Expression of Doublecortin, Glial Fibrillar Acidic Protein, and Vimentin in the Intact Subpallium and after Traumatic Injury to the Pallium in Juvenile Salmon, Oncorhynchus masou.

Fetalization associated with a delay in development and the preservation of the features of the embryonic structure of the brain dominates the ontogeny of salmonids. The aim of the present study was to comparatively analyze the distribution of the glial-type aNSC markers such as vimentin and glial fibrillar acidic protein (GFAP) and the migratory neuronal precursors such as doublecortin in the telencephalon subpallium of juvenile masu salmon, Oncorhynchus masou, in normal conditions and at 1 week after an injury to the dorsal pallium. Immunohistochemical labeling of vimentin, GFAP, and doublecortin in the pallium of intact juvenile masu salmon revealed single cells with similar morphologies corresponding to a persistent pool of neuronal and/or glial progenitors. The study of the posttraumatic process showed the presence of intensely GFAP-labeled cells of the neuroepithelial type that form reactive neurogenic zones in all areas of the subpallial zone of juvenile masu salmon. A comparative analysis of the distribution of radial glia in the dorsal, ventral, and lateral zones of the subpallium showed a maximum concentration of cells in the dorsal part of subpallium (VD) and a minimum concentration in the lateral part of subpallium VL. An essential feature of posttraumatic immunolabeling in the masu salmon subpallium is the GFAP distribution patterns that are granular intracellular in the apical periventricular zone (PVZ) and fibrillar extracellular in the subventricular (SVZ) and parenchymal zones (PZ). In contrast to those in intact animals, most of the GFAP+ granules and constitutive neurogenic niches in injured fish were localized in the basal part of the PVZ. With the traumatic injury to the subpallium, the number of Vim+ cells in the lateral and ventral regions significantly increased. At 1 week post-injury, the total immunolabeling of vimentin cells in the PVZ was replaced by the granular pattern of Vim immunodistribution spreading from the PVZ to the SVZ and deeper parenchymal layers of the brain in all areas of the subpallium. A significant increase in the number of DC+ cells was observed also in all areas of the subpallium. The number of cells increased both in the PVZ and in the SVZ, as well as in the deeper PZ. Thus, at 1 week after the injury to the dorsal pallium, the number of DC, Vim, and GFAP expressing cells of the neuroepithelial type in the subpallium of juvenile masu salmon increased, and additionally GFAP+ radial glia appeared in VD, which was absent from intact animals.

Mechanical Brain Injury Increases Cells' Production of Cystathionine ß-Synthase and Glutamine Synthetase, but Reduces Pax2 Expression in the Telencephalon of Juvenile Chum Salmon, Oncorhynchus keta.

The considerable post-traumatic brain recovery in fishes makes them a useful model for studying the mechanisms that provide reparative neurogenesis, which is poorly represented in mammals. After a mechanical injury to the telencephalon in adult fish, lost neurons are actively replaced due to the proliferative activity of neuroepithelial cells and radial glia in the neurogenic periventricular zone. However, it is not enough clear which signaling mechanisms are involved in the activation of adult neural stem cells (aNSC) after the injury (reactive proliferation) and in the production of new neurons (regenerative neurogenesis) from progenitor cells (NPC). In juvenile Pacific salmon, the predominant type of NSCs in the telencephalon are neuroepithelial cells corresponding to embryonic NSCs. Expression of glutamine synthetase (GS), a NSC molecular marker, was detected in the neuroepithelial cells of the pallium and subpallium of juvenile chum salmon, Oncorhynchus keta. At 3 days after a traumatic brain injury (TBI) in juvenile chum salmon, the GS expression was detected in the radial glia corresponding to aNSC in the pallium and subpallium. The maximum density of distribution of GS+ radial glia was found in the dorsal pallial region. Hydrogen sulfide (H2S) is a proneurogenic factor that reduces oxidative stress and excitotoxicity effects, along with the increased GS production in the brain cells of juvenile chum salmon. In the fish brain, H2S producing by cystathionine ß-synthase in neurogenic zones may be involved in maintaining the microenvironment that provides optimal conditions for the functioning of neurogenic niches during constitutive neurogenesis. After injury, H2S can determine cell survivability, providing a neuroprotective effect in the area of injury and reducing the process of glutamate excitotoxicity, acting as a signaling molecule involved in changing the neurogenic environment, which leads to the reactivation of neurogenic niches and cell regeneration programs. The results of studies on the control of the expression of regulatory Sonic Hedgehog genes (Shh) and the transcription factors Paired Box2 (Pax2) regulated by them are still insufficient. A comparative analysis of Pax2 expression in the telencephalon of intact chum salmon showed the presence of constitutive patterns of Pax2 expression in neurogenic areas and non-neurogenic parenchymal zones of the pallium and subpallium. After mechanical injury, the patterns of Pax2 expression changed, and the amount of Pax2+ decreased (p < 0.05) in lateral (Dl), medial (Dm) zones of the pallium, and the lateral zone (Vl) of the subpallium compared to the control. We believe that the decrease in the expression of Pax2 may be caused by the inhibitory effect of the Pax6 transcription factor, whose expression in the juvenile salmon brain increases upon injury.

Proliferation, Adult Neuronal Stem Cells and Cells Migration in Pallium during Constitutive Neurogenesis and after Traumatic Injury of Telencephalon of Juvenile Masu Salmon, Oncorhynchus masou.

A study of the lateral pallium in zebrafish and the visual tectum of the medaka revealed a population of adult neuroepithelial (NE) cells supported from the early stage of development to various postembryonic stages of ontogenesis. These data emphasize the importance of non-radial glial stem cells in the neurogenesis of adult animals, in particular fish. However, the distribution, cell cycle features, and molecular markers of NE cells and glial progenitors in fish are still poorly understood at the postembryonic stages of ontogenesis. Fetalization predominates in the ontogenetic development of salmon fish, which is associated with a delay in development and preservation of the features of the embryonic structure of the brain during the first year of life. In the present work, we studied the features of proliferation and the migration of neuronal precursors in the pallial proliferative zone of juvenile Oncorhynchus masou. The aim of the study is a comparative analysis of the distribution of glial-type aNSCs markers, such as vimentin and glial fibrillar acid protein GFAP, as well as the proliferation marker BrdU and migratory neuronal precursor doublecortin, in the pallial zone of the intact telencephalon in juvenile O. masou normal and after mechanical injury. The immunohistochemical IHC labeling with antibodies to vimentin, GFAP and doublecortin in the pallium of intact fish revealed single, small, round and oval immunopositive cells, that correspond to a persistent pool of neuronal and/or glial progenitors. After the injury, heterogeneous cell clusters, radial glia processes, single and small intensely labeled GFAP+ cells in the parenchyma of Dd and lateral part of pallium (Dl) appeared, corresponding to reactive neurogenic niches containing glial aNSCs. A multifold increase in the pool of Vim+ neuronal precursor cells (NPCs) resulting from the injury was observed. Vim+ cells of the neuroepithelial type in Dd and Dm and cells of the glial type were identified in Dl after the injury. Doublecortine (Dc) immunolabeling after the injury revealed the radial migration of neuroblasts into Dm from the neurogenic zone of the pallium. The appearance of intensely labeled Dc+ cells in the brain parenchyma might indicate the activation of resident aNSCs as a consequence of the traumatic process.