N. V. Gulyaeva

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.

Single intracerebroventricular administration of amyloid-beta (25-35) peptide induces impairment in short-term rather than long-term memory in rats.

Ample experimental evidence indicates that intracerebral injection or infusion of amyloid-beta peptides (Abeta) to rodents induces learning and memory impairments as well as neurodegeneration in brain areas related to cognitive function. In the present study, we assessed the effects of a single intracerebroventricular (i.c.v.) injection of aggregated Abeta fragment (25-35) at a dose of 15nmol/rat on short-term and long-term memory in rats during the 6-month post-surgery period. The results demonstrate that Abeta(25-35)-induced memory impairments in spontaneous alternation behavior in a Y-maze at 17, 36, and 180 days after the surgery as well as in a social recognition task 110 days post-surgery. Abeta(25-35) also impaired spatial memory in an 8-arm radial maze, but did not influence performance of the step-down passive avoidance task. These results suggest that Abeta(25-35) preferably induces impairments of spatial and non-spatial short-term (working) memory rather than long-term memory in rats.

Caspase activity is essential for long-term potentiation

Slices from rat hippocampus were incubated with the caspase‐3 inhibitor N‐benzyloxycarbonyl‐Asp‐Glu‐Val‐Asp fluoromethylketone (Z‐DEVD‐FMK) or with the inactive peptide N‐benzyloxycarbonyl‐Phe‐Ala fluoromethylketone (Z‐Phe‐Ala‐FMK) for 30 min. The peptides changed neither input–output curves nor paired‐pulse effects at 70‐msec interpulse intervals, nor amplitudes of pop spikes in the CA1 region 1.0–6.9 hr after the incubation. Slices taken 1.0–1.4 hr after Z‐DEVD‐FMK or inactive peptide treatment demonstrated similar long‐term potentiation (LTP) curves; however, LTP was suppressed significantly (P < 0.001) 1.5–3.4 hr after Z‐DEVD‐FMK treatment when compared to the corresponding inactive peptide group. LTP magnitude correlated with time after Z‐DEVD‐FMK (r = −0.74; P < 0.02) but did not depend on time after the inactive peptide treatment. After 3.5 hr, LTP was blocked completely. Z‐DEVD‐FMK did not have a significant effect on presynaptic function. The results are the first evidence that inhibition of caspase‐3 significantly decreases or fully blocks LTP in the CA1 region and suggest that caspase‐3 is essential for LTP. Candidate caspase‐3 substrates that may be cleaved for LTP induction and maintenance are discussed.

Rodent Models of Depression: Neurotrophic and Neuroinflammatory Biomarkers

Rodent models are an indispensable tool for studying etiology and progress of depression. Since interrelated systems of neurotrophic factors and cytokines comprise major regulatory mechanisms controlling normal brain plasticity, impairments of these systems form the basis for development of cerebral pathologies, including mental diseases. The present review focuses on the numerous experimental rodent models of depression induced by different stress factors (exteroceptive and interoceptive) during early life (including prenatal period) or adulthood, giving emphasis to the data on the changes of neurotrophic factors and neuroinflammatory indices in the brain. These parameters are closely related to behavioral depression-like symptoms and impairments of neuronal plasticity and are both gender- and genotype-dependent. Stress-related changes in expression of neurotrophins and cytokines in rodent brain are region-specific. Some contradictory data reported by different groups may be a consequence of differences of stress paradigms or their realization in different laboratories. Like all experimental models, stress-induced depression-like conditions are experimental simplification of clinical depression states; however, they are suitable for understanding the involvement of neurotrophic factors and cytokines in the pathogenesis of the disease—a goal unachievable in the clinical reality. These major regulatory systems may be important targets for therapeutic measures as well as for development of drugs for treatment of depression states.

Effects of tumor necrosis factor-alpha central administration on hippocampal damage in rat induced by amyloid beta-peptide (25-35).

Male Wistar rats received unilateral intrahippocampal injections of 3 nmol (3.18 μg) aggregated Aβ(25–35), intracerebroventricular bilateral injections of 0.5 μg human recombinant TNFα or both (Aβ(25–35) + TNFα‐treated animals). Seven days after the surgery brain sections were stained with cresyl violet (Nissl), for fragmented DNA (TUNEL), glial fibrillar acidic protein (GFAP) and isolectin B4‐reactive microglia. In addition, caspase‐3 activity in brain regions was measured fluorometrically. The morphology of the hippocampus after the injection of Aβ(25–35) or both Aβ(25–35) and TNFα (but not TNFα alone) showed cell loss in the CA1 pyramidal cell layer. The extension of neuronal degeneration measured in the CA1 field was significantly larger in Aβ(25–35)‐treated groups compared to the contralateral hemisphere of both vehicle‐treated controls and animals injected with TNFα alone. TNFα augmented the Aβ(25–35)‐induced damage, significantly increasing the extension of degenerating area. Administration of Aβ(25–35) caused reactive gliosis in the ipsilateral hemisphere as demonstrated by upregulation of GFAP expression and the presence of hypertrophic astrocytes in the hippocampus. This effect was much more prominent in the hippocampi of rats treated with Aβ(25–35) + TNFα but absent after administration of TNFα alone. In both Aβ(25–35)‐treated groups, the damaged area of the hippocampal CA1 field and lateral band of dentate gyrus displayed many darkly stained round isolectin B4‐positive phagocyte‐like microglial cells. Sparse TUNEL‐positive nuclei were found in the hippocampi of rats treated with Aβ(25–35) alone or together with TNFα, but not in the control brain sections or in brain sections of TNFα‐injected animals. The activity of caspase‐3 increased significantly in the ipsilateral hippocampus after the injection of Aβ(25–35). Surprisingly, administration of TNFα into the cerebral ventricles prevented this Aβ(25–35)‐induced increase in hippocampal caspase‐3 activity. The results are discussed from the perspective of dual (neuroprotective and neurodestructive) roles of TNF in the brain.

Amyloid-β (25-35) increases activity of neuronal NO-synthase in rat brain

Nitric oxide (NO) is a free radical with multiple functions in the nervous system. NO plays an important role in the mechanisms of neurodegenerative diseases including Alzheimers disease. The main source of NO in the brain is an enzymatic activity of nitric oxide synthase (NOS). The aim of the present study was to analyze the expression and activity of both neuronal (nNOS) and inducible (iNOS) isoenzymes in the cerebral cortex and hippocampus of rats after intracerebroventricular administration of amyloid-beta (A beta) peptide fragment A beta(25-35). NADPHd histochemistry as well as immunohistochemistry were also used to investigate nNOS and iNOS expression in rat brain. The data presented here show that A beta(25-35) did not influence levels of nNOS or iNOS mRNA or protein expression in both structures studied. A beta(25-35) activated nNOS in the cerebral cortex and hippocampus without effect on iNOS activity. A beta(25-35) decreased the number of NADPHd-expressing neurons in the neocortex, but it did not significantly influence the number NADPHd-positive cells in the hippocampus. The peptide had no effect on the number of nNOS containing cells. We hypothesize that increased synthesis of NO induced by A beta(25-35) is related to qualitative alterations of nNOS molecule, but not to changes in NOS protein expression.

Studies of the effects of central administration of beta-amyloid peptide (25-35): pathomorphological changes in the Hippocampus and impairment of spatial memory.

The possible link between amnesia induced by central administration of β-amyloid (25–35) (Aβ(25–35)) and neurodegenerative changes in the hippocampus was studied. Male Wistar rats received single intracerebroventricular injections of Aβ(25–35) at a dose of 15 nmoles and one month later were trained in an eight-arm radial maze. Training was followed by histological assessment of the state of the hippocampus on brain sections stained with hematoxylin and eosin. Aβ(25–35) induced impairments in long-term (reference) and working memory on testing in the maze. There was a moderate reduction in the number of neurons in hippocampal field CA1; there was no change in the number of cells in field CA3. The numbers of errors made by the animals on testing in the maze were found to correlate negatively with the numbers of nerve cells in hippocampal field CA1. Thus, this is the first demonstration that impairments of learning and memory induced by single doses of Aβ(25–35) are specifically associated with neurodegenerative changes in hippocampal field CA1 in rats.

Administration of aggregated beta-amyloid peptide (25-35) induces changes in long-term potentiation in the hippocampus in vivo.

Intracerebroventricular administration of aggregated β-amyloid protein fragment (25–35) (7.5 nmol/ventricle) was followed one month later by significant changes in the dynamics of long-term potentiation in the hippocampus in vivo, expressed as powerful and stable increases in the amplitude of evoked potentials. This phenomenon may be associated with oxidative stress in the hippocampus, which has previously been demonstrated in this model, and, thus, with disturbances in ion homeostasis.

Pentylenetetrazole kindling induces neuronal cyclin B1 expression in rat hippocampus.

The purpose of the study was to explore the involvement of cell cycle events in the neuronal death induced by repeated seizures. Pentylenetetrazole (PTZ) kindling was used as a model of seizure-induced hippocampal neurodegeneration. Immunohistochemical approach was applied to detect cell cycle markers (cyclins and cycline-dependent kinases) in hippocampus. PTZ-kindling in rats induced moderate neuronal cell loss in hippocampal fields CA1, CA 3, CA 4, and dentate gyrus. The majority of damaged cells in hippocampi of PTZ-kindled rats were cycline B1 positive, while no expression of either other cell cycle markers or TUNEL-positive (apoptotic) nuclei could be revealed. Since cycline B1 expression has been described in hippocampal neurons of patients with temporal lobe epilepsy by [Z. Nagy, M.M. Esiri, Neuronal cyclin expression in the hippocampus in temporal lobe epilepsy, Exp. Neurol. 150 (1998) 240-247], it is suggested that PTZ-kindling may be a suitable model to study the mechanisms of seizure-induced neuronal death.

Maternal separation and early social deprivation in octodon degus: quantitative changes of nicotinamide adenine dinucleotide phosphate-diaphorase-reactive neurons in the prefrontal cortex and nucleus accumbens

The influence of postnatal socio-emotional deprivation on the development of nicotinamide adenine dinucleotide phosphate (NADPH)-diaphorase-reactive neurons in prefrontal cortical areas and in subdivisions of the nucleus accumbens was quantitatively investigated in the precocious rodent Octodon degus. Forty-five-days-old O. degus from two animal groups were compared: (i) degus which were repeatedly separated from their mothers during the first three postnatal weeks and after weaning reared in complete isolation; and (ii) degus which were reared under normal undisturbed social conditions. Socially-deprived animals displayed a significant decrease of NADPH-diaphorase-containing neurons in anterior cingulate cortex (85.5%), the same tendency was observed in the infralimbic, precentral medial and prelimbic prefrontal areas. Similarly, the core region of nucleus accumbens expressed reduced NADPH-diaphorase-reactive neuron numbers in deprived animals (70%), whereas the shell region remained unchanged. Since during normal postnatal development the number of NADPH-diaphorase-reactive neurons gradually decreases in all prefrontal cortical and accumbal regions, the observed deprivation-induced changes may reflect either an excessive reduction of NADPH-diaphorase-positive neurons or a down-regulation of the enzyme in neurons that normally express it. Since some NADPH-diaphorase-containing neurons in the prefrontal cortex have been shown to be GABAergic, it is tempting to speculate that a reduction of these inhibitory neurons in the anterior cingulate cortex may result in an enhanced excitatory output activity of disinhibited projection neurons in this cortical region, including those that project to the core region of the nucleus accumbens. Our results indicate a link between early adverse socio-emotional experience and the maturation of NADPH-reactive neurons and further studies are required to analyse the functional implication for this experience-induced brain pathology.