Cardiovascular Performance and Excitation-Contraction Couplng in Tuna Hearts

金枪鱼以其独特的生理和形态适应性而闻名，使这一群体（金枪鱼科，鲭鱼科）与其他硬骨鱼鱼类区别开来。这些特点包括独特的游泳运动学、高代谢率、高心输出量和局部恒温。许多研究已经证明了金枪鱼心脏在器官和组织水平上的卓越表现，然而对金枪鱼心脏在细胞水平上的功能知之甚少。这个项目的实验将测试金枪鱼心脏在兴奋-收缩（EC）耦合方面的特殊性，这是增强性能的基础。心脏性能将在分子，细胞和有机体水平上检查吸热金枪鱼（蓝鳍金枪鱼和黄鳍金枪鱼）及其恒温亲属（鲭鱼和鲣鱼）。心脏收缩依赖于Ca2+的精确传递到肌丝，而松弛依赖于Ca2+从细胞质中去除。在哺乳动物和鸟类的心脏细胞中，肌浆网（SR）的细胞内Ca2+储存减少了扩散距离并缩短了Ca2+循环的时间过程，从而允许比低等四足动物更高的心率。在鱼类中，内部和外部Ca2+储存的贡献因物种和温度而异，但没有关于金枪鱼心脏性能的细胞研究。该项目将解决这样一个假设，即金枪鱼的高最大心率部分是通过增加对细胞内SR储存的依赖和EC耦合期间钙诱导的Ca2+释放的增加而实现的。此外，还将实验比较寒、暖温带金枪鱼及其异温姊妹类群对急性温度变化的心血管功能。这样的比较将验证一种假设，即生活在热带和暖温带水域的黄鳍金枪鱼体内的Ca2+循环比蓝鳍金枪鱼对温度更敏感。因此，一个综合的方法利用分子，生化，结构和整个有机体生理技术将被采用。全细胞电压钳技术将通过l型Ca2+电流表征肌膜Ca2+进入，并评估其在金枪鱼和鲭鱼中的温度敏感性。共聚焦显微镜将用于观察从不同物种分离的满载氟-4的肌细胞在不同温度下的全细胞Ca2+瞬态，整个生物体的性能作为温度的函数将通过测量灌注的蓝鳍金枪鱼和鲣鱼心脏的原位心血管性能来检查。总的来说，本研究将提供关于Scombridae鱼类心脏功能的比较数据，并将具体表征来自恒温和恒温姐妹类群的心脏细胞中电压敏感的Ca2+通道，SR Ca2+释放通道和Ca2+ atp酶泵的热关系。结果将增加我们对硬骨鱼心脏如何在急性温度变化中实现高代谢率和维持功能的理解。长期的目标是了解在杂交种谱系中恒温和高代谢率的进化。拟议的研究还将产生关于心脏特化在增加蓝鳍金枪鱼生态位呼吸中所起作用的见解。这项研究将有助于了解地球上最具经济价值的动物之一的一小群鱼类的基本知识。这个项目的信息将通过出版物、流行文章、网站和蒙特利湾水族馆的公共展览来传播。金枪鱼研究与保护学院的教职员工和学生将全年提供公开讲座，并为蒙特利湾水族馆的工作做出贡献，向公众宣传金枪鱼生物学和保护知识。

Tunas are well known for the exceptional physiological and morphological adaptations that set this group (Thunnini, Scombridae) apart from other teleost fishes. These specializations include unique swimming kinematics, high metabolic rates, high cardiac outputs, and regional endothermy. A number of studies have demonstrated the exceptional performance of the tuna heart at the organ and tissue levels, however little is known about tuna cardiac function at the cellular level. Experiments in this project will test the hypothesis that tuna hearts have specializations in excitation-contraction (EC) coupling that underlie the enhanced performance. Cardiac performance will be examined at the molecular, cellular and organismal levels in endothermic tunas (bluefin and yellowfin) and their ectothermic relatives (mackerel and bonito). Cardiac contraction depends on the precise delivery of Ca2+ to the myofilaments, while relaxation depends on removal of Ca2+ from the cytoplasm. In heart cells of mammals and birds, the intracellular Ca2+ stores of the sarcoplasmic reticulum (SR) reduce diffusion distances and shorten the time course of Ca2+ cycling, permitting higher heart rates than in lower tetrapods. In fish, the contribution of internal and external Ca2+ stores varies with species and temperature, but no cellular studies of tuna heart performance exist. This project will address the hypothesis that the high maximal heart rates of tunas are achieved in part by increased reliance on intracellular SR stores and increased calcium-induced Ca2+ release during EC coupling. In addition, experimental comparisons will be made of cardiovascular function in cold and warm temperate tuna and their ectothermic sister taxa in response to acute temperature change. Such comparisons will test the hypothesis that Ca2+ cycling in yellowfin tunas, which are confined to tropical and warm temperate waters, is more temperature sensitive than in bluefin tunas. Thus, an integrative approach utilizing molecular, biochemical, structural and whole organismal physiological techniques will be employed. Whole-cell voltage-clamp techniques will be used to characterize sarcolemma Ca2+ entry via the L-type Ca 2+ current, and assess its temperature sensitivity in tunas and mackerel. Confocal microscopy will be used to visualize whole cell Ca2+ transients at different temperatures in fluo-4 loaded myocytes isolated from different species Whole organism performance as a function of temperature will be examined by measuring in situ cardiovascular performance of perfused bluefin tuna and bonito hearts. Overall this study will provide comparative data on cardiac function in fishes of the family Scombridae, and will characterize specifically the thermal relationships of voltage-sensitive Ca2+ channels, SR Ca2+ release channels, and the Ca2+ ATPase pump in heart cells from endothermic and ectothermic sister taxa. The results will increase our understanding of how teleost hearts achieve high metabolic rate and maintain function during acute temperature changes. The long-term objective is to understand the evolution of endothermy and high metabolic rate in the scombrid lineage. The proposed research also will yield insights about the role played by cardiac specializations in increasing niche breath of bluefin tuna. The research will contribute to basic knowledge about a small group of fishes that are among the most economically valuable animals on Earth. Dissemination of information from this project will occur by publications, popular articles, a website and public exhibits at the Monterey Bay Aquarium. The Tuna Research and Conservation faculty and students will provide public lectures throughout the year and contribute to efforts at the Monterey Bay Aquarium to educate the public about tuna biology and conservation.