Transcriptomic reprogramming and tissue injury in the rainbow trout brain under sustained hypoxia

Prolonged hypoxia challenges brain energy homeostasis in hypoxia-sensitive fish, yet how stress-responsive transcriptional changes relate to tissue injury remains unclear. Rainbow trout (Oncorhynchus mykiss) were exposed to normoxia (8.0 mg/L dissolved oxygen) or hypoxia (3.0 mg/L) for 48 h. Responses were assessed using RNA sequencing, reverse transcription quantitative PCR (RT-qPCR), and oxidative stress indices in whole-brain samples, together with histological examination and TUNEL staining of the midbrain. Hypoxia induced 417 differentially expressed genes, including 169 upregulated and 248 downregulated genes. Upregulated genes were enriched in stress responses, glutathione metabolism, and xenobiotic metabolism, whereas downregulated genes were associated with extracellular matrix organization, focal adhesion, and cell-cycle regulation. Gene set enrichment analysis indicated relative transcriptional downregulation of gene sets associated with oxidative phosphorylation, the citrate cycle, and carbon metabolism, together with enrichment of autophagy, mitophagy, MAPK, FoxO, and neuroactive ligand-receptor signaling under hypoxia. RT-qPCR supported the RNA-seq patterns of 12 selected genes. Malondialdehyde (MDA) content increased by 73.6%, indicating enhanced lipid peroxidation. The activities of total superoxide dismutase, catalase, and glutathione peroxidase increased by 55.5%, 52.8%, and 117.3%, respectively, reflecting an enhanced antioxidant response. Hypoxic brains also exhibited localized parenchymal loosening, vacuole-like areas, and more TUNEL-positive cells, consistent with increased DNA fragmentation. Collectively, the findings showed transcriptional and enzymatic changes consistent with metabolic conservation, antioxidant defense, and cellular quality control, together with elevated MDA content and localized tissue alterations. This pattern links pathway-level transcriptional reprogramming with biochemical and histological indicators of hypoxia-associated brain injury.