Neuroprotection by intrathecal application of liposome-entrapped fasudil in a rat model of ischemia.

Pharmacological treatment for cerebral ischemia cannot attain sufficiently high concentrations of the drugs in the cerebrospinal fluid (CSF) without precipitating systemic side effects. The objective of this study is the development of a liposomal drug delivery system that maintains effective concentrations of protein kinase inhibitors fasudil in the CSF, resulting in neuroprotection against cerebral ischemia. Focal cerebral ischemia in rats was induced by middle cerebral artery occlusion using an intraluminal suture technique. Treated rats received 0.25 mg liposome-entrapped fasudil via the cisterna magna 2 hours after ischemic insult. Control rats received drug-free liposomes. Neurological condition and the infarct size were assessed at 24 and 72 hours after ischemia. The concentration of liposome-entrapped fasudil in the CSF was measured before sacrifice. Treated animals showed significantly improved neurological outcomes after the 24-hour observation period compared to the control group (p < 0.001). Treatment with 0.25 mg liposomal fasudil resulted in a reduction in the infarct area (24 hours: 29.0 +/- 4.4%, 72 hours: 28.1 +/- 3.9% of total brain slices) compared to controls (49.6 +/- 4.6%, p < 0.001), but there was no statistical difference between 24 and 72 hours. At 24 hours post-administration, CSF concentrations of liposome-entrapped fasudil were 45.4 +/- 31.5 micrograms/ml (20% of the injected dose). A single intrathecal injection of liposomal fasudil can maintain a therapeutic drug concentration in the CSF over a period of time, significantly decreasing infarct size in a rat model of acute ischemia.

Vertebral artery dissection: warning symptoms, clinical features and prognosis in 26 patients.

BACKGROUND AND OBJECTIVES: Internal carotid artery dissection has been extensively studied and well-described. Although there has been a recent increase in the number of reported cases of vertebral artery (VA) dissection, the clinical variety of presentation and the early warning symptoms have not been well-described before. Our objectives in this study include: (1) To determine the early symptoms and warning signs which may help the clinician in the early identification and treatment of patients with VA dissection. (2) To explore the variety of clinical presentation of VA dissection and its relation to prognosis. DESIGN AND SETTING: Retrospective analysis of hospital records in a tertiary academic centre for the period 1989-1999. RESULTS: Twenty-six patients were identified (13 men and 13 women). The mean age was 48. Possible precipitating factors were identified in 14 patients (53%). Sporting activity and chiropractic manipulations were the most common (15% and 11% respectively). Headache and/or neck pain was the prominent feature in 88% of patients and was a warning sign in 53%, preceding onset of stroke by up to 14 days. The most common clinical features included vertigo (57%), unilateral facial paresthesia (46%), cerebellar signs (33%), lateral medullary signs (26%) and visual field defects (15%). Bilateral VA dissection presented in six patients (24%). The most common region of dissection was the C1-C2 level (16 arteries, 51%). Intracranial VA dissection was found in eight arteries (25%). The majority of patients (83%) had favorable outcome. Poor prognosis was associated with (1) bilateral dissection; (2) intracranial VA dissection accompanied by subarachnoid hemorrhage. Only two patients reported stroke recurrence. CONCLUSIONS: Our findings show that VA dissection affects mainly middle age persons and involves both sexes equally. Headache and/or neck pain followed by vertigo or unilateral facial paresthesia is an important warning sign that may precede onset of stroke by several days. Although the majority of patients will have excellent prognosis, this was less likely in patients presenting with subarachnoid hemorrhage or bilateral VA dissection. Recurrence rate was low.

The role of taurine in neuronal protection following transient global forebrain ischemia.

Osmoregulation and post ischemic glutamate surge suppression (PIGSS) are important mechanisms in the neuroprotective properties of taurine. We studied the role of taurine in PIGSS following transient global forebrain ischemia (TGFI). A group of gerbils received a high dose of continuous intracerebral taurine during the peri-ischemic period. Beta-alanine was given similarly to a negative control group. The control group consisted of animals undergoing only TGFI. On the fourth day following commencement of drug administration, TGFI was induced. Concurrently, half the animals from each group receiving an agent had intracerebral microdialysis. All animals underwent histological assessment at day 7. The microdialysis and histological data was analyzed. Our results showed that taurine treatment did not cause PIGSS. The histological difference between the three groups was statistically insignificant. We conclude that intracerebral taurine in the dosage administered during peri-ischemic period, does not result in PIGSS or histologically evident neuroprotection.

Attenuated glutamate release during ischemia in ethanol-administered gerbils.

Previous studies have found an association between prior ethanol consumption and aggravated stroke outcome. Gerbils were intermittently given ethanol injections (s.c.) for 21 days at doses of 1 and 4 g/kg. After cessation of injections and appropriate weight gain, subjects underwent bilateral carotid occlusion while amino acid neurotransmitter levels in the hippocampus were monitored. Both the low and high dose ethanol groups demonstrated significantly decreased glutamate release compared with saline-treated controls during ischemia (p < 0.05). These results are consistent with a long-lasting ethanol-induced decrease in synaptic density in the hippocampus. That no intergroup differences on histological or neurobehavioral measures was found may suggest a functional dissociation of glutaminergic involvement in the pathogenesis of aggravated stroke outcome with alcoholism.

Neuroprotection with felbamate: a 7- and 28-day study in transient forebrain ischemia in gerbils.

The use of glutamate antagonists and GABA agonists may protect neurons from the effects of transient ischemia. Felbamate is a new antiepileptic drug with glutamate antagonist and GABA agonist properties. We tested the efficacy of felbamate in a gerbil model of transient forebrain ischemia. Damage assessment was done with silver staining at 7 and 28 days after 5 min of bilateral carotid occlusion. Cerebral cortex, hippocampus (CA1 and CA4), thalamus and striatum were evaluated on a 4-point scoring system. The animals sacrificed at 28 days were also tested in a water-maze task to assess recovery of function. The initial dose of felbamate (300 mg/kg) was given 30 min before the ischemic insult in one set of animals and 30 min after the insult in another set of animals. There were 8 animals tested per group (total: 48 animals). There was significant neuronal protection with the use of felbamate, both before and after ischemia in all regions of the brain. Protection was seen in animals sacrificed at 7 and 28 days. Protection was moderate when felbamate was used before ischemia. It was highly significant when felbamate was given 30 min after the insult. Behavioral studies however did not show any difference in the felbamate treated animals versus the saline treated controls. The structural protection with felbamate was very significant when used in the post-ischemic period. This window for protection merits further evaluation in relation to the clinical setting of stroke.

Acetyl-L-carnitine attenuates neuronal damage in gerbils with transient forebrain ischemia only when given before the insult.

The underlying mechanisms leading to neuronal damage in cerebral ischemia are multifactoral. In this study, we evaluated the neuroprotective effects of acetyl-L-carnitine, a medication that may enhance metabolic recovery after cerebral ischemia. The 5-minute transient forebrain ischemia model in gerbils was used. Acetyl-L-carnitine was given 30 minutes before the insult in one set of animals and 30 minutes after the insult in a second set of animals with histological evaluation at 7 days (Group A) and 28 days (Group B). Damage assessment was done using a 4-point damage score and Mann-Whitney U test was used for statistical analysis. Compared to the controls, there was significant protection in the cerebral cortex, hippocampus and the striatum in animals treated with the medication before the insult in Group A and Group B. Post-ischemic therapy showed little evidence of neuronal protection in either group. Behavioral tests in the Group B animals showed no significant differences between the treated or the saline controls. Our study shows, that pre-ischemic treatment with acetyl-L-carnitine results in neuronal protection. This may have clinical significance in situations (such as bypass surgery) where treatment could be initiated prior to the insult.

Baclofen is cytoprotective to cerebral ischemia in gerbils.

The release of the neurotransmitter, glutamate, and the activation of receptor operated calcium channels, may increase the degree of damage in ischemic brain tissue. Inhibition of excitatory neurotransmitters should therefore result in cytoprotection of ischemic brain tissue. In this study we evaluated the effect of baclofen, an inhibitor of presynaptic glutamate release, on ischemic gerbil cortex, hippocampus (CA 1 and CA4), striatum and thalamus. Histological evaluation was done in a blind manner in 4 groups (total 36 animals): a control group (9 animals) and three groups (27 animals) with varying doses of baclofen. For cerebral ischemia, we used single episode of five minutes of arterial occlusion of the carotid arteries. Baclofen in doses of 0, 25, 50, and 100 mg/kg were given to different groups five minutes prior to ischemic insult. This was followed by intraperitoneal injections given 24 and 48 hours after the initial insult. Statistically significant histological cytoprotection was demonstrated. Doses of 25 mg/kg appeared to demonstrate significant protection of the cortex (p = 0.0002), the CA1 and CA4 regions of the hippocampus (p = 0.0004 and 0.0001) respectively. At a dose of 50 mg/kg, significant cytoprotection was demonstrated at the hippocampus (CA1 and CA4 regions), in particular at the CA4 region (p = 0.0029). The 100 mg/kg dose appeared to have most significant protection at the CA1 and CA4 regions of the hippocampus (both p = 0.0001), striatum (p = 0.0011), and the thalamus (p = 0.0008). All statistical comparisons were done using non-parametric tests (Mann-Whitney U test). Our study demonstrates that baclofen is cytoprotective to ischemic neuronal cells, especially in the hippocampus.(ABSTRACT TRUNCATED AT 250 WORDS)

Neuronal protection with superoxide dismutase in repetitive forebrain ischemia in gerbils.

The underlying mechanism for severe damage with repetitive ischemia is not fully understood. Because of prolonged periods of reperfusions between the brief insults, we speculated that the severe damage may be secondary to excessive generation of oxygen free radicals. In this study we tested the efficacy of peg-superoxide dismutase (SOD) in a model of repeated ischemia in gerbils. Superoxide dismutase (SOD) or vehicle (saline) was delivered through osmotic pumps into the lateral ventricles continuously from the onset of the insult until the gerbils were sacrificed 6 days later. Three doses of SOD were used in the experiments (110, 150, and 190 units per microliter). Damage was assessed using a 0-4 point scoring system and statistical comparisons were done using the Mann-Whitney U-test. There was significant protection in the hippocampus (p < 0.05), striatum (p < 0.001), and substantia nigra reticulata (p < 0.05) in the lowest dose SOD-treated group (110 units per microliter). Animals treated with 150 units showed lesser (but significant) protection in the thalamus, medial geniculate nucleus, and striatum. In the animals treated with the higher dose of SOD (190 units per microliter), the extent of damage was no different than vehicle-treated controls in the cortex, striatum, and hippocampus. Compared to controls, neuronal damage was, however, significantly more severe in the medial geniculate nucleus and the thalamus in the high-dose SOD-treated animals (p < 0.05). Our experiments suggest that the SOD may have a small therapeutic window. Higher doses may either have no neuroprotective effects or may be harmful.

Hypothyroidism protects the brain during transient forebrain ischemia in gerbils.

The mechanisms by which brain cells die after brief episodes of cerebral ischemia are not fully understood. In certain brain regions this damage may not be apparent for days. Hypothyroidism is known to decrease cerebral metabolism. We postulated that this slowing in cerebral metabolism may be neuroprotective after transient cerebral ischemia. To test this hypothesis, a total of 10 gerbils had thyroidectomies performed 2 weeks prior to ischemia. Six gerbils served as euthyroid controls. All animals were exposed to 5 min of transient ischemia and sacrificed 7 days after the insult. Silver degeneration staining was used for histological evaluation. Hippocampal damage [subiculum (P < 0.001), CA1 (P = 0. < .001), CA3 (P < 0.05), and CA4 (P < 0.001)] was significantly less in the hypothyroid animals. There was also significantly less damage in the cerebral cortex (P < 0.05) and thalamus (P < 0.05) in the hypothyroid animals. The exact mechanism of this protection is not fully understood but could be secondary to a decrease in the metabolic activity, or a reduced generation of free radicals (as is seen with protection from ischemia in kidney and liver under hypothyroid conditions). Further studies are required in order to gain a better understanding of the protective effects of hypothyroidism on cerebral ischemia.

GABA agonist "muscimol" is neuroprotective in repetitive transient forebrain ischemia in gerbils.

The damaging effects from transient forebrain ischemia may be a result of excessive excitability or loss of inhibitory influences. In the brain, GABA acts as the major inhibitory neurotransmitter and its loss may be an important factor leading to delayed neuronal damage in the substantia nigra reticulata (SNr). In this study, we looked at the protective effects of muscimol, a GABA A agonist in a gerbil model of repetitive forebrain ischemia. For cerebral ischemia, we used three episodes of 2 min with a reperfusion period of 1 h between the insults. Histological evaluations were done 7 days after the insult using silver degeneration staining. Muscimol was infused into the third ventricle continuously for 7 days beginning just prior to the insult. There were a total of 20 animals, 12 treated with muscimol and the other 8 serving as controls. At 7 days, there was significant protection in the cortex (P = 0.007), hippocampus [CA1 (P = 0.01), CA4 (P = 0.015)], substantia nigra reticulata (P = 0.007), striatum (P = 0.049), and thalamus (P = 0.012). All statistical comparisons were done using nonparametric tests (Mann-Whitney U test). Our study shows that potentiation of inhibitory mechanisms may be important mechanisms of neuronal protection from the effects of repetitive ischemia and the effects are not limited to the SNr. Further studies are needed to better understand their mechanism of action.