Field M L, Azzawi A, Unitt J F, Seymour A M, Henderson C, Radda G K
Department of Biochemistry, University of Oxford, Oxfordshire, UK.
J Mol Cell Cardiol. 1996 Jan;28(1):65-77. doi: 10.1006/jmcc.1996.0007.
Fluorescence and 31P magnetic resonance spectroscopy have been used to monitor simultaneously, the [Ca2+]i staircase and high energy phosphate metabolism in isolated Langendorff-perfused rat heart paced at 2, 4 and 6 Hz. In order to investigate further the relationship between high energy phosphate metabolism and the calcium staircase we perturbed the intracellular phosphocreatine (PCr)/creatine concentration with dietary beta-guanidinopropionic acid (beta-GPA). We have observed that: (a) At 2 Hz stimulation, the ventricular -Ca2+-i-dependent fluorescence decay is biexponential and continues to decay throughout the interstimulus interval; (b) at 4 Hz and 6 Hz, the [Ca2+]i decay is monoexponential; (c) end-diastolic [Ca2+]i is elevated at higher stimulation frequencies; (d) net [Ca2+]i flux per cycle is reduced at higher stimulation frequencies and is therefore correlated inversely with stimulation frequency and end-diastolic [Ca2+]i; (e) "heart rate * [Ca2+]i flux product" which is a measure of the work done in cycling calcium, is directly proportional to stimulation frequency; (f) the hysteresis between peak ventricular isovolumic pressure and peak fluorescence is decreased at higher stimulation frequencies; (g) no correlation was detected between the PCr/ATP ratio and stimulation frequency; (h) despite a 60% decrease in the myocardial PCr/ATP ratio after beta-GPA feeding, rat heart is able to maintain the end-diastolic [Ca2+]i-dependent fluorescence, and therefore the [Ca2+]i staircase relationship, similar to that of normal rat heart. In conclusion, using a physiological stimulation range and substrate supply we have observed a negative staircase of both [Ca2+]i and isovolumic pressure in whole heart which is not hypoxic. We propose that the inability of the sarcoplasmic reticulum to sequester sufficient cytosolic calcium at high stimulation frequencies leads to an elevation in end-diastolic [Ca2+]i, decreased net calcium flux per cycle resulting in a negative [Ca2+]i staircase and thus a negative isovolumic pressure-frequency relationship. We did not detect any correlation between steady-state high energy phosphate metabolism and stimulation frequency.
荧光和31P磁共振波谱已被用于同时监测在2、4和6Hz频率起搏的离体Langendorff灌注大鼠心脏中的[Ca2+]i阶梯和高能磷酸代谢。为了进一步研究高能磷酸代谢与钙阶梯之间的关系，我们用膳食β-胍基丙酸（β-GPA）扰乱细胞内磷酸肌酸（PCr）/肌酸浓度。我们观察到：（a）在2Hz刺激下，心室Ca2+依赖的荧光衰减是双指数的，并且在整个刺激间期持续衰减；（b）在4Hz和6Hz时，[Ca2+]i衰减是单指数的；（c）在较高刺激频率下舒张末期[Ca2+]i升高；（d）在较高刺激频率下每个周期的净[Ca2+]i通量降低，因此与刺激频率和舒张末期[Ca2+]i呈负相关；（e）“心率×[Ca2+]i通量乘积”（衡量钙循环所做的功）与刺激频率成正比；（f）在较高刺激频率下，心室等容压力峰值与荧光峰值之间的滞后减小；（g）未检测到PCr/ATP比值与刺激频率之间的相关性；（h）尽管喂食β-GPA后心肌PCr/ATP比值降低了60%，大鼠心脏仍能够维持舒张末期Ca2+依赖的荧光，因此维持[Ca2+]i阶梯关系，与正常大鼠心脏相似。总之，使用生理刺激范围和底物供应，我们在非缺氧的全心中观察到了[Ca2+]i和等容压力的负阶梯现象。我们提出，在高刺激频率下肌浆网无法摄取足够的胞浆钙导致舒张末期[Ca2+]i升高，每个周期的净钙通量降低，从而导致[Ca2+]i负阶梯，进而导致等容压力-频率负相关。我们未检测到稳态高能磷酸代谢与刺激频率之间的任何相关性。
