Ultrastructural Organization and Immunohistochemical Analysis of Bcl-2, Caspase 3, and NeuroN Localization in the Spinal Cord of Juvenile Chum Salmon Oncorhynchus keta in Intact Growth and After Traumatic Injury

The localization of the proapoptotic (caspase 3) and the antiapoptotic B-cell lymphoma 2 (Bcl-2) factors in the spinal cord of 2-year-old juvenile chum salmon Oncorhynchus keta, as well as the neuron-specific marker NeuroN, was studied using IHC labeling. Scanning electron microscopy was used to obtain three-dimensional, highly detailed images of the surface structure of the primary motor neurons (MT1) and secondary motor neurons (MT2) of the ventral spinal cord column (VSCC), as well as nerve endings and interneuronal connections in the VSCC. The spatial organization of VSCC neurons, astrocytes, and the surface characteristics of membranes and cell connections in the spinal cord were examined. The localization of apoptotic markers and NeuroN was studied in intact juveniles and three days after traumatic spinal cord injury. The results showed that the pro-apoptotic factor caspase 3 (cas3) in the spinal cord of intact animals had a rather limited localization in the perikarya of primary and secondary motor neurons of the VSCC. Cas3 immunolocalization in the spinal cord of juvenile chum salmon was represented in astrocytes located within the VSCC, as well as among the fibers of the white matter. SEM revealed the morphological features of perineuronal astrocytes of the VSCC. Cas3+ microvascular sites were identified in the spinal cord, and their reactivity increased markedly after traumatic injury. Cas3 synthesis increased in the synaptic projections of VSCC neurons, as well as in blood vessels after traumatic injury. Along with various types of neurons, cas3 was expressed after injury in fibers and small adult neural stem and progenitor cells (aNSPCs) located in the dorsal horn and submarginal regions. The antiapoptotic factor Bcl-2 in intact juvenile chum salmon is relatively widespread in the perikarya of VSCC neurons, white matter interneurons, and dorsal horn cells located in the submarginal zone. In the post-traumatic period, Bcl-2 expression increased significantly both in VSCC neurons and in cells and fibers of the dorsal horn. Along with proapoptotic and antiapoptotic factors, the number of undifferentiated NeuroN-producing cells in the spinal cord of juvenile chum salmon increased dramatically in the post-traumatic period, indicating an active neurogenic post-traumatic process. Thus, in intact animals, proapoptotic and antiapoptotic factors are synthesized during growth; the balance between them determines the successful elimination of cells that have not established connections during ontogenetic growth and regulates programmed cell death. An increase in the number of intensely immunolabeled cells with an undifferentiated phenotype in the post-traumatic period indicates an intensive regenerative process, consistent with the results obtained in other models.