D. E. Korzhevskii

Structural Organization of Striatal Microgliocytes after Transient Focal Ischemia

Microgliocytes are known not to be structurally stable cellular elements; altering the lengths of their processes, they control the synaptic organization of the brain. The aim of the present work was to study the structural organization of the microglia in the striatum and periventricular area of the telencephalon of the rat after transient focal ischemia. Ischemic lesions without cerebral infarction were found to lead to activation of the microglia, promoting reorganization of the striatal neuropil. Activation of microgliocytes was associated with changes in their shape, from the characteristic dendritic shape to the oval and round shapes typical of the ameboid microglia. Microgliocytes were able to proliferate after ischemia.

Иммуногистохимическая характеристика нейронов черного вещества головного мозга человека

AIMnTo determine the cytochemical characteristics of unchanged neurons of the human substantia nigra using a wide range of immunocytochemical markers some of which (glutamate decarboxylase-65, PGP 9.5, non-phosphorylated neurofilament proteins, alpa-tubulin) have never been used for study of human dopaminergic neurons.nnnMATERIAL AND METHODSnFragments of human midbrain (17 men and women, aged from 28 to 78 years) from the archives of the Department of General and Specific Morphology of the Institute of Experimental Medicine were used. The study was performed using classical neurohistological techniques and immunocytochemistry using antibodies to 15 different proteins.nnnRESULTSnMost neurons in substantia nigra exhibited a reduced expression of common neuronal markers such as neuronal nuclear protein NeuN, PGP 9.5 protein, and neuron-specific enolase. GABAergic (GAD65-immunopositive) neurons were not found in the substantia nigra. Single cholinergic neurons without neuromelanin were identified in the dorsal part of the substantia nigra. Calcium-binding proteins calbindin and calretinin were not found in the majority of nigral cells although calbindin was rarely seen in some neurons of the dorsal part and calretinin in the ventral one. Nitric oxide synthase (NOS) was present in the substantia nigra both in neuropil and neuronal bodies.nnnCONCLUSIONnThe results suggest the unique cytochemical properties of the nigral neurons, which may be related to their increased susceptibility to lesion and degeneration.

Characterization of amyloid deposits found in internal organs of mdx mice

The mdx mouse strain is the most widely used experimental model of Duchenne muscular dystrophy (DMD). Although it was previously shown that muscle biopsy specimens obtained from patients with different types of muscular dystrophy contain amyloid, no available publications have analyzed the presence of amyloid aggregates in tissues of DMD patients or mdx mice. The objective of the present work was to verify whether the internal organs of mdx mice might accumulate amyloid. The study was performed in the myocardium, kidney, and liver specimens obtained from male and female mdx mice aged from 2 to 18 months. Using histochemical staining with Congo red, amyloid aggregates were detected in mouse organs studied, and their morphology and location were analyzed. Mass spectrometry data suggest that the most probable components of amyloid aggregates found in mdx mice are vitronectin and apolipoprotein A-II.

A Method of Enhanced Immunofluorescence by Amplification of the Immunoperoxidase Reaction

The aim of the present work was to develop a simple and effective means of converting the immunoperoxidase reaction into an immunofluorescence reaction. Use of fluorochrome-conjugated antibodies to horseradish peroxidase significantly increased the intensity of the immunofluorescence reaction as compared with the indirect Coons method, which was confirmed by quantitative evaluation of fluorescence intensity. The conversion method developed here does not require the protocol to be expanded by a large number of steps or any reagent substitutions, which facilitates the transition from the immunoperoxidase reaction to the immunofluorescent reaction.

Glutamine Synthetase in Rat Brain Cells

Objective. To use an immunohistochemical method to study glutamine synthetase (GS)-synthesizing brain cells. Materials and methods. Enzyme was detected on frontal rat brain sections (n = 10) using mouse monoclonal antibodies. Specimens were analyzed by light and confocal laser microscopy. Results. GS was found to be expressed in all areas of the brain, mainly by two types of cells, with different structures and topographies. The main type of cell with immunopositive reactions to glial fibrillary acidic protein were identical to astrocytes. The other structural and locational type of cell differed from typical astrocytes. Conclusions. The data obtained here provide evidence that GS is not a selective marker for any particular population of rat brain cells.

Mitochondrial HSP60 Protein in Rat Medulla Oblongata Cells

HSP60 is a member of the heat shock protein (HSP) group and is located predominantly in cell mitochondria. Dysfunction and insufficient synthesis of HSP60 in the nervous system leads to neurodegeneration. The aim of the present work was to conduct immunohistochemical determination of the location of HSP60 protein in neurons and gliocytes in the medulla oblongata in rats. Studies were performed on mature male Wistar rats weighing 200–250 g (n = 7). The results provided evidence that HSP60 has a discrete localization in cytoplasmic structures in neurons and gliocytes. In large neurons, local accumulation of HSP60 allows fine structures to be detected, probably mitochondria. An important observation is that of the absence of any cytosolic reaction in medulla oblongata neurons and gliocytes. This provides grounds for using the reaction for HSP60 in complex studies of apoptosis in nerve cells, as this is associated with an accumulation of HSP60 and displacement of immunopositive product to the plasmalemma.

Fluorene Derivative Disodium Salt as a New Fluorescent Dye for Identification of Amyloid Deposits in Myocardium of mdx Mice

The aim of this study was to develop a new method for the detection of amyloid deposits in laboratory animals using an analogue of Congo red synthesized on the basis of diaminofluorene. The analogue is disodium salt of 2,7-(1-amino-4-sulfo-2-naphthylazo)fluorene (DSNAF). Myocardial samples from mdx mice of both sexes aged 1–1.5 years (n = 8) were used as the material for this study. The main result of this study was the development of an optimal protocol for amyloid staining with DSNAF. It has been shown that the sensitivity and specificity of amyloid detection by this method is comparable with Congo-red staining. The clear advantages of using DSNAF are stability of staining, high fluorescence intensity of amyloid deposits, and total lack of background fluorescence, which greatly simplifies the procedure of quantitative evaluation of obtained results. The method of amyloid staining with DSNAF is characterized by simplicity and good reproducibility. Further research will show the possibility to apply this method for diagnosis of amyloidosis in the practice of clinical research.

Nucleolin and Nucleoli in Ependymocytes and Tanycytes of the Third Ventricle of the Rat Brain

The aim of the present study was to compare structure of the nucleoli of ependymocytes, tanycytes, and secretory cells of the subcommissural organ using immunohistochemical staining for nucleolin and confocal laser microscopy. The study was performed in samples from the diencephalon of adult male Wistar rats (n = 6). The samples were fixed in zinc–ethanol–formaldehyde, a fixative providing a high level of preservation of antigen determinants. In the present study, we estimated diameters of nucleoli and their number in various types of cells lining the third ventricle. We compared for the first time the nucleoli of different subpopulations of tanycytes and report data on the distribution of nucleolin protein in the cells lining the ventricles. The content and location of nucleolin reflect the functional state of the cell. Our data will promote understanding of the interrelationships between the indices of the nucleolar apparatus and the functional state of the cell under various conditions, including stress, neoplastic transformation, and other pathological conditions.

Cell Contact Protein β-Catenin in Ependymal and Epithelial Cells in the Choroid Plexus of the Lateral Ventricles of the Brain

The aim of the present work was to study the distribution of cell contact protein β-catenin in the choroid plexus and ependyma of the lateral ventricles of the brain. Experiments were performed using frontal sections of brains from Wistar rats (n = 10) using polyclonal antibodies to β-catenin. Preparations were analyzed under the microscope in transmitted light and by confocal laser microscopy. The distribution of β-catenin in different projections was studied by three-dimensional reconstruction. The results identified differences in the distributions of this protein in the ependyma and choroid plexus. In contrast to the ependyma, β-catenin is distributed in choroid plexus cells in the same way as in monolayer epithelial tissues (at the basal and lateral margins of the cells). This may be evidence that the ependyma and epithelium of the choroid plexus are of different tissue types, despite their common origin.

Intercellular Contact Protein β-Catenin in Neurons of the Habenula of the Rat Brain

Studies were performed on frontal brain sections (n = 5) from Wistar rats using immunocytochemical methods. This is the first report on the detection of the protein β-catenin in the nuclei of mature neurons in the medial part of the habenula. As the intranuclear localization of this protein points to its involvement in the canonical Wnt signal pathway, which is expressed during embryogenesis, it can be suggested that habenular neurons retain properties not typical of mature neurons.