[Fusion of frog cerebellar granule cells induced by toxic effects of glutamate and NO-generating compound].

The ultrastructural changes of cerebellar granule cells were studied under stroke modeling conditions, after the toxic effects of glutamate (Glu) and NO-generating compound. Glu toxic doses were shown to induce two types of nuclear chromatin changes. In some cases, the appearance of practically completely decondensed nuclear chromatin was detected, while in the others the nuclei contained partially decondensed chromatin. Pathological fusion of granule cells was observed in both cases. The toxic effect of NO-generating compound on granule cells also caused the appearance of the cells with both completely and partially decondensed (flocculent) chromatin. Granule cells with different chromatin type were able to fuse with each other. Thus, Glu and NO, causing changes in nuclear chromatin, activate the processes of cell clustering with the following cytoplasmic fusion and the formation of multinuclear conglomerates. The possible physiological role of granule cells fusion induced by high concentrations of Glu and NO-generating compound is discussed. This process is considered as a realization of the compensatory-adaptive reactions under extreme conditions observed in the stroke and oxidative stress.

Role of glycogen in processes of cerebellar glial cells under conditions of its damage with sodium nitrite.

Ultrastructure of processes of glial cell, astrocytes of the molecular layer of cerebellar cortex in Rana temporaria frog, under conditions of damage to the cerebellum caused by NO-generating compound sodium nitrite was studied under an electron microscope. It was found that astrocytes have at least two types of processes: the first (fibrillar) primarily contained numerous fibrils and few glycogen granules and the second (granular) primarily containing glycogen granules. In the presence of NO-generating compound in toxic doses, fibrillar processes are damaged or completely degrade more rapidly than granular ones. The processes containing glycogen can protect both damaged synapses and individual synaptic buttons by forming a compact structure, wrapping, around them. We analyzed the possible role of glycogen of cerebellar glial cell processes in neuroglial interactions in the presence of sodium nitrite.

Neuron-glial contacts formed in the cerebellum during electrical stimulation in the presence of an NO-generating compound.

The molecular layer of the cerebellum of the frog Rana temporaria was studied by light and electron microscopy after electrical stimulation in the presence of an NO-generating compound. In these conditions, there was severe swelling of granule cell axon terminals (boutons) and astrocyte processes (AP), with loss of cytoplasmic elements. However, along with damaged structures, there were also undamaged structures: boutons with synaptic vesicles and AP with glycogen granules. It is suggested that these persisting viable AP may form 1) glial "wrappings" around damaged synapses or boutons and 2) neuron-glial contacts, which form when synaptic vesicles cross damaged bouton membranes to AP containing glycogen granules. It is also suggested that the presence of glycogen in AP in conditions of oxygen and glucose deficiency may provide a source of high-energy substrates such as glucose and ATP, thus providing conditions for neuron survival in pathological states (ischemia/hypoxia).

Effect of NO-generating compound NaNO2 on ultrastructure of synaptic vesicles of glutamatergic synapses.

Ultrastructure of synaptic vesicles in axon terminals of granule cells from isolated cerebellum of Rana temporaria frogs under the influence of NO-generating compound NaNO2 in various concentrations and electrical stimulation was evaluated by the method of electron microscopy. NO-generating compound in low concentration induced translocation of synaptic vesicles and formation of small clusters. The size and structure of synaptic vesicles remained unchanged under these conditions. Increasing the concentration of NaNO2 led to swelling of synaptic vesicles, formation of arranged heaps from individual vesicles or fusion of their content. Electrical stimulation of the cerebellum in the presence of NaNO2 increased damage to synaptic vesicles. These experimental data model some stages observed in stroke. The formation of clusters from synaptic vesicles is a compensatory and adaptive response maintaining the structure of synaptic vesicles and protecting neurons from high concentrations of glutamate. Glutamate produces a toxic effect on nerve cells and glial cells of the cerebellum under pathological conditions, which is accompanied by impairment of signal transduction from presynaptic to postsynaptic neurons.

[Neuron-glial junction formation in cerebellum after electrical stimulation in presence of no-generating substance].

Molecular layer of frog (Rana temporaria) cerebellum was studied using light and electron microscope after electrical stimulation of parallel fibers in presence of NO-generating compound. Under these conditions, significant swelling of axonal terminals (boutons) of granular cells and astrocyte processes (AP) with a loss of cytoplasmic elements. However, along with the damaged structures, intact boutons were found with synaptic vesicles and APs containing glycogen granules. It is suggested that the remaining viable APs are capable of forming 1) protective glial "wrappings" around damaged synapses or boutons, and 2) neuron-glial junctions, that are formed due to transmission of synaptic vesicles through the damaged membrane of bouton into AP containing glycogen granules. It is also proposed that the presence of glycogen in APs under conditions of oxygen and glucose deficit may serve as the source of such energy-containing substrates, such as glucose and ATP, and thus may provide for neuronal survival in pathological states (ischemia/hypoxia).

[Protective role of autotypic contacts under cerebellar neural net injury by toxic doses of NO-generative compounds].

In the present work, cerebellar neural net injury was induced by toxic doses of NO-generative compound (NaNO2). A protective role of glial cells was revealed in such conditions. The present results were compared with those of the previous work concerning the action of high concentration glutamate on the frog cerebellum (Samosudova et al., 1996). In both cases we observed the appearance of spiral-like structures--"wrappers)"--involving several rows of transformed glial processes with smaller width and bridges connecting the inner sides of row (autotypic contact). A statistic analysis was made according to both previous and present data. We calculated the number and width of rows, and intervals between bridges depending on experimental conditions. As the injury increased (stimulation in the NO-presence), the row number in "wrappers" also increased, while the row width and intervals between bridges decreased. The presence of autotypic contacts in glial "wrappers" enables us to suppose the involvement of adhesive proteins--cadherins in its formation. The obtained data suggested that the formation of spiral structures--"wrappers" may be regarded as a compensative-adaptive reaction on the injury of cerebellar neural net glutamate and NO-generative compounds.

[Nitric oxide as a contrast modulator of basic elements of the cytoskeleton]. / Oksid azota kak moduliator kontrastnosti osnovnykh élementov tsitoskeleta.

Early it was shown that nitric oxide induced in the cerebellum neuronal net both degenerative and compensatory-adaptive changes: 1) bouton encapsulation of spines, and 2) spiral wraps formed by glial cell processes around synapses and boutons. All these morphological changes were produced with cytoskeleton involvement. In the present work we have found that a NO-generative compound enhanced the contrast of cytoskeleton elements which depended on the concentration of this compound. The best contrast was observed at 1 mM concentration. The reason of the contrast enhance may be due presumably to protein transition from a soluble to a membrane-bound state. Using the contrast enhance effect we carried out a comparative analysis of cytoskeleton elements (CE) composition. Results of the analysis showed the specificity of CE in different cell structures: bouton, spine, glial cell. The obtained data support our proposal about the leading role of cytoskeleton in compensatory-adaptive morphological changes in extremal conditions.

[Increase in the efficiency of parallel fiber synapses on Purkinje cells of the frog after simultaneous activation of climbing and parallel fibers]. / Uvelichenie éffektivnosti sinapsov parallel'nykh volokon na kletkakh Purkin'e liagushki posle odnovremennoi aktivatsii liannykh i parallel'nykh volokon.

Ability of the Purkinje cells to long-term plasticity changes after joint stimulation of parallel and climbing fibres has been studied in the isolated cerebellum-medulla oblongata preparation of frog. 18 neurons have been chosen on the ground of the presence of clearly identifiable monosynaptic response to parallel fibre stimulation and stability of the background activity within two hours. These neurons belonged to three distinct categories: those having the clear climbing fibre response, those having the climbing fibre-like response and those without the climbing fibre response. The neurons of the first group showed a significant increase of the ability to respond to parallel fibre stimulation after joint stimulation of both types of fibres. The stimulating current strength which was necessary for the Purkinje cell firing index 1/2 was reduced after conditioning procedures in this neuron group to 0.7 of its original value. There were no substantial changes in parallel fibre stimulation efficiency in two other neuron groups.