Cardiovascular Performance and Excitation-Contraction Couplng in Tuna Hearts

金枪鱼以其特殊的生理和形态适应能力而闻名，这使它有别于其他硬骨鱼。这些特长包括独特的游泳运动学、高代谢率、高心输出量和区域吸热。许多研究已经证明了金枪鱼心脏在器官和组织水平上的非凡表现，但人们对金枪鱼心脏在细胞水平上的功能知之甚少。这个项目中的实验将检验金枪鱼心脏在兴奋-收缩(EC)耦合方面具有专门化的假设，这是增强性能的基础。吸热型金枪鱼(蓝鳍金枪鱼和黄鳍金枪鱼)和它们的外温型近亲(马克鱼和鳄鱼)的心脏功能将在分子、细胞和组织水平上进行检查。心肌收缩依赖于钙离子向肌丝的精确输送，而松弛依赖于胞浆中钙离子的去除。在哺乳动物和鸟类的心肌细胞中，肌浆网(SR)的细胞内钙库缩短了钙循环的扩散距离，缩短了钙循环的时间进程，使心率高于低等四足动物。在鱼类中，体内和外部钙储存的贡献随着物种和温度的不同而不同，但目前还没有关于金枪鱼心脏功能的细胞研究。这个项目将解决这一假设，即金枪鱼的高最大心率部分是通过增加对细胞内SR存储的依赖和在EC偶联过程中增加钙诱导的钙释放来实现的。此外，还将对冷温带金枪鱼和暖温带金枪鱼及其体温较低的姐妹类群的心血管功能对急性温度变化的反应进行实验比较。这种比较将检验这样一种假设，即黄鳍金枪鱼的钙循环比蓝鳍金枪鱼更敏感。黄鳍金枪鱼仅限于热带和暖温带水域。因此，将采用一种综合的方法，利用分子、生化、结构和整体生物体生理技术。采用全细胞电压钳技术，研究金枪鱼和鲱鱼肌膜钙通道的L钙电流，并评价其对温度的敏感性。共聚焦显微镜将被用来显示从不同物种分离的携带Fluo-4的心肌细胞在不同温度下的全细胞钙瞬变，整个生物体的表现将通过原位测量灌流的蓝鳍金枪鱼和鳄鱼心脏的心血管表现来检验。总体而言，这项研究将提供关于Scomridae科鱼类心脏功能的比较数据，并将具体描述吸热和外温姐妹类心脏细胞中电压敏感的钙通道、SR钙释放通道和钙ATPase泵的热关系。这一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。长期的目标是了解吸热和高代谢率在梳状血统中的进化。这项拟议的研究还将深入了解心脏专门化在增加蓝鳍金枪鱼生态位呼吸方面所起的作用。这项研究将有助于对地球上最具经济价值的动物之一的一小群鱼的基本知识。该项目的信息将通过出版物、热门文章、网站和蒙特利湾水族馆的公开展览进行传播。金枪鱼研究和保护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教育公众有关金枪鱼生物学和保护的努力做出贡献。

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.