min ischemia. MK-801 administrated immediately after ischemia significantly decreased this hyperactivity. In all ischemic-treated animals, behavioural suppression by MK-801 was observed already 1 day after occlusion and was lasting as far as observed ischemia-dependent hypermotor responses. Beneficial effect of MK-801 was also confirmed by morphological and neurological status data. These findings suggest that sustained ischemia-induced hyperactivity is related to abnormalities in NMDA glutamatergic function, as well as its manifestation could be completely abolished by NMDA receptor blockade immediately after ischemic insult.

Use of confocal microscopy in the study of ischemia-induced hippocampal neuronal damage

The present study was undertaken to reveal by means of confocal laser microscopy the cytoarchitecture of hippocampal CA3 neurons in Mongolian gerbils before and after cerebral ischemia of different duration. The common carotid arteries of gerbils were occluded for 5, 10, or 15 min. On the 4th, 14th and 28th day after reperfusion, neuronal damage was examined by laser scanning confocal microscopy in the CA3 region of hippocampus (30 ?m slices). Slices were stained with fluorescent Nissl staining and fluorescent membrane tracer DiI. Increased duration of cerebral ischemia resulted in a progressive loss of hippocampal CA3 neurons. Four days after the ischemic insult, neuronal damage in the hippocampal CA3 region was mild but visible. On the 28th day after reperfusion, neuronal damage in the observed brain structure was most severe. These results demonstrate the temporal profile of neuronal damage after an ischemic insult as observed using confocal microscopy.

Time course of motor behavior changes in Mongolian gerbils submitted to different durations of cerebral ischemia

In addition to morphological changes, global cerebral ischemia leads to functional changes that can be assessed by behavioral examination. The purpose of this study was to investigate the impact of the duration of global cerebral ischemia on the time course of a comprehensive set of motor behaviors in Mongolian gerbils. The common carotid arteries of gerbils were occluded either for 5 min, 10 min, or 15 min. Gerbil motor behavior was recorded in the open field at 24 h, 48 h, 4 days, 7 days, 14 days, 21 days, and 28 days after reperfusion. Each session lasted for 60 min and was composed of six intervals of 10 min. Our results revealed that ischemic gerbils quickly develop locomotor and stereotypic hyperactivity, with the expected decrease of resting time. The most evident effect was observed in gerbils submitted to a 15 min ischemia, whose locomotor activity returned to nearly normal values after 7 days. In contrast, the duration of global cerebral ischemia had no effects on rearing, clockwise, or counter-clockwise rotation. These findings indicate that exposure to global cerebral ischemia induces changes in locomotion, stereotypy, and resting time. The magnitude and duration of these effects depend on the duration of ischemia.

Protective effects of iptakalim, a novel ATP-sensitive potassium channel opener, on global cerebral ischemia-evoked insult in gerbils1

To investigate the protective role of iptakalim, a novel ATP sensitive potassium channel opener, on global cerebral ischemia-evoked insult in gerbils and glutamate-induced PC12 cell injury. Global cerebral ischemia was induced by occluding the bilateral common carotid arteries in gerbils for 5 min. The open field maze and T-maze were employed to investigate the experimental therapeutic value of iptakalim on ischemic brain insult (n=8). The pyramidal cells in the hippocampal CA1 regions were counted to assess the protective effects of iptakalim. Glutamate released from the gerbil hippocampus and PC12 cells were determined by HPLC. Intracellular calcium was measured by Fluo-3 AM with A Bio-Rad Radiance 2100TM confocal system in conjunction with a Nikon TE300 microscope. Astrocyte glutamate uptake measurements were determined by liquid scintillation counting. Iptakalim (0.5-4.0 mg/kg per day, ip) could reduce the high locomotor activity evoked by ischemia and improve global cerebral ischemia-induced working memory impairments. Histological studies revealed that iptakalim could increase the survival neuron in the hippocampus CA1 zone in a dose-dependent manner. Moreover, iptakalim could reverse ischemia-evoked increases of glutamate in the hippocampus of gerbils. In an in vitro study, iptakalim protected PC12 cells against glutamate-induced excitotoxicity, reduced the [Ca(2+)](i) increases, and enhanced the glutamate uptake activity of primary cultured astrocytes. Iptakalim plays a key role in preventing global cerebral ischemia-evoked insults in gerbils and glutamate-induced PC12 cell injury by anti-excitotoxicity. Iptakalim might be a promising novel candidate for the prevention and/or treatment of stroke.

Long-term treadmill exercise overcomes ischemia-induced apoptotic neuronal cell death in gerbils

It has been suggested that exercise may ameliorate neurologic impairment by impeding neuronal loss following various brain insults. In the present study, the effect of long-term treadmill exercise on short-term memory and apoptotic neuronal cell death in the hippocampus following transient global ischemia in gerbils was investigated. A step-down inhibitory avoidance task, terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) assay, and immunohistochemistry for caspase-3 were used for this study. Ischemia was induced by occlusion of both the common carotid arteries of gerbils for 5 min. Gerbils in the exercise groups were forced to run on a treadmill for 30 min once a day for 4 consecutive weeks. The present results reveal that treadmill exercise for 4 weeks improved short-term memory by suppressing the ischemia-induced apoptotic neuronal cell death in the hippocampus. Here in this study, we show that long-term treadmill exercise for 4 weeks overcomes the ischemia-induced apoptotic neuronal cell death and thus facilitates the recovery of short-term memory impairment induced by ischemic cerebral injury.

Glial glutamate transporter GLT-1 down-regulation precedes delayed neuronal death in gerbil hippocampus following transient global cerebral ischemia

Glial (GLT-1 and GLAST) and neuronal (EAAC1) high-affinity transporters mediate the sodium dependent glutamate reuptake in mammalian brain. Their dysfunction leads to neuronal damage by allowing glutamate to remain in the synaptic cleft for a longer duration. The purpose of the present study is to understand their contribution to the ischemic delayed neuronal death seen in gerbil hippocampus following transient global cerebral ischemia. The protein levels of these three transporters were studied by immunoblotting as a function of reperfusion time (6 h to 7 days) following a 10 min occlusion of bilateral common carotid arteries in gerbils. In the vulnerable hippocampus, there was a significant decrease in the protein levels of GLT-1 (by 36–46%, P<0.05; between 1 and 3 days of reperfusion) and EAAC1 (by 42–68%, P<0.05; between 1 and 7 days of reperfusion). Histopathological evaluation showed no neuronal loss up to 2 days of reperfusion but an extensive neuronal loss (by ∼84%, P<0.01) at 7 days of reperfusion in the hippocampal CA1 region. The time frame of GLT-1 dysfunction (1–3 days of reperfusion) precedes the initiation of delayed neuronal death (2–3 days of reperfusion). This suggests GLT-1 dysfunction as a contributing factor for the hippocampal neuronal death following transient global cerebral ischemia. Furthermore, decreased EAAC1 levels may contribute to GABAergic dysfunction and excitatory/inhibitory imbalance following transient global ischemia.

Neuroprotective Properties of the Novel Antiepileptic Lamotrigine in a Gerbil Model of Global Cerebral Ischemia

Elevated glutamate levels are thought to be a primary cause of neuronal death after global cerebral ischemia. The purpose of this study was to investigate the potential neuroprotective effects of lamotrigine, a novel antiepileptic drug that inhibits the release of glutamate in vitro, with both behavioral and histological measures of global ischemia in gerbils. The common carotid arteries of gerbils were occluded for either 5, 10, or 15 minutes. Twenty-one days after reperfusion, gerbils were tested for impairments in a spatial memory task (Morris water maze). After water maze testing the animals were killed, and damage to hippocampal pyramidal cells was assessed. The effect of lamotrigine on the behavioral and histological outcome of either 5 or 15 minutes of global ischemia was evaluated. Bilateral occlusion of the common carotid arteries for 5 minutes resulted in severe degeneration of hippocampal CA1 and CA2 pyramidal cells. Lamotrigine significantly prevented loss of hippocampal CA1 neurons when administered acutely (100 mg/kg PO) immediately after reperfusion or when administered in two equal doses of 30 or 50 mg/kg 2 hours before and immediately after reperfusion. Gerbils subjected to 5 minutes of ischemic insult were not impaired in their ability to solve a spatial memory task 21 days after cerebral ischemia. However, gerbils subjected to 10 and 15 minutes of carotid artery occlusion showed significant impairment in their ability to solve a water maze task. Lamotrigine significantly protected against the cognitive deficits associated with 15 minutes of cerebral ischemia. Histologically, increased durations of cerebral ischemia resulted in a progressive loss of CA1, CA2, and CA3 pyramidal cells. Lamotrigine completely protected gerbils exposed to 15 minutes of cerebral ischemia against CA3 cell loss and greatly reduced damage to the CA1 and CA2 cell tracts of the hippocampus. Lamotrigine also reduced the mortality associated with 15 minutes of ischemia. Lamotrigine had neuroprotective effects in a gerbil model of global cerebral ischemia. Lamotrigine protected gerbils against behavioral deficits resulting from 15 minutes of carotid occlusion and also prevented histological damage resulting from 5 and 15 minutes of global cerebral ischemia.

Effects of recombinant superoxide dismutase on manganese superoxide dismutase gene expression in gerbil hippocampus after ischemia.

We reported that recombinant human superoxide dismutase ameliorates delayed neuronal death in the postischemic gerbil hippocampus. Since postischemic induction of copper-zinc superoxide dismutase messenger RNA was abolished by this treatment, oxygen radicals generated on reperfusion may induce the expression of this gene. In the present study we examined whether oxygen radicals also induce the expression of manganese superoxide dismutase messenger RNA in the postischemic brain. We induced transient cerebral ischemia by occluding the bilateral common carotid arteries of gerbils. Recombinant human superoxide dismutase (8 x 10(5) U/kg) or apo-superoxide dismutase was administered intravenously 1 minute before a 5-minute occlusion of the carotid arteries. We analyzed both copper-zinc and manganese superoxide dismutase RNA by in situ hybridization histochemistry and by Northern and dot blot analyses using radioisotope-labeled oligonucleotide probes. Hybridization with the manganese superoxide dismutase messenger RNA occurred at the limit of detection in normal CA1 neurons. We observed striking increases in the labeling of CA1 up to 24 hours after 5 minutes of ischemia. The hybridization occurred anew in glial cells of the CA1 layer during 3 to 7 days. Pretreatment with recombinant human superoxide dismutase had no effect on the postischemic induction of manganese superoxide dismutase messenger RNA, whereas the same treatment significantly attenuated (P < .01) the increase in copper-zinc superoxide dismutase messenger RNA. Our results demonstrated temporal postischemic induction of manganese superoxide dismutase messenger RNA. The inducer may not be superoxide radicals but may be other chemical mediators such as cytokines.

Experimental Basis of Multi-Infarct Dementia: Memory Impairments in Rodent Models of Ischemia

Memory is an intellectual function that is initially and consistently impaired in patients with vascular dementia. Experimental approaches to vascular dementia have so far been confined to investigations of memory impairments in rodent ischemic models. Unilateral middle cerebral artery (MCA) occlusion, multiple small embolization or transient four-vessel occlusion in rats produced acute single or multiple infarctions. In such rats, significant memory impairments occurred during the subacute or chronic phases, but were partially reversible. Permanent stenosis of both common carotid arteries in gerbils caused no ischemic changes at 1 day after stenosis but induced multiple infarctions after 1 week of stenosis, probably due either to chronic recurrent ischemia resulting from transient repetitive obstruction of the carotid arteries or to chronic low perfusion. The memory impairments in this model were persistent. Permanent bilateral common carotid artery occlusion in rats produced multiple infarctions plus white matter changes after 1 week of occlusion. Marked memory impairment was also observed in this model. The results of the above studies suggest that memory impairments due to ischemic causes may be partially reversible provided that the infarctions occur only once and are followed by flow recovery. Memory impairments, however, appear to persist if the brain is exposed to chronic recurrent ischemia or chronic moderately low perfusion. A repetitive or persistent low-flow state appears to be an important factor in determining the irreversibility of cognitive impairments.

Selective sparing of NADPH‐diaphorase–somatostatin–neuropeptide Y neurons in ischemic gerbil striatum

In the present study we investigated the relative vulnerability of neuronal subsets in the striatum to ischemia that was induced by bilateral transient ligation of the common carotid arteries in gerbils. After 4 days of survival, brains were evaluated using histochemical methods (NADPH-diaphorase and silver degeneration procedures) and neurochemical methods with radioimmunoassays for somatostatin-, neuropeptide Y-, and substance P-like immunoreactivity and measurements of amino acids using high-pressure liquid chromatography with electrochemical detection. NADPH-diaphorase-positive neurons were strikingly preserved in the ischemic dorsolateral portion of the striatum, in which there was severe neuronal loss. There was no significant depletion of NADPH-diaphorase-positive neurons in the striatum or cerebral cortex. Concentrations of neuropeptide Y-like and somatostatin-like immunoreactivity were unchanged despite a significant 25% depletion of substance P-like immunoreactivity and gamma-aminobutyric acid. Ischemic brain damage may be mediated by a neurotoxic effect of glutamate acting at the N-methyl-D-aspartate (NMDA) receptor. Previous studies of NMDA excitotoxin lesions in rat striatum have shown a sparing of neurons containing NADPH-diaphorase, somatostatin, and neuropeptide Y. The similar sparing of these neurons following ischemic lesions in gerbil striatum provides further evidence that NMDA receptor activation may play a role in ischemic injury.

Propentofylline increases the survival rate in gerbils after brain ischemia: Influence on serotonin metabolism

AbstractFollowing the bilateral occulusion of common carotid arteries in gerbils for 15 min, the survival rate and serotonin metabolism were evaluated in groups of animals treated over a period 7 days by propentofylline or pentobarbitalor both propentofylline fo pentobarbital. In the non‐treated group as well as in animals treated by pentobarbital only, survival rate was about 29%, and serotonin metabolism was inhibited, both in the cerebral cortex and basal ganglia. Contrary to that, survival rate of animals treated by propentofylline or propentofylline and pentobarbital was about 70% at the 7th day after 15 min of ischemia, and serotonin metabolism was at the normal rate. These results show the superior effect of propentofylline in comparison to pentobarbital on the survival rate and serotonin metabolism after brain ischemia.

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.

Prediction of stroke before and after unilateral occlusion of the common carotid artery in gerbils.

A method was developed to predict the severity of cerebral ischemia before permanent occlusion of a common carotid artery in gerbils by observing the diameter and appearance of the artery after temporary occlusion and observing clinical signs after permanent occlusion. The severity of cerebral ischemia was confirmed by a sensitive immunohistochemical method and measurement of focal cerebral blood flow after 30 minutes' ischemia. All gerbils with greater than 40% reduction of the diameter and a white arterial margin distal to temporary occlusion developed severe neurologic signs following permanent occlusion, but no gerbils with reduction of less than 30% and a red arterial margin developed neurologic signs. With the cumulative neurologic score, gerbils could be divided into classes with no, mild, moderate, and severe symptoms, mostly after 10 minutes. Severely symptomatic gerbils were identified in 3 minutes. Extensive ischemic damage was observed in severely symptomatic gerbils, but no immunohistochemical lesion was detected in mildly symptomatic gerbils. Cerebral blood flow was markedly reduced in severely symptomatic gerbils but more selectively reduced in the cortical structures of moderately symptomatic gerbils. This prediction method is useful for investigating early cerebral ischemia and for evaluating the effectiveness of pharmacologic agents.