Satellite cells of sensory neurons after various types of sciatic nerve trauma in the rat.

Sciatic nerve crushing, transection, and ligation models were used in rats to study the reactions of and changes in the numbers of satellite cells (SC) in spinal dorsal root ganglia in the lumbar segment. Nerve transection was followed by the appearance of neurons surrounded by two layers of SC. The thickness of SC processes and the areas of contacts with neurons increased as a result of invaginations into neuron perikarya. After nerve ligation, SC and their processes were located around parts of large and intermediate neurons in several tightly appressed layers; the area of contact between SC and neuron perikarya showed increased development of invaginations such that lamellar structures appeared in the SC cytoplasm, along with contacts with SC processes surrounding neighboring neurons. The greatest increases in SC numbers were seen after ligation of the nerve. Transection was followed by increases in the numbers of small and intermediate neurons surrounded by vimentin-positive SC. The number of large neurons surrounded by these cells decreased. At all time points following ligation of the nerve, all neurons in the study ganglia were surrounded by vimentin-positive SC. Post-traumatic changes in structure and numbers differed in SC associated with sensory neurons of individual size populations and depended on the type of trauma applied to efferent conductors.

Morphofunctional analysis of experimental model of esophageal achalasia in rats.

We carried out a detailed analysis of rat model of esophageal achalasia previously developed by us. Manifest morphological and functional disorders were observed in experimental achalasia: hyperplasia of the squamous epithelium, reduced number of nerve fibers, excessive growth of fibrous connective tissue in the esophageal wall, high contractile activity of the lower esophageal sphincter, and reduced motility of the longitudinal muscle layer. Changes in rat esophagus observed in experimental achalasia largely correlate with those in esophageal achalasia in humans. Hence, our experimental model can be used for the development of new methods of disease treatment.

[Satellite cells of the sensory neurons after different types of sciatic nerve trauma in rat].

The reaction of satellite cells (SC) and the changes in their numbers were studied in rat lumbar dorsal root ganglia using the models of sciatic nerve crush, transection and ligation. After the nerve transection, the neurons surrounded by two layers of SC were found. This was accompanied by the increased SC branch thickness and contact area due to invaginations into neuronal perikarya. After the nerve ligation, both SC and their branches were found to form several closely adjacent layers around the part of the large and medium neurons, the area of SC contact with the perikarya of neurons of these populations was increased due to more developed invaginations, there appeared the multilamellar structures in SC cytoplasm and the contacts with the branches of SC, which surrounded the neighboring neurons. The most pronounced increase in SC numbers was demonstrated after the nerve ligation. After the nerve transection, the numbers of small and medium neurons, surrounded by vimentin-positive SC, was increased. At the same time, the number of large neurons surrounded by these cells, was decreased. At all time intervals after the nerve ligation, all the neurons in the ganglia studied were surrounded by vimentin-positive SC. Post-traumatic changes in structure and numbers were different in SC, associated with specific populations of sensory neurons and depended on type of afferent conductor injury.

Posttraumatic survival of sensory neurons during allotransplantation of rat embryonic tissues into the nerve.

Allotransplantation of the liver, hindlimb, and bone marrow tissues from 14-day-old embryos into the sciatic nerve had a modulatory effect on survival of various populations of axotomized neurons in L5 spinal ganglion in an adult rat.

[Effect of xymedone on high-voltage-activated Ca2+ currents in pyramidal neurons of the entorhinal cortex].

The effect of xymedon on Ca2+ currents in entorhinal cortex LIII pyramidal neurons was studied using brain slices from 10-17-day old rats, which were analyzed by means of the infrared video assisted whole cell patch clamp recording. The sample slices were superfused with artificial cerebrospinal fluid containing tetrodotoxin, 4-aminopyridine, and tetraethylammonium for the blocking of Na+ and K+ channels, respectively. Xymedone was added to artificial cerebrospinal fluid and to all extracellular solutions. The slices were exposed to different concentrations of xymedone for 3 hours followed by patch-clamp recordings. Control recordings were run with the vehicle. Xymedone in a concentration of 0.01 mM decreased the maximum voltage-dependent Ca2+ current amplitude by 39.8 %, while 1 mM of xymedone inhibited the Ca2+ currents almost completely. The obtained data showed for the first time that xymedone exhibits a calcium channel blocker activity in neurons. Possible neuroprotective mechanisms of xymedone are discussed.

Survival and phenotypic characteristics of axotomized neurons in spinal ganglia.

Trauma to the peripheral processes of sensory neurons of different subpopulations was followed by indirect immunohistochemical analysis of the expression of Bcl-X(L) and Bax, which are, respectively, antiapoptotic and proapoptotic proteins of the Bcl-2 family, and also of the cytokine interleukin-1beta, with the aim of identifying the roles of these substances in controlling apoptosis. The survival abilities of these neurons after central and peripheral axotomy were compared by studying the expression of the high molecular weight component of the neurofilament triplet NF200 and isolectin B4 (IB4). By day 30 after central axotomy, there were no changes in the total numbers of neurons in ganglia L(IV)-L(V) in rats, though there were significant reductions in the numbers of NF200+ neurons. In spinal ganglion L(V) of mice, the proapoptotic protein was detected in the nuclei of 46% of small neurons, which account for 20% of all neurons in the ganglion. By day 30 after nerve compression, Bax was expressed in the nuclei of 30% of neurons and the cytoplasm of 20% of neurons. In intact animals, the antiapoptotic protein Bcl-X(L) was seen in the cytoplasm of 30% of small neurons, as well as in satellite cells surrounding large and intermediate neurons. By day 30 after nerve trauma, Bcl-X(L) was not expressed in spinal ganglion L(V). Interleukin-1beta was present in the cytoplasm of 17% of neurons belonging to the subpopulations of large and intermediate neurons. By day 30 after nerve compression, interleukin-1 beta+ neurons were not identified.

Post-traumatic survival of sensory neurons of different subpopulations.

Immunohistochemical studies were performed to address the expression of the high-molecular-weight component of the neurofilament triplet NF200 (a marker of neurons forming A fibers) and the binding of isolectin B4 (IB4) by neurons of the L4-5 spinal ganglia after ligation or section of the sciatic nerve in rats. A total of 15% of neurons in the ganglia of intact rats expressed NF200. By 90 days after nerve ligation, the proportion of NF200+ neurons decreased two-fold; administration to these rats of the nerve regeneration stimulator xymedone increased the number of NF200+ neurons by 50.7% compared with controls (ligation, no treatment). In intact rats, 23.6% of neurons bound IB4. The proportion decreased by 2.6% 30 days after nerve ligation and to undetectable levels by 90 days; xymedone increased the proportion of surviving IB(4)+ neurons more than eight-fold. IB(4)+ neurons were more likely to enter post-traumatic apoptosis. Ligation of the nerve was followed by survival of fewer NF200+ and IB(4)+ neurons than section of the nerve, which suggests that axon lengthening is a factor maintaining neuron survival. The pyrimidine derivative xymedone increased the survival of neurons of both subpopulations, especially IB(4)+ neurons.

[Survival and phenotypic characterisctic of axotomized dorsal root ganglion neurons]. / Vyzhivanie i fenotipicheskaia kharakteristika aksotomirovannykh neironov spinal'nykh gangliev.

Using indirect inmunohistochemical method, the expression of Bcl-XL and Bax, anti- and proapoptotic proteins of Bcl-2 family, as well as of cytokine IL-1beta were studied to demonstrate the role of these substances in apoptosis regulation of sensory neurons of different subpopulations after the severance of their peripheral processes. For comparison of the capacity of these neurons to survive after central and peripheral axotomy, the expression of high-molecular component of neurofilament triplet NF200 and isolectin B4 (IB4) binding were studied. At day 30 after central axotomy, the total number of neurons in LIV-LV ganglia of rat was not changed, but the number of NF200+ neurons was decreased. In mouse Lv dorsal root, proapoptotic BaX protein was demonstrated in the nuclei of 46% of small neurons that accounts for 20% of the total neuronal number in this ganglion. By day 30 after the nerve crush separate Bax expression was found in the nuclei of 30% and in the cytoplasm of 20% of neurons. In intact animals, dorsal root ganglion antiapoptotic Bcl-XL protein was expressed in the cytoplasm of 30% of small neurons, as well as in satellite cells surrounding large and intermediate neurons. At day 30 after the nerve crush Bcl-XL expression in LV ganglion was not detected. IL-1beta was present in the cytoplasm of 17% of neurons belonging predominantly to the subpopulations of large and intermediate neurons. By day 30 after the nerve crush IL-1(+-neurons were not found.

[Role of transthyretin in posttraumatic regeneration of the sciatic nerve in mouse]. / Rol' transtiretina v posttravmaticheskoi regeneratsii sedalishchnogo nerva myshi.

Although thyroid hormones are known to have a significant influence on the development of nervous system, the absence of changes in the brain of mice deficient in transthyretin--a protein providing thyroid hormone transport across the blood-brain and blood-nerve barrier--remains unexplained. Therefore, the aim of this study was to determine the effect of transthyretin on the formation of myelinated and unmyelinated nerve fibers in sciatic nerve of mice. The myelinated and unmyelinated nerve fibers were counted in sciatic nerve of 3-months-old normal and transthyretin-knockout (transthyretin(-/-)) mice 15 and 30 days after nerve crushing. No differences were detected in the number of myelinated and unmyelinated nerve fibers in intact control (wild-type) animals group vs. transthyretin(-/-) mice. By days 15 and 30 after nerve crushing the number of myelinated nerve fibers was diminished by 54.7 and 71.8%, respectively, in transthyretin(-/-) mice, as compared to that in control animals. The number of unmyelinated nerve fibers at day 15 after the injury was not different in transthyretin(-/-) and control mice, however, by day 30 the number of these fibers in control group was found to increase significantly, exceeding that one in transthyretin(-/-) mice by 27.9%. These results indicate the important contribution of transthyretin, as a thyroxin carrier protein, to the process of posttraumatic regeneration of sciatic nerve. The absence of changes in nerve fiber numbers in transthyretin-knockout mice in postembryonic period suggests the presence of transthyretin-independent mechanism of thyroxin transport into the peripheral nervous system.

[Post-traumatic survival in different subpopulations of sensory neurons]. / Posttravmaticheskoe vyzhivanie chuvstvitel'nykh neironov razlichnykh subpopuliatsii.

An expression of high molecular component of neurofilament triplet NF200 (marker of neurons forming A-fibers) and binding of isolectin B4 (IB4) was examined immunohistochemically in LIV-LV dorsal root ganglia (DRG) neurons after ligation or transection of the sciatic nerve in rat. NF200 immunoreactivity was detected in 15% of all neurons in DRG of intact rats. Ligation of sciatic nerve caused a two-fold decrease in number of NF200-positive neurons by 90th day after nerve injury, however in animals treated with peripheral nerve regeneration stimulator xymedon the number of surviving NF200-positive neurons was increased by 50.7% as compared with control group (nerve ligation without treatment). In DRG of intact rats 23.6% of neurons showed IB4 binding. Of the DRG neurons 2.6% were labeled by IB4 at 30th day after ligation of the nerve. At 90th day after ligation no IB4-positive neurons were revealed in DRG of untreated rats, while xymedon treatment was shown to result in more than 8-fold increase in the number of surviving IB4-positive neurons. IB4-positive neurons have greater probability of entering the posttraumatic apoptosis. After nerve ligation the survival of NF200- and IB4-positive neurons was less than that one following nerve transection, suggesting that axon lengthening could be a the factor supporting neuronal survival. Pyrimidine derivative xymedon promoted the survival of neurons in both subpopulations with predominant effect on IB4-positive neurons.

[Interaction of sensory neurons and satellite cells during stimulation of the nerve regeneration]. / Vzaimodeistvie chuvstvitel'nykh neironov i kletok-satellitov pri stimuliatsii regeneratsii nerva.

Using the model of pharmacological stimulation of rat sciatic nerve regeneration, the effect of pyrimidine derivative ximedon on survival of different populations of LIV-LV dorsal root ganglion neurons, their expression of antiapoptotic Bcl-2 protein and number of neurons surrounded by vimentin-positive satellite cell, was studied. 90 days after nerve cutting, ximedon was found to reduce posttraumatic death of neurons having both medium-sized and large perikarya by 35.5% and 42.8%, respectively, to support Bcl-2 expression in all neuronal populations and to modify vimentin expression in satellite cells. After nerve cutting, the number of small and medium-sized neurons with vimentin-expressing satellite cells was significantly increased while the population of large neurons was diminished. Nerve regeneration stimulation by ximedon induced the increase in the number of large neurons, surrounded by vimentin-expressing satellite cells, up to the levels found in intact animals.

Effect of stimulation of nerve regeneration on posttraumatic neuronal survival in dorsal root ganglia.

The survival of neurons is a key condition for complete posttraumatic regeneration of the peripheral nerve. In experiments on rats we studied survival capacity of different neuronal subpopulations in L(IV)-L(V) dorsal root ganglia after ligation or transection and suturing of the sciatic nerve. Experiments with nerve ligation showed that IB4+ neurons are more sensitive to the injury than NF200+ neurons. By day 90 after ligation of the sciatic nerve IB4+ neurons were virtually not detected in the dorsal root ganglia. By day 90 after nerve transection the number of surviving NF200+ and IB4+ neurons decreased by 26.1 and 21.4%, respectively, in comparison with intact animals. Treatment with xymedon, a regeneration stimulator, led to a 48.5% increase in the number of surviving NF200+ neurons by day 30 after ligation of the nerve and a 50.7% increase by day 90. The number of surviving IB4+ neurons increased more than 8-fold by this term after ligation of the nerve and drug stimulation. Xymedon had a neuroprotective effect towards both neuron subpopulations, more intensely preventing apoptosis of IB4+ neurons.

Effect of xymedon on cell survival in the system sensory neuron Schwann cell.

Pyrimidine derivative xymedon inhibits neuronal death in L4-L5 spinal ganglia 30 days after ligation of rat sciatic nerve. After treatment with xymedon the number of neurons on the operated side decreased by 22.1% compared to that on the contralateral side, while in the control group this parameter decreased by 28.7%. At the same terms, the number of Schwann cells on the operated side after xymedon injection increased by 27.7% in comparison with that on the contralateral side, while in the control group this parameter decreased by 57.3%.

[Drug stimulation of the peripheral nerve regeneration]. / Stimuliatsiia regeneratsii perifericheskogo nerva lekarstvennymi sredstvami.

A comparative study of the effects of xymedone, riboxin, and piracetam upon the regeneration of myelinated axons and the number of surviving sensory neurons in spinal ganglia L4-L5 was studied on the model of sciatic nerve transection in rats. The three drugs decrease the number of neurons entering the posttraumatic apoptosis. Riboxin and xymedone stimulate the regeneration of myelinated axons, the latter drug being more effective and producing a 21.3 and 14.7% increase in the number of regenerated myelinated axons and a 29.3 and 37.7% increase in the relative number of sensory neurons (the transected/contralateral side ratio) in spinal ganglia L4-L5 relative to control by the 60th and 90th day upon transection, respectively.

[Sensory neurons and Schwann cells during pharmacologic stimulation of the nerve regeneration]. / Chuvstvitel'nye neirony i Shvanovskie kletki pri farmakologicheskoi stimuliatsii regeneratsii nerva.

Using the model of rat sciatic nerve transection and crush injury we studied influence of pyrimidine derivative xymedon on efficacy of regeneration of myelinated axons, number and phenotype of surviving sensory neurons (expressing GAP-43 and Bcl-2) and Schwann cells (S100, GAP-43, PCNA) on the 7th, 15th, 30th, 60th and 90th day after nerve injury. We found out that xymedon counteracts posttraumatic death of sensory neurons, stimulates regeneration of myelinated fibres and proliferation of Schwann cells.