CYTOSOLIC ADENYLATES AND ADENOSINE RELEASE IN PERFUSED WORKING HEART - COMPARISON OF WHOLE TISSUE WITH CYTOSOLIC NONAQUEOUS FRACTIONATION ANALYSES
CYTOSOLIC ADENYLATES AND ADENOSINE RELEASE IN PERFUSED WORKING HEART - COMPARISON OF WHOLE TISSUE WITH CYTOSOLIC NONAQUEOUS FRACTIONATION ANALYSES
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DOI：
10.1111/j.1432-1033.1986.tb09854.x
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发表时间：
1986-08-15
期刊：
影响因子：
--
通讯作者：
SOBOLL, S
中科院分区：
文献类型：
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作者：
BUNGER, R;SOBOLL, S
检测游离胞质腺苷酸与豚鼠灌注心脏释放的腺苷和肌苷的关系。将冷冻夹持心脏的全组织腺苷酸数据与非水介质中匀浆心肌亚细胞分离获得的相应值进行定量比较。采用高效液相色谱法测定心脏静脉积液中的腺苷和肌苷。心脏在其自然流动时，受到各种负荷、底物和儿茶酚胺的影响，在广泛的生理范围内改变心肌能量代谢和呼吸。非水细胞质ATP和磷酸肌酸（CrP）分别占心肌总含量的80%以上。当细胞外Pi校正后，胞浆CrP/Piratio与整体组织CrP/Piratio接近定量一致。这是确凿的证据，证明ATP、CrP和pii主要位于充氧良好的心肌细胞的细胞质中。心肌摄氧量（MVO2）与CrP （[CrP]/[Cr] × [Pi]）和ATP （[ATP]/[ADP] × [Pi]）的磷酸化状态呈负相关，假设肌酸激酶在接近恒定的pH值为7.2的接近平衡状态。另一方面，在ATP水平几乎不变或仅略有下降的情况下，计算出的平均游离胞浆ADP浓度随着mvo2的增加基本线性增加至三倍；这是根据整个组织和特殊的亚细胞分离数据发现的。采用肌激酶质量作用比，用平均游离胞质ADP浓度代替心脏总ADP，结果表明，平均游离胞质AMP浓度在纳摩尔范围内，即比整体组织AMP含量低3个数量级。因此，我们提出，在常氧心脏中，AMP主要位于线粒体室。然而，胞内游离AMP浓度和腺苷+肌苷的释放明显与心脏呼吸速率呈正方函数或更高幂函数。另一方面，嘌呤核苷平均释放量与计算的胞质游离AMP浓度呈线性相关（r= 0.920）。我们的观察似乎表明，胞浆中游离ADP和AMP的浓度是肌苷和冠状动脉血管舒张剂腺苷产生的主要决定因素。可以想象，AMP降解的这种动力学控制可以使游离腺苷酸/腺苷系统对机械性能、总能量输出和/或心肌的能量状态具有高度适应性。自由ATP/ADP系统的能量相关变化似乎也在受刺激的呼吸和随自由ADP浓度增加而降低的胞质ATP电位之间产生了一种强制性的关联。
Free cytosolic adenylates were examined in relation to adenosine plus inosine released from perfused working guinea‐pig hearts. Whole‐tissue adenylate data from freeze‐clamped hearts were quantitatively compared with corresponding values obtained by subcellular fractionation of homogenized myocardium in non‐aqueous media. Adenosine and inosine in venous cardiac effuents were measured by high‐performance liquid chromatography. Hearts, perfused at their natural flows, were subjected to various workloads, substrates and catecholamines to alter myocardial energy metabolism and respiration over a wide physiological range. Non‐aqueous cytosolic ATP and creatine phosphate (CrP) accounted for more than 80% of the respective total myocardium content. The cytosolic CrP/Piratio was in near‐quantitative agreement with the overall tissue CrP/Piratio when the latter parameter was corrected for extracellular Pi. This was conclusive evidence that ATP, CrP and Piwere pre‐dominantly located in the cytosol of the well‐oxygenated cardiomyocyte. Measured myocardial oxygen uptake (MVO2) was reciprocally related to the phosphorylation state of CrP ([CrP]/[Cr] × [Pi]) and hence that of ATP ([ATP]/[ADP] × [Pi]) assuming the creatine kinase at near‐equilibrium at a near‐constant pH of 7.2. On the other hand, calculated mean free cytosolic ADP concentrations increased essentially linearily up to threefold with increasing MVO2in the presence of virtually unchanged or only slightly decreased ATP levels; this was found both according to the whole tissue and the special subcellular fractionation data. Employing the myokinase mass‐action ratio and substituting total cardiac ADP by the mean free cytosolic ADP concentrations, the mean free cytosolic AMP concentrations proved to be in the nanomolar range, i.e. up to three orders of magnitude lower than the overall tissue AMP content. We propose, therefore, that in the normoxic heart, AMP is located predominantly in the mitochondrial compartment. Nevertheless, both free cytosolic AMP concentration and release of adenosine plus inosine were apparently square or even higher‐power functions of the rate of cardiac respiration. On the other hand, the mean purine nucleoside release seemed linearily correlated (r= 0.920) with the calculated free cytosolic AMP concentration.Our observations seem to suggest that the concentrations of free ADP and AMP in the cytosol are major determinants of the production of inosine and coronary vasodilator adenosine. Such kinetic control of AMP degradation can conceivably render the free adenylate/adenosine system highly adaptive to mechanical perfor‐mance, total energy output, and/or the energy state of the myocardium. The energy‐linked changes in the free ATP/ADP system also seemed to produce an obligatory association between stimulated respiration and decreased cytosolic ATP potential concomitant with an increase in the concentration of free ADP.