Transcriptomic Profiling Revealed Signaling Pathways Associated with the Spawning of Female Zebrafish under Cold Stress

As one of the critical abiotic factors, temperature controls fish development and reproduction. However, the effects of low temperature on the transcriptional regulation of zebrafish reproduction remain largely unclear. In this study, the fecundity of zebrafish was examined after exposure to cold temperatures at 19.5 °C, 19 °C, 18.5 °C, or 18 °C. The temperature at 19 °C showed no significant influence on the fecundity of zebrafish, but temperature at 18.5 °C or 18 °C significantly blocked the spawning of females, suggesting the existence of a low temperature critical point for the spawning of zebrafish females. Based on these observations, the brains of anesthetized fish under cold stress at different cold temperatures were collected for high-throughput RNA-seq assays. Key genes, hub pathways and important biological processes responding to cold temperatures during the spawning of zebrafish were identified through bioinformatic analysis. The number of down-regulated and up-regulated genes during the temperature reduction from egg-spawning temperatures at 19.5 °C and 19 °C to non-spawning temperatures at 18.5 °C and 18 °C were 2588 and 2527 (fold change ≥ 1.5 and p-value ≤ 0.01), respectively. Venn analysis was performed to identify up- and down-regulated key genes. KEGG enrichment analysis indicated that the hub pathways overrepresented among down-regulated key genes included the GnRH signaling pathway, vascular smooth muscle contraction, C-type lectin receptor signaling pathway, phosphatidylinositol signaling system and insulin signaling pathway. GO enrichment analysis of down-regulated key genes revealed the most important biological processes inhibited under non-spawning temperatures at 18.5 °C and 18 °C were photoreceptor cell outer segment organization, circadian regulation of gene expression and photoreceptor cell maintenance. Furthermore, 99 hormone-related genes were found in the brain tissues of non-spawning and spawning groups, and GnRH signaling pathway and insulin signaling pathway were enriched from down-regulated genes related to hormones at 18.5 °C and 18 °C. Thus, these findings uncovered crucial hormone-related genes and signaling pathways controlling the spawning of female zebrafish under cold stress.