Hypoxia is a serious issue that affects the marine environment, with a growing number of hypoxic “dead zones” occurring each year. Reports have indicated that hypoxia is detrimental to the reproductive function and sexual development of fish via the disruption of endocrine signaling in organs involved in the hypothalamus–pituitary–gonad axis, including the brain. While we previously reported that hypoxia induces transcriptome-wide alterations in the brain of marine medaka (Oryzias melastigma), whether these effects were reflected at the protein level remains unclear. Therefore, the present study used high-throughput proteomic sequencing along with bioinformatics analysis to assess the short-term and multi-generational effects of hypoxia on the brain proteome of O. melastigma. We identified 36,567 peptides and 7,599 proteins (1% false discovery rate in brain samples), with functions involved in cellular and metabolic processes such as signaling and reproductive processes as well as energy production and conversion. Furthermore, we determined that hypoxia resulted in the significant differential expressions of 33 upregulated and 69 downregulated proteins in the short-term exposure group and 24 upregulated and 52 downregulated proteins in the multi-generational exposure group. Pathway enrichment analysis of the deregulated proteins indicated that hypoxia could impair brain function by altering arachidonic acid metabolism, tight junctions, and adrenergic signaling under short-term hypoxic exposure and by altering p53 and PI3K–Akt signaling under multi-generational hypoxic exposure, which may lead to the onset of neurodegenerative disorders including Alzheimer’s disease and amyotrophic lateral sclerosis. Ingenuity pathway analysis of the deregulated proteins showed that hypoxia affected common signaling pathways in the brain (e.g., integrin, paxillin, and epithelial adherens junction signaling) under both short-term and multi-generational exposures. Hypoxia also deregulated pathways specific to short-term exposure (including integrin-linked kinase, calcium, and integrin signaling) and multi-generational exposure (including sphingosine-1-phosphate signaling, endocannabinoid neuronal synapse pathway, and endoplasmic reticulum stress pathway). Overall, our results provide additional insights into the mechanisms of hypoxia disrupting neuronal function at the protein level in marine medaka.
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