Thermal sensitivity of membrane fluidity and integrity in hearts of Antarctic fishes that vary in expression of hemoglobin and myoglobin

Antarctic notothenioids are noted for extreme stenothermy, yet underpinnings of their thermal limits are not fully understood. We hypothesized that properties of ventricular membranes could explain previously observed differences among notothenioids in temperature onset of cardiac arrhythmias and persistent asystole. Microsomes were prepared using ventricles from six species of notothenioids, including four species from the hemoglobin-less (Hb-) family Channichthyidae (icefishes), which also differentially express cardiac myoglobin (Mb), and two species from the (Hb+) Nototheniidae. We determined membrane fluidity and structural integrity by quantifying fluorescence depolarization of 1,6-diphenyl-1,3,5-hexatriene (DPH) and leakage of 5(6)-carboxyfluorescein, respectively, over a temperature range from ambient (0 °C) to 20 °C. Compositions of membrane phospholipids and cholesterol contents were also quantified. Membranes from all four species of icefishes exhibited greater fluidity than membranes from the red-blooded species N. coriiceps. Thermal sensitivity of fluidity did not vary among species. The greatest thermal sensitivity to leakage occurred between 0 and 5 °C for all species, while membranes from the icefish, Chaenocephalus aceratus (Hb-/Mb-) displayed leakage that was nearly 1.5-fold greater than leakage in N. coriiceps (Hb+/Mb+). Contents of phosphatidylethanolamine (PE) were approximately 1.5-fold greater in icefishes than in red-blooded fishes, and phospholipids had a higher degree of unsaturation in icefishes than in Hb + notothenioids. Cholesterol contents were lowest in Champsocephalus gunnari (Hb-/Mb-) and highest in the two Hb+/Mb + species, G. gibberifrons and N. coriiceps. Our results reveal marked differences in membrane properties and indicate a breach in membrane fluidity and structural integrity at a lower temperature in icefishes than in red-blooded notothenioids.

南极南极大头鱼亚目（Antarctic notothenioids）以极端狭温性（extreme stenothermy）著称，但其热耐受极限的内在机制尚未完全明晰。我们提出假说：心室膜（ventricular membranes）的特性可解释此前观测到的南极大头鱼亚目物种间，在心律失常（cardiac arrhythmias）发生温度阈值与持续性心搏停止（persistent asystole）临界温度上的差异。本研究以6种南极大头鱼亚目鱼类的心室组织为材料制备微粒体（microsomes），其中4种隶属于无血红蛋白（Hb-）的冰鱼科（Channichthyidae，俗称冰鱼），该类群的心肌肌红蛋白（Mb）表达水平存在分化；另外2种属于有血红蛋白（Hb+）的南极鱼科（Nototheniidae）。我们在0℃至20℃的温度梯度下，分别通过定量1,6-二苯基-1,3,5-己三烯（DPH）的荧光去极化（fluorescence depolarization）值，以及5(6)-羧基荧光素（5(6)-carboxyfluorescein）的泄漏量，来表征膜流动性（membrane fluidity）与结构完整性（structural integrity），同时定量分析了膜磷脂（phospholipids）组成与胆固醇（cholesterol）含量。4种冰鱼的心室膜流动性均显著高于红血鱼种南极岩本鱼（N. coriiceps），不同物种的膜流动性热敏感性并无显著差异。所有物种的膜泄漏量热敏感性峰值均出现在0~5℃区间；其中革首冰鱼（Chaenocephalus aceratus，Hb-/Mb-）的膜泄漏量约为南极岩本鱼（N. coriiceps，Hb+/Mb+）的1.5倍。冰鱼的磷脂酰乙醇胺（PE）含量约为红血鱼种的1.5倍，且冰鱼膜磷脂的不饱和程度高于有血红蛋白的南极大头鱼亚目物种。贡氏颌须冰鱼（Champsocephalus gunnari，Hb-/Mb-）的膜胆固醇含量最低，而2种Hb+/Mb+物种——驼背南极鱼（G. gibberifrons）与南极岩本鱼的胆固醇含量最高。本研究结果揭示了不同南极大头鱼亚目物种间膜特性的显著差异，并表明冰鱼的膜流动性与结构完整性会在更低温度下就出现破损，这一临界温度早于红血的南极鱼科物种。