Neuronal Damage In Rats Research Articles

Effects of transcranial focal electrical stimulation alone and associated with a sub-effective dose of diazepam on pilocarpine-induced status epilepticus and subsequent neuronal damage in rats

Experiments were conducted to evaluate the effects of transcranial focal electrical stimulation (TFS) applied via tripolar concentric ring electrodes, alone and associated with a sub-effective dose of diazepam (DZP) on the expression of status epilepticus (SE) induced by lithium-pilocarpine (LP) and subsequent neuronal damage in the hippocampus. Immediately before pilocarpine injection, male Wistar rats received TFS (300Hz, 200-μs biphasic square charge-balanced 50-mA constant current pulses for 2min) alone or combined with a sub-effective dose of DZP (0.41mg/kg, i.p.). In contrast with DZP or TFS alone, DZP plus TFS reduced the incidence of, and enhanced the latency to, mild and severe generalized seizures and SE induced by LP. These effects were associated with a significant reduction in the number of degenerated neurons in the hippocampus. The present study supports the notion that TFS combined with sub-effective doses of DZP may represent a therapeutic tool to induce anticonvulsant effects and reduce the SE-induced neuronal damage.

Evaluation of Drug Induced Neuronal Damage in Rats

Evaluation of Drug Induced Neuronal Damage in Rats

Hippocampal neuronal damage in rats exposed to a double hit: irradiation and hyperthermia.

In utero irradiation models induce diffuse neuronal damage. Experimental studies have shown that hyperthermia induced seizures are easily elicited and have high mortality accompanied by neuronal loss. Neuronal damage and loss are the results of cell death coupled with cortical development in altered cellular development. The aim of the study was to investigate the changes in hippocampus that was exposed to irradiation and hyperthermia. Four groups were studied: 1) The irradiation group was exposed to 225 cGy irradiation on the 17th gestational day; 2) The hyperthermia group was exposed to hyperthermia on the 10th postnatal day; 3) The hyperthermia plus irradiation group was exposed to in utero irradiation and postnatal hyperthermia; 4) The control group was sham operated. Animals were examined 3 and 6 months later. The hippocampus was atrophic with neuronal loss in CA regions and ectopic neurons were in irradiation group. Severe damage with the most atrophy was demonstrated in all regions of the irradiation plus hyperthermia group. In long term, damage was severe in all groups. This study demonstrated more damage in hippocampi exposed to both irradiation and hyperthermia that may be taken as an evidence for the double hit hypothesis in the development of hippocampal damage.

Hippocampal neuronal damage in rats exposed to a double hit: irradiation and hyperthermia

AIm: In utero irradiation models induce diffuse neuronal damage. Experimental studies have shown that hyperthermia induced seizures are easily elicited and have high mortality accompanied by neuronal loss. Neuronal damage and loss are the results of cell death coupled with cortical development in altered cellular development. The aim of the study was to investigate the changes in hippocampus that was exposed to irradiation and hyperthermia. mAterIAl and methOds: Four groups were studied: 1) The irradiation group was exposed to 225 cGy irradiation on the 17th gestational day; 2) The hyperthermia group was exposed to hyperthermia on the 10th postnatal day; 3) The hyperthermia plus irradiation group was exposed to in utero irradiation and postnatal hyperthermia; 4) The control group was sham operated. Animals were examined 3 and 6 months later. results: The hippocampus was atrophic with neuronal loss in CA regions and ectopic neurons were in irradiation group. Severe damage with the most atrophy was demonstrated in all regions of the irradiation plus hyperthermia group. In long term, damage was severe in all groups. COnClusIOn: This study demonstrated more damage in hippocampi exposed to both irradiation and hyperthermia that may be taken as an evidence for the double hit hypothesis in the development of hippocampal damage.

Fasudil mesylate protects PC12 cells from oxidative stress injury via the Bax-mediated pathway.

We previously reported that fasudil mesylate (FM) improves neurological deficit and neuronal damage in rats with ischemia following middle cerebral artery occlusion and reperfusion in vivo. In this study, the properties of FM on hydrogen peroxide (H(2)O(2))-induced oxidative stress insult in cultured PC12 cells as well as the underlying mechanisms were investigated in vitro. Pretreatment with FM (5, 10 μM) prior to H(2)O(2) exposure significantly elevated cell viability, reduced cell apoptosis by MTT assay, LDH assay, Hoechst 33258 dye staining, and FM also decreased the accumulation of reactive oxygen species (ROS) by DCFH-DA staining and NBT test. Furthermore, FM also reversed the upregulation of Bax/Bcl-2 ratio, the downstream cascade following ROS. FM protected PC12 cells from oxidative stress insult via downregulating the Bax/Bcl-2 ratio. These findings indicate that a direct effect of fasudil mesylate on PC12 cells may be partly responsible for its protective effect against oxidative stress injury.

Protective effect of Ilex latifolia, a major component of “kudingcha”, against transient focal ischemia-induced neuronal damage in rats

Ilex latifolia (Aquifoliaceae), a primary component of "kudingcha", has been used in Chinese folk medicine to treat various kinds of diseases including headaches, inflammatory diseases, and cardiac ischemic injury. The present study investigated the protective effect of the ethanol extract of Ilex latifolia against transient, focal, ischemia-induced neuronal damage. Transient focal ischemia was induced by 2 h middle cerebral artery occlusion followed by 24 h reperfusion (MCAO/reperfusion) in rats. After MCAO/reperfusion, brain infarction and neuronal death were measured by triphenyltetrazolium chloride and hematoxylin and eosin staining, respectively. Glutathione concentration and lipid peroxidation rate were measured. The expression levels of phosphorylated mitogen activated proteins kinases (MAPKs), cyclooxygenase 2 (COX-2), and anti-apoptotic and pro-apoptotic proteins were detected by Western blot. Ilex latifolia (50-200 mg/kg) significantly reduced MCAO/reperfusion-induced infarction and edema formation, neurological deficits, and brain cell death. Depletion of glutathione level and lipid peroxidation induced by MCAO/reperfusion were inhibited by administration of Ilex latifolia. The increase of phosphorylated MAPKs, COX-2, and proapoptotic proteins and the decrease of antiapoptotic protein in MCAO/reperfusion rats were significantly inhibited by treatment with Ilex latifolia. Ilex latifolia ameliorated ischemic injury induced by MCAO/reperfusion in rats, and this neuroprotective effect might be associated with its anti-apoptotic effect, resulting from anti-oxidative and anti-inflammatory actions.

Ameliorating effects of Kangen-karyu on neuronal damage in rats subjected to repeated cerebral ischemia

We previously reported that 21-day (14-day pre-ischemic and 7-day post-ischemic) treatment with Kangen-karyu (KGKR) improved spatial memory impairment and hippocampal neuronal death induced by repeated cerebral ischemia (2 x 10-min, 1-h interval) in rats. In the present study, we examined the effect of single and 14-day pre-ischemic KGKR treatment on neuronal damage in the same repeated cerebral ischemia model. Additionally, to determine the mechanisms of neuroprotection by KGKR at glutamatergic neurons, we examined the effects of KGKR on glutamate release induced by repeated cerebral ischemia in vivo, and on cell damage induced by both glutamate and kainate in primary cultured hippocampal neurons in vitro. The 14-day pre-ischemic KGKR (300 mg/kg, oral administration (p.o.)) treatment reduced neuronal damage and astrocyte expression induced by repeated cerebral ischemia. No effect was observed after single pre-ischemic KGKR treatment. Both single and 14-day KGKR treatment decreased glutamate release in the hippocampal CA1 region in intact rats; however, neither pre-ischemic KGKR treatment altered glutamate release during cerebral ischemia. In vitro, KGKR (100-1000 microg/mL) dose-dependently suppressed hippocampal neuronal damage induced by both glutamate (100 microM) and kainate (1 mM). These data suggest neuroprotection with KGKR requires continuous pre-ischemic treatment, and that the mechanisms of protection may be involved in inhibiting the glutamatergic receptors of the post-synaptic neurons.

Vasoregression Linked to Neuronal Damage in the Rat with Defect of Polycystin-2

BackgroundNeuronal damage is correlated with vascular dysfunction in the diseased retina, but the underlying mechanisms remain controversial because of the lack of suitable models in which vasoregression related to neuronal damage initiates in the mature retinal vasculature. The aim of this study was to assess the temporal link between neuronal damage and vascular patency in a transgenic rat (TGR) with overexpression of a mutant cilia gene polycystin-2.MethodsVasoregression, neuroglial changes and expression of neurotrophic factors were assessed in TGR and control rats in a time course. Determination of neuronal changes was performed by quantitative morphometry of paraffin-embedded vertical sections. Vascular cell composition and patency were assessed by quantitative retinal morphometry of digest preparations. Glial activation was assessed by western blot and immunofluorescence. Expression of neurotrophic factors was detected by quantitative PCR.FindingsAt one month, number and thickness of the outer nuclear cell layers (ONL) in TGR rats were reduced by 31% (p<0.001) and 17% (p<0.05), respectively, compared to age-matched control rats. Furthermore, the reduction progressed from 1 to 7 months in TGR rats. Apoptosis was selectively detected in the photoreceptor in the ONL, starting after one month. Nevertheless, TGR and control rats showed normal responses in electroretinogram at one month. From the second month onwards, TGR retinas had significantly increased acellular capillaries (p<0.001), and a reduction of endothelial cells (p<0.01) and pericytes (p<0.01). Upregulation of GFAP was first detected in TGR retinas after 1 month in glial cells, in parallel with an increase of FGF2 (fourfold) and CNTF (60 %), followed by upregulation of NGF (40 %) at 3 months.InterpretationOur data suggest that TGR is an appropriate animal model for vasoregression related to neuronal damage. Similarities to experimental diabetic retinopathy render this model suitable to understand general mechanisms of maturity-onset vasoregression.

Neuroprotective potential of fasudil mesylate in brain ischemia-reperfusion injury of rats.

We previously reported that inhibition of Rho-kinase (ROCK) by hydroxyl fasudil improves cognitive deficit and neuronal damage in rats with chronic cerebral ischemia (Huang et al., Cell Mol Neurobiol 28:757-768, 2008). In this study, fasudil mesylate (FM) was investigated for its neuroprotective potential in rats with ischemia following middle cerebral artery occlusion (MCAO) and reperfusion. The effect of fasudil mesylate was also studied in rat brain cortical and hippocampal slices treated with oxygen-glucose deprivation (OGD) injury. Gross anatomy showed that cerebral infarct size, measured with 2,3,5-triphenyltetrazolium chloride (TTC) staining, was significantly smaller in the FM-treated than in the non-FM-treated ischemic rats. In the brain regions vulnerable to ischemia of ischemic rats, fasudil mesylate was also found to significantly restore the enzyme protein expression level of endothelial nitric oxide synthase (eNOS), which was decreased in ischemia. However, it remarkably reduced the protein synthesis of inducible nitric oxide synthase (iNOS) that was induced by ischemia and reperfusion. In rat brain slices treated with OGD injury, fasudil mesylate increased the neuronal cell viability by 40% for cortex and by 61% for hippocampus, respectively. Finally, in the presence of OGD and fasudil mesylate, superoxide dismutase (SOD) activity was increased by 50% for cortex and by 58% for hippocampus, compared to OGD only group. In conclusion, our in vivo study showed that fasudil mesylate not only decreased neurological deficit but also reduced cerebral infarct size, possibly and at least partially by augmenting eNOS protein expression and inhibiting iNOS protein expression after ischemia-reperfusion.

Blood-Brain Barrier Permeability and Nerve Cell Damage in Rat Brain 14 and 28 Days After Exposure to Microwaves from GSM Mobile Phones

We investigated the effects of global system for mobile communication (GSM) microwave exposure on the permeability of the blood-brain barrier and signs of neuronal damage in rats using a real GSM programmable mobile phone in the 900 MHz band. Ninety-six non-anaesthetized rats were either exposed to microwaves or sham exposed in TEM-cells for 2 h at specific absorption rates of average whole-body Specific Absorption Rates (SAR) of 0.12, 1.2, 12, or 120 mW/kg. The rats were sacrificed after a recovery time of either 14 or 28 d, following exposure and the extravazation of albumin, its uptake into neurons, and occurrence of damaged neurons was assessed. Albumin extravazation and also its uptake into neurons was seen to be enhanced after 14 d (Kruskal Wallis test: p = 0.02 and 0.002, respectively), but not after a 28 d recovery period. The occurrence of dark neurons in the rat brains, on the other hand, was enhanced later, after 28 d (p = 0.02). Furthermore, in the 28-d brain samples, neuronal albumin uptake was significantly correlated to occurrence of damaged neurons (Spearman r = 0.41; p < 0.01).

Improvement of cognitive deficit and neuronal damage in rats with chronic cerebral ischemia via relative long-term inhibition of rho-kinase.

(1) The role of activation of Rho-kinase in the pathogenesis of cognitive deficit and neuronal damage caused by chronic global ischemia is not clear. In this study, hydroxyfasudil, a Rho-kinase inhibitor, was found to improve the learning and memory performance significantly in rats with ischemia induced by chronic cerebral hypoperfusion after permanent bilateral carotid artery ligation (BCAL). This was observed by the administration of hydroxyfasudil (1 mg/kg or 10 mg/kg, once per day for 30 days) to ischemic rats and the measurements of escape latency and time spent in the target quadrant among the ischemic, sham, and ischemic plus hydroxyfasudil rats by the method of Morris water maze. (2) In electrophysiological study, hydroxyfasudil abolished the inhibition of long-term potentiation (LTP) in rats with ischemia. Morphologically, it also markedly reduced pathological changes such as neuronal cells loss and nuclei shrinkage in cortex and hippocampus of ischemic rats. Biochemical analysis showed that the inhibition of Rho-kinase by hydroxyfasudil reduced the amount of MDA and increased the activities of SOD and GPx in ischemic rats that had increased MDA and decreased SOD and GPx activities. (3) To explore mechanism (s) of the beneficial effects of hydroxyfasudil in ischemia, we performed immunohistochemistry and RT-PCR analyses of NMDA NR2B subunit and for the first time found that hydroxyfasudil increased the expression of NR2B in cortex and hippocampus at both protein and mRNA levels. (4) Taken together, our data further support the notion that the inhibition of Rho-kinase provides neuroprotective effects in cerebral ischemia.

Effects of Amelioration of Total Flavonoids from Stems and Leaves of Scutellaria baicalensis Georgi on Cognitive Deficits, Neuronal Damage and Free Radicals Disorder Induced by Cerebral Ischemia in Rats

Previous studies reported that the total flavonoids from the stems and leaves of Scutellaria baicalensis Georgi (TFSS) could enhance and improve learning and memory abilities in experimental animals, and reduce the neuronal pathologic alterations induced by some reagents in mice. The present study examined whether TFSS can improve memory dysfunction, neuronal damage, and abnormal free radicals induced by permanent cerebral ischemia in rats. The permanent cerebral ischemic model in rats was produced by bilateral ligation of the common carotid arteries. The influence of permanent cerebral ischemia on learning and memory was determined in the Morris water maze. The neuronal damage in the hippocampus and cerebral cortex was assessed by the neuronal morphologic observations. The contents of malondialdehyde (MDA) and nitric oxide (NO), and the activities of superoxide dismutase (SOD) and catalase (CAT) in the hippocampus and cerebral cortex were measured using thiobarbituric acid, nitrate reductase, xanthine-xanthine oxidase, and ammonium molybdate spectrophotometric methods, respectively. In learning and memory performance tests, cerebral ischemic rats always required a longer latency time to find the hidden platform and spent a shorter time in the target quadrant in the Morris water maze. TFSS 17.5-70 mg.kg(-1) daily orally administered to ischemic rats for 20 d, from day 16-35 after operation differently reduced the prolonged latency and increased swimming time spent in the target quadrant. In neuronal morphologic observations, daily oral TFSS 17.5-70 mg.kg(-1) for 21 d, from day 16-36 after operation markedly inhibited the ischemia-induced neuronal damage. In addition, the increased contents of MDA and NO, and SOD activity, and the decreased activity of CAT in the hippocampus and cerebral cortex induced by cerebral ischemia were differently reversed. The reference drug piracetam (140 mg.kg(-1) per day for 20-21 d) similarly improved impaired memory and neuronal damage but had no significant effects on free radicals in ligated rats. TFSS can improve memory deficits and neuronal damage in rats after permanent cerebral ischemia, which may be beneficial in the treatment of cerebrovascular dementia.

Evaluation of neuronal damage following hypoxic–ischaemic brain injury in acute and early chronic periods in neonatal rats

This study was undertaken to investigate the effects of neonatal cerebral hypoxic-ischaemic brain injury (HIBI) in acute and early chronic phases in the rat. HIBI was induced in 7-day-old rat pups by ligation of the right common carotid and then the pups were exposed to 1 h of hypoxia in 8% oxygen. They were divided into two groups: 1-day (acute phase, in the first 24 h) and 5-day (early chronic phase, 120 h). Neuropathological evaluation was performed using the hippocampus, cerebral cortex and basal ganglia on the coronal plane. The following values were obtained: (i) the ratio of the infarcted area; (ii) hemispheric atrophy/asymmetry; (iii) patchy lesions confined to the thalamus, caudate and putamen; (iv) the ratio of damaged neurons to all neurons; and (v) the percentage of apoptotic neurons relative to the total neurons in all brain areas. HIBI-induced global cerebral damage and cellular damage findings did not significantly differ between the two groups. However, they showed a tendency to recover/deteriorate in both acute and early chronic phases. The ratio of ipsi- and contra-lateral hemisphere infarct areas (20.7 and 15.7% vs. 40.1 and 26.7%, respectively), basal ganglia patchy lesion ratio (27.5 vs. 36.7%) and hemispheric atrophy/asymmetry (92.4 vs. 84.7%) were found to be lower in the rat pups in the chronic phase than those in the acute phase. In contrast, increases in the ratio of damaged neurons (16.7 vs. 13.3% in the cerebral and dorsal hippocampus, respectively) and in the ratio of apoptotic neurons (ipsi-lateral: 18 vs. 6%; contra lateral hemispheres: 3.5 vs. 1.7%, respectively) were recorded. It is concluded that cellular damage tends to deteriorate (damaged and apoptotic neurons) while global damage (cerebral infarct and patchy damage) improves with the progression of HIBI. However, further studies are needed in order to elucidate this process.

Melatonin protects against neuronal damage induced by 3-nitropropionic acid in rat striatum

In this study, the protective effects of melatonin were evaluated against 3-nitropropionic acid (3-NP)-induced striatal neuronal damage in rats. Lesions were induced in the right striatum of Sprague–Dawley rats by stereotaxic injection with 3-NP and melatonin was intraperitoneally administered both 30 min before and 60 min after 3-NP injection. And rats continuously received melatonin daily for 3 days. As indicators of oxidative damage, lipid peroxidation and protein oxidation in the lesioned striatum were measured at 1 day after 3-NP injection. Levels of malondialdehyde (MDA) and protein carbonyl were significantly increased by 3-NP injection, but reduced in the melatonin-treated rats. Four days post-lesion, large lesions and extensive neuronal damage were produced in the 3-NP-injected striata, as revealed by 2,3,5-triphenyltetrazolium chloride (TTC) staining. In addition, marked ipsilateral rotational behavior following apomorphine challenge and a decrease of dopamine content in the lesioned striatum were observed in the 3-NP-injected rats. However, melatonin treatment significantly attenuated the 3-NP-induced neuronal damage, reduced the degree of asymmetric rotational behavior, and restored the dopamine level in the lesioned striatum. The present results indicate that melatonin effectively protects against the neuronal damage caused by 3-NP in vivo and that the neuroprotective effects of melatonin may be related to antioxidant action.

Sevoflurane-induced preconditioning protects against cerebral ischemic neuronal damage in rats

In the present study, we tested the ability of sevoflurane to induce early and late preconditioning against ischemic neuronal injury using an in vivo model of global cerebral ischemia in the rat. Seven-minute global ischemia was induced by cardiac arrest, followed by resuscitation and recovery for 7 days. Hippocampal slices were then prepared and the amplitude of extracellularly recorded, orthodromically evoked, CA1 population spikes (neuronal function) was quantified. Rats were preconditioned for 30 min with 1.0 minimum alveolar concentration (MAC) of sevoflurane once or on 4 consecutive days, 15 min (single exposure, early) or 24 h (four exposures, late preconditoning) prior to cardiac arrest. After early or late preconditioning, sevoflurane reduced ischemic neuronal damage from 43 ± 3% [sham rats, (mean ± SEM)] to 30 ± 3% and 35 ± 4%, respectively. Histopathology demonstrated a preserved morphology of the CA1 region of preconditioned rats, whereas pyknosis was present in control and sham-treated rats. Sevoflurane-induced preconditioning confers neuroprotection during an early as well as late time window.

Protective Effect of Buckwheat Polyphenols Against Long-Lasting Impairment of Spatial Memory Associated With Hippocampal Neuronal Damage in Rats Subjected to Repeated Cerebral Ischemia

In the present experiment, we studied the action of buckwheat polyphenol (BWP, from Fagopyrum esculentum MOENCH) in a repeated cerebral ischemia model, which induced a strong and long-lasting impairment of spatial memory in 8-arm radial maze with hippocampal CA1 cell death in rats. BWP (600 mg/kg, continuous 21-day p.o.) significantly ameliorated not only the impairment of spatial memory in the 8-arm radial maze, but also necrosis and TUNEL-positive cells in the hippocampal CA1 area subjected to repeated cerebral ischemia (10 min × 2 times occlusion, 1-h interval) in rats. In order to investigate the mechanism of BWP protective action, we measured the release of glutamate and NOx- (NO2- +NO3-) production induced by repeated cerebral ischemia in the rat dorsal hippocampus using microdialysis. A 14-day BWP treatment significantly inhibited the excess release of glutamate after the second occlusion. In addition, the BWP remarkably suppressed a delayed increase in NOx- (NO2- +NO3-) induced by repeated cerebral ischemia in the dorsal hippocampus as determined in vivo by microdialysis. However, the 14-day treatment did not affect hippocampal blood flow in either intact rats or rats subjected to repeated ischemia measured by lasser Doppler flowmeter. These results suggested that BWP might ameliorate spatial memory impairment by inhibiting glutamate release and the delayed generation of NOx- in rats subjected to repeated cerebral ischemia.

Effect of lamotrigine treatment on status epilepticus-induced neuronal damage and memory impairment in rat

Status epilepticus causes neuronal damage that is associated with cognitive impairment. The present study examined whether a novel antiepileptic drug, lamotrigine (LTG), alleviates status epilepticus-induced temporal lobe damage and memory impairment, and compared its efficacy with carbamazepine. Status epilepticus was induced by electric stimulation of the perforant pathway (PP) in rats. Treatment with LTG (12.5 mg/kg, twice a day) was started either 3 days before (preLTG group) or 1 h after (postLTG group) a 60 min PP stimulation. Treatment with carbamazepine (CBZ; 30 mg/kg, twice a day) was started 3 days before (CBZ group) a 60 min PP stimulation. All treatments were continued for 2 weeks. Thereafter, the severity of seizures, seizure-induced neuronal damage, quantitative electroencephalogram (EEG), and memory impairment were compared between vehicle-treated unstimulated and stimulated controls, LTG-treated rats, and CBZ-pretreated rats. Both in the preLTG and postLTG groups, damage to hilar somatostatin-immunoreactive neurons, hippocampal CA3b and CA3a pyramidal cells, and the piriform cortex was mild and did not differ from that in unstimulated controls. Furthermore, CA3c damage in the preLTG group did not differ from that in unstimulated controls. Vehicle-treated stimulated controls and CBZ-pretreated rats, however, had significant damage in the hilus, CA3 subregions, and piriform cortex compared with unstimulated controls ( P<0.05 for the stimulated side, contralateral side, or both). Treatment with LTG or CBZ had no effect on the number or duration of behavioral seizures during PP stimulation. They did not affect the baseline EEG or status epilepticus-induced slowing of the EEG. Also, the status epilepticus-induced spatial memory impairment in the Morris water-maze was not attenuated by treatment with LTG or CBZ. Our data demonstrate that treatment with LTG has a mild neuroprotective effect on status epilepticus-induced neuronal damage in rats even when administered after the beginning of status epilepticus.

Evaluation of cardiac adrenergic neuronal damage in rats with doxorubicin-induced cardiomyopathy using iodine-131 MIBG autoradiography and PGP 9.5 immunohistochemistry.

Doxorubicin is one of the most useful anticancer agents, but its repeated administration can induce irreversible cardiomyopathy as a major complication. The purpose of this study was to investigate doxorubicin toxicity on cardiac sympathetic neurons using iodine-131-metaiodobenzylguanidine (MIBG) and protein gene product (PGP) 9.5 immunohistochemistry, which is a marker of cardiac innervation. Wistar rats were treated with doxorubicin (2 mg/kg, i.v.) once a week for 4 (n=5), 6 (n=6) or 8 (n=7) weeks consecutively. Left ventricular ejection fraction (LVEF), calculated by M-mode echocardiography, was used as an indicator of cardiac function. Plasma noradrenaline (NA) concentration was measured by high-performance liquid chromatography (HPLC). 131I-MIBG uptake of the left ventricular wall (24 ROIs) was measured by autoradiography. 131I-MIBG uptake pattern was compared with histopathological results, the neuronal population on PGP 9.5 immunohistochemistry and the degree of myocyte damage assessed using a visual scoring system on haematoxylin and eosin and Masson's trichrome staining. LVEF was significantly decreased in the 8-week group (P<0.05). The serum NA level also showed no statistical difference until 4 weeks and was significantly increased in the 8-week group (P<0.05). MIBG uptake was decreased in the 6- and 8-week groups (P<0.05), and was closely correlated with the reduction in the number of nerve fibres on PGP 9.5 stain. Myocyte damage was seen only in the 8-week group. Neuronal population and the 131I-MIBG uptake ratio of subepicardium to subendocardium were significantly increased (P<0.05) in the 8-week group as compared with the control group. It may be concluded that radioiodinated MIBG is a reliable marker for the detection of cardiac adrenergic neuronal damage in doxorubicin-induced cardiomyopathy; it detects such damage earlier than do other clinical parameters and in this study showed a good correlation with the reduction in the neuronal population on PGP 9.5 stain. The subendocardial layer appeared to be more vulnerable to doxorubicin than the subepicardium.

Preischemic blood glucose supply to the brain modulates HSP 72 synthesis and neuronal damage in gerbils

Preischemic hyperglycemia is known to aggravate brain damage caused by transient forebrain ischemia. Because heat shock proteins (HSPs) 72 have been proposed to play a protective role against ischemic neuronal injury, we studied the HSP 72 mRNA expression and protein synthesis in gerbils subjected to 10 min bilateral carotid occlusion under normoglycemic, hyperglycemic and fasting conditions. HSP 72 mRNA expression and HSP 72 synthesis were studied using in situ hybridization and immunostaining, respectively. After 8 h of blood recirculation, HSP 72 mRNAs were expressed in all the hippocampal subfields of the three different groups, with higher expression in the hyperglycemic gerbils. After 48 h of reperfusion, HSP 72 mRNAs had almost completely disappeared in the hyper- and normoglycemic groups, and were more strongly expressed in the CA 1 neurons of the fasted group. At this time, fasted gerbils exhibited intense HSP 72 immunoreactivity in the CA 1, whereas an absence of immunoreactivity was observed in that area in the other groups. Finally, ischemia was also associated with marked astrocytic activation, as evidenced by GFAP immunostaining. Overall results indicate that preischemic differences in blood glucose supply to the brain are related to HSP 72 mRNA expression (in terms of duration) and to HSP 72 protein induction (in terms of intensity) in the vulnerable CA 1 neurons of the hippocampus. Ability of CA 1 neurons to synthesize HSP 72 proteins was associated with higher neuronal survival in the fasted group after 48 h of reflow, suggesting a protective role of HSP 72, even though evaluation of neuronal damage at 7 days indicated that neuronal death was mainly delayed in the time.

Ciliary neurotrophic factor protects hippocampal neurons from excitotoxic damage

The loss of neurons is responsible for many acute neurological disorders as well as chronic neurodegenerative diseases. This cell loss might be prevented by a direct delivery of neurotrophic factors. Therefore, we investigated the capacity of ciliary neurotrophic factor (CNTF) and nerve growth factor (NGF) as well as the combination of both growth factors on the glutamate-induced excitotoxic damage in hippocampal cultures. The exposure of hippocampal neuronal/glial co-cultures to 0.5 mM l-glutamate for 1 h induced pronounced neurotoxicity evaluated 18 h later by trypan blue staining and morphological criteria. The damaged neurons showed both apoptotic and necrotic features. However, CNTF (1–1000 pg/ml) reduced neuronal degeneration when administered 6 and 24 h before induction of injury and remained in contact with the cells until evaluation of neuronal damage. Furthermore, NGF (1 ng/ml) also rescued the hippocampal neurons under the same experimental conditions and with a similar to CNTF potency. However, the co-administration of NGF and CNTF (but not either factor alone) restored the neuronal survival to control levels. Our results support the hypothesis that administering neurotrophic factors could represent an alternative strategy for the treatment of acute and chronic brain disorders.