Efforts to study the effects of environmental change on early development of Antarctic fishes would benefit from the ability to control reproduction by in vitro fertilization. Here, we established a spectrophotometric method to determine spermatozoa concentration in the semen of the blackfin icefish Chaenocephalus aceratus and we evaluated the potential for short (refrigeration) and long-term (cryopreservation) storage. The former will maximize the use of semen over several hours and the latter alleviate the risk that semen is unavailable when a spawning female is available. Seminal plasma osmolality and pH of GnRH-injected male semen were 423.65 ± 35.6 mOsm kg−1 and 8.05 ± 0.11, respectively. The predictive power of regression models between spermatozoa concentration and absorbance was determined from 200 to 600 nm and revealed a best-fitted correlation at 505 nm. The standard curve was tested against measured concentrations and the average difference between measured and estimated concentrations was 0.27 ± 10.23%. Semen could be kept on ice for up to 6 h with minimal loss (< 10%) in motility and could also be efficiently cryopreserved with > 90% post-thawing motility using a salt-based extender (pH 7.5–7.85 and ~ 280 mOsm kg−1) and 10% DMSO after 72 h immersed in liquid nitrogen. Spermatozoa morphometry revealed significant inter-individual variability. Average head length, head width, and flagella length were 3.78 ± 1.03, 2.87 ± 0.78, and 17.75 ± 7.15 µm, respectively. Spermatids composed 9.7 ± 0.8% of total spermatogenic cells, probably indicating active spermatogenesis, but perhaps resulting to some degree from the semen extraction technique used. Variations on these baseline methods may facilitate semen preservation and ultimately in vitro fertilization in this and other notothenioid species, thereby enhancing our understanding of the responses of early-life stages of Antarctic fishes to future warming of the Southern Ocean.

中文翻译：

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控制黑鳍冰鱼 Chaenocephalus aceratus (Lönnberg 1906) 的繁殖：精子浓度的测定和精液短期和长期保存的评价

研究环境变化对南极鱼类早期发育影响的努力将受益于体外受精控制繁殖的能力。在这里，我们建立了一种分光光度法来确定黑鳍冰鱼 Chaenocephalus aceratus 精液中的精子浓度，并评估了短期（冷藏）和长期（冷冻保存）储存的潜力。前者将在几个小时内最大限度地利用精液，而后者则可以降低在产卵雌性可用时无法获得精液的风险。注射 GnRH 的男性精液的精浆渗透压和 pH 值分别为 423.65 ± 35.6 mOsm kg-1 和 8.05 ± 0.11。精子浓度和吸光度之间回归模型的预测能力在 200 到 600 nm 范围内确定，并揭示了 505 nm 处的最佳拟合相关性。标准曲线根据测量浓度进行测试，测量浓度和估计浓度之间的平均差异为 0.27 ± 10.23%。精液可以在冰上保存长达 6 小时，运动损失最小（< 10%），也可以使用盐基补充剂（pH 7.5–7.85 和 ~ 280 mOsm）有效冷冻保存 > 90% 的解冻后运动kg-1) 和 10% DMSO 浸入液氮 72 小时后。精子形态测定显示显着的个体间变异性。平均头部长度、头部宽度和鞭毛长度分别为 3.78 ± 1.03、2.87 ± 0.78 和 17.75 ± 7.15 µm。精子组成 9.7 ± 0。占总生精细胞的 8%，可能表明精子发生活跃，但可能在一定程度上是由于使用的精液提取技术造成的。这些基线方法的变化可能会促进该物种和其他类拟海藻物种的精液保存和最终体外受精，从而增强我们对南极鱼类早期生命阶段对未来南大洋变暖的反应的理解。