Two distinct nonapeptide systems, vasotocin- and oxytocin-related peptides, evolved in vertebrates. Their role in male zebrafish reproduction has not been formally investigated. We hypothesized that the teleost nonapeptides vasotocin and isotocin stimulate male zebrafish reproductive physiology and success by affecting central neuronal and/or peripheral endocrine pathways. Pharmacological inhibition experiments revealed that both vasotocin and isotocin contribute significantly to male reproductive success, which in the case of vasotocin correlated significantly with indices of male courtship behavior. Interestingly, co-administration of vasotocin and isotocin antagonists completely abolished male reproductive success without affecting male courtship behavior and endocrine indices, possibly linked to a synergistic action of nonapeptides on male pheromone release. To further probe the nonapeptides' role in male zebrafish reproduction, we subsequently tested whether male zebrafish nonapeptide systems were acutely activated by the female releaser pheromone PGF_2α, a strong chemoattractant and important reproductive cue in males which stimulates courtship behavior. Male zebrafish attracted to PGF_2α in a choice assay exhibited acute increases in neuronal activation marker p-ERK immunoreactivity in the ventral glomerulus of the olfactory bulb and the preoptic area, however no co-localization with isotocin was observed. Conversely, PGF_2α time-dependently stimulated whole brain isotocin mRNA abundance, suggesting secondary longer-term effects of PGF_2α exposure on the central isotocinergic system. While the current lack of vasotocin-specific antibodies for zebrafish does not allow to probe acute activation of vasotocinergic neurons, whole brain vasotocin mRNA was not significantly affected by PGF_2α exposure. Together, our results identify a role for nonapeptides in male zebrafish reproductive physiology and success.

脊椎动物进化出两种截然不同的九肽系统，即与血管毒素和催产素有关的肽。它们在雄性斑马鱼繁殖中的作用尚未得到正式研究。我们假设硬骨鱼九肽血管毒素和异麦芽酮素通过影响中枢神经元和/或外周内分泌途径刺激雄性斑马鱼的生殖生理和成功。药理抑制实验表明，vasotocin和isotocin均对男性生殖成功做出了重要贡献，在vasotocin的情况下，这与男性求爱行为指标显着相关。有趣的是，同时使用血管毒素和异氰酸酯拮抗剂可以完全消除男性的生殖成功，而不会影响男性的求爱行为和内分泌指标，可能与九肽对雄性信息素释放的协同作用有关。为了进一步探讨九肽在雄性斑马鱼繁殖中的作用，我们随后测试了雌性释放信息素PGF是否能急性激活雄性斑马鱼的九肽系统。_2α，一种强烈的趋化性和重要的生殖线索，在男性中刺激求爱行为。雄性斑马鱼吸引到PGF _2α在选择分析中显示在嗅球和视前区的腹侧肾小球中的神经元激活标记P-ERK的免疫反应性增加急性然而没有观察到共定位isotocin。相反，_PGF2α时间依赖性地刺激了全脑同位素MRNA的丰度，表明_PGF2α暴露对中枢同位素能系统具有长期的次级影响。虽然目前缺乏针对斑马鱼的血管生成素特异性抗体无法探测到血管生成素能神经元的急性激活，但全脑血管生成素的mRNA并未受到PGF的明显影响_2α暴露。总之，我们的结果确定了九肽在雄性斑马鱼生殖生理和成功中的作用。