Tamar Kadar

Age-related structural changes in the rat hippocampus: correlation with working memory deficiency.

Age-related histopathological changes in the hippocampal formation were correlated with cognitive performance, evaluated in rats at the 8-arm radial maze. Experiments were conducted using young (3 months), mature (12 months), middle-aged (17 months) and aged (24 months) Wistar rats. Significant memory impairments were already observed at the age of 12 months in all the measured parameters (correct choices, percent errors and total time). No further decline was observed between 12 and 17 months of age, while at 24 months additional decline was monitored mainly in the percent errors parameter. Morphometric analysis revealed a decrease in the area of cells within the hippocampus and the number of cells in the CA3 subfield. This pattern of morphological changes with age corresponded well with the cognitive impairments, with high correlation especially to lesions at the CA3 subfield. It had also been confirmed in this study that lipofuscin appeared to be a good histochemical marker for CNS cell degeneration. It is concluded that 12-month-old Wistar rats may serve as the animal model of choice for the study of specific age-related behavioral deficits and that the hippocampal CA3 region might play a major role in the age-dependent cognitive decline.

Cognitive deficits induced in young rats by long-term corticosterone administration

Corticosterone slow-release pellets, implanted for 9 weeks in young Fischer 344 rats, resulted in continuous high plasma levels of the hormone which are comparable to those of rats under mild stress. One week following termination of the drug treatment, the rats were tested in an eight-arm radial maze. During the initial acquisition stages, corticosterone-treated rats exhibited cognitive impairments in contrast to placebo-treated rats. The deficits were observed in all three parameters which were monitored, the total number of errors, the number of correct entries out of the first eight, and the total time needed to complete the test. This study is the first to report specific behavioral decrements related to the previously observed morphological hippocampal changes induced by long-term corticosterone administration.

Sarin-induced neuropathology in rats

Sarin, a highly toxic cholinesterase (ChE) inhibitor, administered at near 1 LD50 dose causes severe signs of toxic cholinergic hyperactivity in both the peripheral and central nervous systems (CNS). The present study evaluated acute and long-term neuropathology following exposure to a single LD50 dose of sarin and compared it to lesions caused by equipotent doses of soman described previously. Rats surviving 1 LD50 dose of sarin (95 micrograms/kg; IM), were sacrificed at different time intervals post exposure (4 h-90 days) and their brains were taken for histological and morphometric study. Lesions of varying degrees of severity were found in about 70% of the animals, mainly in the hippocampus, piriform cortex, and thalamus. The damage was exacerbated with time and at three months post exposure, it extended to regions which were not initially affected. Morphometric analysis revealed a significant decline in the area of CA1 and CA3 hippocampal cells as well as in the number of CA1 cells. The neuropathological findings, although generally similar to those described following 1 LD50 soman, differed in some features, unique to each compound, for example, frontal cortex damage was specific to soman poisoning. It is concluded that sarin has a potent acute and long-term central neurotoxicity, which must be considered in the design of therapeutic regimes.

NG-nitro-L-arginine enhances neuronal death following transient forebrain ischemia in gerbils.

Experiments were performed with Mongolian gerbils to study the effect of the specific nitric oxide (NO) synthase inhibitor NG-nitro-L-arginine (L-NNA) on ischemic brain damage induced by 5 min bilateral carotid occlusion. A single i.p. injection of L-NNA did not result in any neuronal loss in the central nervous system. In animals undergoing ischemia, a selective destruction of hippocampal CA1 cells was observed whereas pretreatment with 50 mg/kg L-NNA 4 h before administration of ischemia produced significantly more extensive cell damage in the hippocampus and other brain regions. These findings demonstrate that in this model inhibition of nitric oxide generation augments ischemia-induced neuronal cell injury in the brain.

Aging has a complex effect on a rat model of ischemic stroke.

Stroke in humans is usually associated with advanced age. Nevertheless, almost all animal models of ischemic stroke are based on young animals. The present study was designed to assess the effect of age on the development of ischemic injury in a model of focal brain ischemia in rats. Two age groups of Wistar rats were used: young adult (3 months) and old (24-26 months). Under halothane anesthesia, polyethylene microspheres (50 microm in diameter) were injected into the left common carotid artery following a temporary occlusion of the external carotid artery. Sham-operated rats underwent the same procedure but were injected with an identical volume (100 microl) of saline only. Rats of both experimental groups displayed neurological impairment after surgery. However, contrary to expectation, the young rats were more affected than the old rats. Young rats displayed an abrupt 30% decrement in neurological functions in the first week and then showed a partial functional recovery into a 12% decrement from the second week on. Old rats developed the neurological impairment gradually over a 2-week period (6.3% in the first week and 11% in the second week and thereafter). One month later, rats were tested in a water maze task. Again, performance was more impaired in the young ischemic rats than in the old rats. Histological evaluation revealed more extensive neurological damage in young ischemic as compared to old rats. Thus, although increased age has a critical effect on the evolution of the neurological impairment following focal brain ischemia and stroke, its effects in the rat model were more pronounced in the young animals.

Sub-regional hippocampal vulnerability in various animal models leading to cognitive dysfunction.

Summary. Various animal models, involving different brain insults, lead to memory deficits, which can be measured using behavioral tests. In numerous studies, using five different experimental models in rats, we have found that cognitive dysfunction is invariably accompanied by hippocampal CA1 and CA3 pyramidal cells degeneration. However, of these two, the most affected area changes from one model to the other. The present manuscript describes and compares the morphological alterations within the hippocampus in the following experimental models: normal aging, hypoxia, prolonged corticosterone administration, brain ischemia and cholinesterase (ChE) inhibition. In all the above, many hippocampal neurons were severely damaged, however, CA3 pyramidal cells were mostly affected in normal aging and following hypobaric hypoxia, whereas CA1 cells were especially affected following corticosterone administration, global ischemia and ChE inhibition.Several mechanisms, which might be involved in the diverse courses of the lesions are being considered: cerebral oxygen and glucose, glutamate neurotoxicity and calcium involvement. It is anticipated that elucidation of the specific role of CA1 and CA3 hippocampal sub-fields in the various experimental models might help in understanding processes such as age-related neuronal degeneration and assist in their prevention.

Ocular injuries following sulfur mustard exposure: Pathological mechanism and potential therapy

Sulfur mustard (SM) is a potent vesicant, known for its ability to cause incapacitation and prolonged injuries to the eyes, skin and respiratory system. The toxic ocular events following sulfur mustard exposure are characterized by several stages: photophobia starting a few hours after exposure, an acute injury phase characterized by inflammation of the anterior segment and corneal erosions and a delayed phase appearing following a clinically silent period (years in human). The late injury appeared in part of the exposed eyes, expressed by epithelial defects and corneal neovascularization (NV), that lead to vision deficits and even blindness. During the last years we have characterized the temporal development of ocular lesions following SM vapor exposure in rabbits and have shown the existence of two sub-populations of corneas, those exhibiting delayed ocular lesions (clinically impaired) and those exhibiting only minor injuries if at all (clinically non-impaired). The aim of the present study was to investigate the pathological mechanism underlying the delayed injury by focusing on the unique characteristics of each sub-population and to test the efficacy of potential treatments. Clinically impaired corneas were characterized by chronic inflammation, increased matrix metalloproteinase (MMP) activity, poor innervation and limbal damage. Moreover, using impression cytology and histology, we identified the delayed lesions as typical for an ocular surface disorder under the category of limbal epithelial stem cell deficiency (LSCD). These results point to therapeutic directions, using anti-inflammatory drugs, MMPs inhibitors, neurotrophic factors and amniotic membrane transplantation. Topical anti-inflammatory drugs, either steroid (Dexamycin, DEX) or non-steroidal anti-infllammatory drug (NSAID, Voltaren Ophtha) were found to be beneficial in ameliorating the initial inflammatory response and in postponing the development of corneal NV, when given during the first week after exposure. When DEX was administered as a symptomatic treatment against NV, a significant regression in the angiogenic process was observed, however, the effect was temporal and blood vessels reappeared after therapy ceased. Chronic administration (8 weeks) of the MMP inhibitor Doxycycline was also effective in attenuation of the acute and delayed injury. Preliminary results, using amniotic membrane transplantation revealed some decrease of corneal edema with no effect on corneal NV. It is suggested that the chronic inflammation and prolonged impairment of corneal innervation are playing a role in the pathogenesis of the delayed LSCD following SM exposure by creating a pathological microenvironment to limbal epithelial stem cells, thus, leading to their slow death and to a second cascade of pathological events eventually resulting in severe long-term injuries. As of today, only topical anti-inflammatory drugs reached the criteria of an applicable efficient post-exposure ocular treatment for SM injuries. Further studies are required to investigate the effects of SM on epithelial stem cells and their involvement in the pathogenesis of the long-term injuries.

Characterization of acute and delayed ocular lesions induced by sulfur mustard in rabbits

Purpose. To establish an experimental model for sulfur mustard-induced acute and delayed ocular lesions in rabbits. Methods. Rabbit eyes were exposed to sulfur mustard (HD) vapor (370, 420 µg/l) for a period of two minutes. A three months follow-up study was carried out, based on the evaluation of clinical, biochemical and histological parameters. Results. HD exposure initiated typical clinical symptoms within 2–6 hrs, characterized by eye closure, eyelid swelling, conjunctival hyperemia, corneal erosions and inflammation. The clinical signs were significantly dose-dependent and reached a peak at 24–72 hrs post exposure. Biochemical evaluation of the aqueous humor exhibited an inflammatory reaction and oxidative stress at 4 hrs after exposure, subsiding at 28 hrs after exposure. Histological examination of corneas at 48 hrs revealed epithelial denudation and marked stromal edema, accompanied by cellular infiltration. Epithelial regeneration started after 72 hrs, and recovery was almost completed within 1Ð2 weeks, depending on the HD dose. A second phase of pathological processes started as early as two weeks post exposure and was characterized by corneal edema, opacity, recurrent erosions and neovascularization. The delayed injuries were found in 25 and 40% of the eyes respectively, and when appearing, were more severe than the initial ones. Conclusions. The development of HD-induced ocular lesions in rabbits is similar to the lesions described in human casualties. Quantitative analysis of the various clinical parameters emphasizes the contribution of each tissue to the overall toxic picture. Our experimental model is useful for studying the pathological mechanisms of HD-ocular lesions, and may serve for testing potential therapies.

Nimodipine Improves Spatial Working Memory and Elevates Hippocampal Acetylcholine in Young Rats

The calcium channel blocker nimodipine has been reported to improve cognitive performance in aged and brain-damaged animals. In the present study, the effects of nimodipine and placebo on spatial working memory and hippocampal acetylcholine were studied in young Fischer-344 rats. Nimodipine or placebo was administered via subcutaneously implanted, sustained-release pellets. Each active pellet contained 20 mg of nimodipine and released the drug over approximately 21 days. Two days after the drug or placebo pellets were implanted, training in the 8-arm radial maze started and continued for 12 days. Rats were required to learn a win-shift surgery. Nimodipine-treated animals learned the maze more rapidly than a placebo-treated group as indicated by the number of correct choices out of the first eight arms visited (p less than 0.001). Treated rats also made twice as many choices per unit time during the first week of training (p = 0.005). To assess hippocampal acetylcholine release, in vivo microdialysis was performed while animals were awake and unrestrained, 19-21 days after pellet implantation. A probe with a 3 mm semipermeable tip was placed in the hippocampus (CA1 and dentate gyrus), and individual microliters dialysate samples were collected at 2 microliters/min and immediately analyzed by high performance liquid chromatography with electrochemical detection. Significantly higher extracellular ACh levels were found in nimodipine-treated rats (71.4 +/- 3.6 nM; n = 4) compared to controls (52.5 +/- 2.5 nM; n = 5) (p = 0.003) and in another group of rats of the same age that received identical drug treatment.(ABSTRACT TRUNCATED AT 250 WORDS)

Long-Term Study of Brain Lesions Following Soman, in Comparison to DFP and Metrazol Poisoning

The long-term histopathological effects of acute lethal (95 μg kg-1) and sublethal (56 μg kg-1) doses of soman were studied in rats and were compared to lesions caused by equipotent doses of either another cholinesterase (ChE) inhibitor, DFP (1.8 mg kg-1), or a non-organophosphorus convulsant, metrazol (100 mg kg-1). Severe toxic signs were noted following one LD50 dose administration of all the compounds, yet only soman induced brain lesions. Moreover, even when administered at a sublethal dose (0.5 LD50), soman induced some histological changes without any clinical signs of intoxication. Soman-induced brain lesions were assessed quantitatively using a computerized image analyser. The analysis was carried out for up to 3 months following administration, and a dynamic pattern of pathology was shown. The cortical thickness and area of CA1 and CA3 cells declined significantly as early as 1 week post-exposure. No pathological findings were detected following DFP and metrazol administration. It is therefore suggested that brain lesions are not common for all ChE inhibitors and that convulsions per se are not the only factor leading to brain damage following the administration of soman. The degenerative process (found also with the sublethal dose of soman) might be due to a secondary effect, unrelated to somans clinical toxicity, but leading to long-term brain injuries.