Yulia Moiseeva

Amyloid-β(25–35)-induced memory impairments correlate with cell loss in rat hippocampus

Amyloid beta-peptide (Abeta) plays an important role in the pathophysiology of Alzheimers disease. The relationship between amnesia induced by central administration of aggregated Abeta(25-35) and neurodegeneration in the hippocampus was investigated. One month after a single intracerebroventricular injection of Abeta(25-35) (15 nmol), male Wistar rats were tested in an eight-arm radial maze. A quantitative evaluation of cell number in hippocampal regions was carried out on H&E-stained brain sections of rats used in the behavioral study. Indices of free radical-mediated processes in the hippocampus were evaluated in additional groups of animals 1, 3, 5, and 30 days after surgery. Abeta(25-35) induced impairments of working and reference memory (RM) as well as neurodegeneration in the CA1 but not in the CA3 field of the hippocampus. A significant correlation between both reference and working memory (WM) impairments and the neuronal cell loss in the hippocampal CA1 region was demonstrated. A gradually developing oxidative stress was evident in the hippocampus of rats treated with Abeta(25-35) as indicated by the increase in 2-thiobarbituric acid (TBARS) reactive substances and superoxide generation. These data suggest the involvement of oxidative stress in Abeta(25-35)-induced neurodegeneration and a relation between memory impairment and neurodegeneration in the CA1 subfield of the hippocampus.

Role of nitric oxide in the ethylcholine aziridinium model of delayed apoptotic neurodegeneration in vivo and in vitro.

The involvement of nitric oxide in neurodegenerative processes still remains incompletely characterized. Although nitric oxide has been reported to be an important mediator in neuronal degeneration in different models of cell death involving NMDA-receptor activation, increasing evidence for protective mechanisms has been obtained. In this study the role of nitric oxide was investigated in a model of NMDA-independent, delayed apoptotic cell death, induced by the neurotoxin ethylcholine aziridinium ethylcholine aziridinium both in vivo and in vitro. For the in vivo evaluation rats received bilateral intracerebroventricular injections of ethylcholine aziridinium (2nmol/ventricle) or vehicle. In the hippocampus a transient decrease in nitric oxide synthase activity occurred, reaching its lowest levels three days after ethylcholine aziridinium treatment (51.7+/-9.8% of controls). The decrease coincided with the maximal reduction in choline acetyltransferase activity as marker for the extent of cholinergic lesion. The effect of pharmacological inhibition of nitric oxide synthase was tested by application of various nitric oxide synthase inhibitors with different selectivity for the nitric oxide synthase-isoforms. Unspecific nitric oxide synthase inhibition resulted in a significant potentiation of the loss of choline acetyltransferase activity in the hippocampus measured seven days after ethylcholine aziridinium application, whereas the specific inhibition of neuronal or inducible nitric oxide synthase was ineffective. These pharmacological data are suggestive for a neuroprotective role of nitric oxide generated by endothelial nitric oxide synthase. In vitro experiments were performed using serum-free primary neuronal cell cultures from hippocampus, cortex and septum of E15-17 Wistar rat embryos. Ethylcholine aziridinium-application in a range of 5-80microM resulted in delayed apoptotic neurodegeneration with a maximum after three days as confirmed by morphological criteria, life-death assays and DNA laddering. Nitric oxide synthase activity in harvested cells decreased in a dose- and time-dependent manner. Nitric oxide production as determined by measurement of the accumulated metabolite nitrite in the medium was equally low in controls and in ethylcholine aziridinium treated cells (range 0.77-1.86microM nitrite). An expression of inducible nitric oxide synthase messenger RNA could not be detected by semiquantitative RT-PCR 13h after ethylcholine aziridinium application. The present data indicate that in a model of delayed apoptotic neurodegeneration as induced by ethylcholine aziridinium neuronal cell death in vitro and in vivo is independent of the cytotoxic potential of nitric oxide. This is confirmed by a decrease in nitric oxide synthase activity, absence of nitric oxide production and absence of inducible nitric oxide synthase expression. In contrast, evidence for a neuroprotective role of nitric oxide was obtained in vivo as indicated by the exaggeration of the cholinergic lesion after unspecific nitric oxide synthase inhibition by N-nitro-L-arginine methylester.

Tongue protrusion: a simple test for neurological recovery in rats following focal cerebral ischemia

A simple tongue protrusion (TP) test is described for rats following focal ischemia induced by middle cerebral artery occlusion (MCAO). MCAO resulted in a dramatic decrease in TP that correlated with a concomitant decline in neurological performance in standard 5- and 20-point tests and deficits in performance in the Morris water maze and the accelerating rotarod. TP values also correlated with infarct size at 7 and 24 days following MCAO. This simple and inexpensive test, that monitors the ability of rats to lick food out of a glass tube, is easily administered, can be administered frequently without changing baseline performance, is not susceptible to behavioral compensation and should not interfere with other tests used concurrently to evaluate neurological deficit. The TP test may, therefore, serve as a useful addition to the battery of tests commonly used to assess neurological damage in rats, particularly in models of stroke.

Region-specific changes in activities of cell death-related proteases and nitric oxide metabolism in rat brain in a chronic unpredictable stress model.

Effects of a chronic combined unpredictable stress on activities of two cell death-related proteases, calpain and cathepsin B, were studied along with indices of nitrergic system in rat brain structures. Male Wistar rats were subjected to a 2-week-long combined stress (combination of unpaired flash light and moderate footshock associated with a white noise session). Stress resulted in a significant loss in the body and thymus weight and increased defecation in the open field test, though neither motor and exploratory activity, nor plasma corticosterone differed from the respective control levels. Decreased calpain activity and increased cathepsin B activity were demonstrated in the hippocampus of stressed rats (previously we have shown that caspase-3 activity was significantly suppressed in the brain of rats subjected to same type of stress). A significant reduction in the number of NOS-containing neurons was accompanied by a chronic stressinduced decline in NOS activity in the neocortex. Similar changes were observed in the hippocampus. However, levels of NO metabolites were elevated in both structures. Thus, stress-induced structural modifications in the brain may be mediated by disturbances in the nitrergic system and increased lysosomal proteolysis.

Nitric oxide synthase activity in Fasciola hepatica: a radiometric study.

The activity of neuronal nitric oxide synthase (nNOS) in homogenates of adult Fasciola hepatica was measured by the direct radiometric assay of the production of L-[3H]citrulline. This is the first radiometric study of the activity of nNOS in a fluke. The effect of arginase was tested. In the presence of L-valine, which is an inhibitor of arginase, the formation of L-[3H]citrulline decreased from 12% to 38%, depending on the time of incubation. This means that the arginase activity in the worm is high, and has to be taken into consideration when measuring the activity of nNOS. When co-factors, such as H4B, and NADPH, were omitted the formation of L-[3H]citrulline decreased significantly (29%). The effects of several nNOS inhibitors were tested. N(omega)-nitro-L-arginine (L-NAME), aminoguanidine and S-methyl-L-thiocitrulline added at a concentration of 1 mM inhibited the L-[3H]citrulline formation by 28%, 15% and 14%, respectively. Chelation of Ca2+ with 1 mM EGTA resulted in a 40% decrease in the formation of L-[3H]citrulline. These results indicate the presence of nNOS activity in homogenates of F. hepatica.

Interval Hypoxic Training Prevents Oxidative Stress in Striatum and Locomotor Disturbances in a Rat Model of Parkinsonism

In Parkinson’s disease, a progressive degeneration of nigro-striatal dopaminergic neurons results in akinesia, muscular rigidity and tremor. MPTP (l-methyl-4-phenyl-1,2,3,6-tetrahydropyridine) is a neurotoxin that is responsible for the Parkinson-like symptoms seen in humans using illicit synthetic opiate analogs of meperidine containing MPTP as a contaminant (Langston et al., 1983). Rodents are highly susceptible to the neurotoxic effects of MPTP and may be useful animal models for Parkinson’s disease (Hallmann et al., 1985; Takada et al., 1987). Administration of MPTP to rodents results in a significant loss of nervous cells in the substantia nigra (Takada et al., 1987) and a marked reduction in neostriatal content of dopamine and its metabolites (Heikkila et al., 1984; Fuller and Hemrick-Luecke, 1984).

Intracerebroventricular Administration of Beta-Amyloid Peptide (25-35) Induces Oxidative Stress and Neurodegeneration in Rat Brain

The cytotoxic action of beta-amyloid has been considered to be the primary determinant of the neurodegeneration observed in Alzheimer’s disease. Many aspects of the molecular mechanisms associated with neurotoxic activity of beta-amyloid are being investigated using the synthetic peptide—the active fragment of beta amyloid protein containing residues from 25 to 35 of the parent compound [beta (25–35)].

Specific Activity Features in the Forced Swim Test: Brain Neurotrophins and Development of Stress-induced Depressive-like Behavior in Rats

Selective vulnerability or resilience to mood disorders is related to individual differences or personality. In the present study forced swim test (FST) was used as a tool for division of male rats according to their immobility behavior. The animals were subjected to a chronic unpredictable mild stress (CUS). Depressive-like behavior and modifications in brain neurotrophin system of were examined after CUS exposure. The low immobile (LI) and high immobile (HI) rats demonstrated elusive differences in expression of BDNF ExVI mRNA and TrkA mRNA which was higher in the hippocampus and frontal cortex, respectively, of HI rats as compared to LI animals. Exposure to CUS resulted in development of depressive-like phenotype and increased anxiety in both subgroups; however, immobility in FST specifically decreased in the initially HI animals. In hippocampus of stressed LI rats, the contents of total BDNF mRNA decreased. In hippocampus of stressed HI rats, the content of TrkA mRNA increased whereas in frontal cortex, the content of BDNF exon I mRNA decreased in both LI and HI rats. The levels of BDNF ExIX and ExI as well TrkB mRNAs were higher in the hippocampus of HI rats as compared to LI rats. In general, the response of hippocampus to CUS was much more expressed as compared to frontal cortex. Thus, initially different stress coping strategies of rats in the FST (HI, LI) were associated with the development of similar behavioral phenotypes after chronic unpredictable stress; however, these phenotypes were associated with different alterations in neurotrophin systems of the brain.

P2-001: Pathomorphological correlates of behavioral abnormalities in amyloid injection rodent model of Alzheimer’s-like pathology

upon symptoms in the Mood cluster at both weeks 12 (P .034) and 24 (P .033), with 65.5% of patients in the memantine group showing a positive response at week 24. Memantine also had a significant effect over placebo (OC) upon symptoms of Psychosis at both weeks 12 (P .006) and 24 (P .001), with 80.7% of patients in the memantine group showing a positive response in this domain at week 24. The response difference (OC) between memantine and placebo patients at week 24 was 12.2% and 18.9% for Mood and Psychosis clusters, respectively. LOCF analysis yielded comparable results. Effects of memantine on Frontal symptoms were not significant, while the effects on Other symptoms were significant at week 24 using LOCF analysis (P .037), but not OC analysis (P .058). Conclusions: Taken together, these results suggest that memantine provides specific behavioral benefits for mood and psychosis-related symptoms associated with AD.