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Corpus cerebelli in juvenile chum salmon is a multiprojective region of the brain connected via afferent and efferent projections with the higher regions of the brainstem and synencephalon, as well as with multiprojection regions of the medulla oblongata and spinal cord. During the postembryonic development of the cerebellum in chum salmon, Oncorhynchus keta , the lateral part of the juvenile cerebellum gives rise to the caudomedial part of the definitive cerebellum, which is consistent with the data reported for zebrafish and mouse cerebellum. Thus, the topographic organization of the cerebellum and its efferents are similar between fish (chum salmon and zebrafish) and mammals, including mice and humans. The distributions of recombinant adeno-associated viral vectors (rAAVs) after an injection of the base vector into the cerebellum have shown highly specific patterns of transgene expression in bipolar neurons in the latero-caudal lobe of the juvenile chum tectum opticum . The distribution of rAAVs in the dorsal thalamus, epithalamus, nucleus rotundus , and pretectal complex indicates the targeted distribution of the transgene via the thalamo-cerebellar projections. The detection of GFP expression in the cells of the epiphysis and posterior tubercle of juvenile chum salmon is associated with the transgene's distribution and with the cerebrospinal fluid flow, the brain ventricles and its outer surface. The direct delivery of the rAAV into the central nervous system by intracerebroventricular administration allows it to spread widely in the brain. Thus, the presence of special projection areas in the juvenile chum salmon cerebellum, as well as outside it, and the identification of the transgene's expression in them confirm the potential ability of rAAVs to distribute in both intracerebellar and afferent and efferent extracerebellar projections of the cerebellum.

To date, data on the presence of adenoviral receptors in fish are very limited. In the present work, we used mouse recombinant adeno-associated viral vectors (rAAV) with a calcium indicator of the latest generation GCaMP6m that are usually applied for the dorsal hippocampus of mice but were not previously used for gene delivery into fish brain. The aim of our work was to study the feasibility of transduction of rAAV in the mouse hippocampus into brain cells of juvenile chum salmon and subsequent determination of the phenotype of rAAV-labeled cells by confocal laser scanning microscopy (CLSM). Delivery of the gene in vivo was carried out by intracranial injection of a GCaMP6m-GFP-containing vector directly into the mesencephalic tegmentum region of juvenile (one-year-old) chum salmon, Oncorhynchus keta . AAV incorporation into brain cells of the juvenile chum salmon was assessed at 1 week after a single injection of the vector. AAV expression in various areas of the thalamus, pretectum, posterior-tuberal region, postcommissural region, medial and lateral regions of the tegmentum, and mesencephalic reticular formation of juvenile O. keta was evaluated using CLSM followed by immunohistochemical analysis of the localization of the neuron-specific calcium binding protein HuCD in combination with nuclear staining with DAPI. The results of the analysis showed partial colocalization of cells expressing GCaMP6m-GFP with red fluorescent HuCD protein. Thus, cells of the thalamus, posterior tuberal region, mesencephalic tegmentum, cells of the accessory visual system, mesencephalic reticular formation, hypothalamus, and postcommissural region of the mesencephalon of juvenile chum salmon expressing GCaMP6m-GFP were attributed to the neuron-specific line of chum salmon brain cells, which indicates the ability of hippocampal mammal rAAV to integrate into neurons of the central nervous system of fish with subsequent expression of viral proteins, which obviously indicates the neuronal expression of a mammalian adenoviral receptor homolog by juvenile chum salmon neurons.