Korge P, Campbell K B
Department of Veterinary and Comparative Anatomy, Pharmacology, and Physiology, Washington State University, Pullman 99164-6520.
Am J Physiol. 1994 Aug;267(2 Pt 1):C357-66. doi: 10.1152/ajpcell.1994.267.2.C357.
Ca2+ pump function of skeletal muscle sarcoplasmic reticulum (SR) vesicles was measured by monitoring Ca2+ uptake and efflux with a Ca(2+)-sensitive minielectrode and adenosinetriphosphatase (ATPase) activity of the same preparation under the same conditions. The efficiency of Ca2+ transport into SR vesicles, defined by the amount of Ca2+ transported per ATP hydrolyzed (coupling ratio), varied significantly depending on assay conditions. Coupling ratio increased in parallel with increase in precipitating anion concentration, which is supposed to decrease accumulation of free Ca2+ inside vesicles and its subsequent efflux. Membrane-bound creatine kinase-creatine phosphate (CK-CP) system, acting as a ADP sensor and local ATP regenerator, significantly improved Ca2+ pump function when the pump worked with low efficiency (coupling ratio < 1). The effect of CK-CP system on Ca2+ pump function was also dependent on extravesicular Ca2+ concentration ([Ca2+]o), the effect being most significant at high initial [Ca2+]o. Under conditions in which SR vesicles were allowed to decrease [Ca2+]o, as occurs also during muscle relaxation, plateau values of Ca(2+)-ATPase activity were reached at significantly higher [Ca2+]o (54 +/- 5.7, n = 6), compared with leaky vesicles or the condition in which [Ca2+]o was maintained. By preventing local accumulation of ADP, generated in ATPase reactions, CK-CP system also inhibited Ca2+ efflux under conditions in which this efflux was stimulated by the increase of free Ca2+ inside vesicles. This effect was at least partially responsible for the CK-CP-supported increase in Ca2+ uptake and coupling ratios that were more expressed at low precipitating anion concentration. We hypothesize that local ATP regeneration by CK-CP system is one mechanism the cell can use to improve Ca2+ uptake by SR in emergency conditions, where excessive increase in cytoplasmic [Ca2+] may have deleterious effects.
通过使用钙敏微电极监测钙摄取和流出，并在相同条件下测量同一制剂的腺苷三磷酸酶（ATP酶）活性，来测定骨骼肌肌浆网（SR）囊泡的Ca2+泵功能。Ca2+转运到SR囊泡中的效率，由每水解一分子ATP所转运的Ca2+量（偶联比）定义，其会因测定条件的不同而有显著变化。偶联比随着沉淀阴离子浓度的增加而平行增加，这被认为会减少囊泡内游离Ca2+的积累及其随后的流出。膜结合肌酸激酶-磷酸肌酸（CK-CP）系统，作为一种ADP传感器和局部ATP再生器，当泵效率较低（偶联比<1）时，能显著改善Ca2+泵功能。CK-CP系统对Ca2+泵功能的影响也取决于囊泡外Ca2+浓度（[Ca2+]o），在高初始[Ca2+]o时这种影响最为显著。在允许SR囊泡降低[Ca2+]o的条件下，就像肌肉松弛时也会发生的那样，与渗漏囊泡或维持[Ca2+]o的条件相比，Ca(2+)-ATP酶活性的稳定值在显著更高的[Ca2+]o（54±5.7，n = 6）时达到。通过防止ATP酶反应中产生的ADP局部积累，CK-CP系统在囊泡内游离Ca2+增加刺激这种流出的条件下，也抑制了Ca2+流出。这种效应至少部分解释了在低沉淀阴离子浓度下更明显的CK-CP支持的Ca2+摄取增加和偶联比增加。我们推测，在紧急情况下，当细胞质[Ca2+]过度增加可能产生有害影响时，CK-CP系统进行的局部ATP再生是细胞可用来改善SR对Ca2+摄取的一种机制。
