The calcium sensitivity of force production of cardiac muscle fibres is altered by certain drugs. The sites of action of three such compounds (pimobendan, sulmazole, isomazole) within the myofibril have been investigated. Calmodulin antagonists, perhexilene and bepridil, which have been shown to alter the calcium dependence of myofibrillar ATPase activity and oxmetidine, an H2-receptor antagonist which binds to calmodulin, were also studied. 2. The rates of dissociation of calcium from both the regulatory and high affinity sites on bovine isolated cardiac troponin C (cTnC) were measured in a stopped-flow fluorimeter. The rates of dissociation were found to be 136.5 +/- 16 s-1 and 1.3 +/- 0.20 s-1 (mean +/- s.e.mean, n = 11 determinations; conditions: 100 mM KCl, 10 mM MOPS, 3 mM MgCl2, 0.1 mM dithriothreitol, pH 7.0, 15 degrees C). Sulmazole, isomazole and perhexiline (final concentration of 50 microM) had no effect on the rate of Ca2+ dissociation from the regulatory Ca2+ site, indicating that these compounds do not act on cTnC directly. 3. The rate of dissociation of Ca2+ from the regulatory site was slightly reduced (approximately 20%) by pimobendan (50 and 100 microM) and was somewhat increased by oxmetidine (28% at 100 microM). 4. Bepridil (25 microM) reduced the rate of dissociation by 50%, indicating a direct effect of bepridil on TnC. 5. Sulmazole, isomazole, perhexiline, pimobendan (50 microM) and bepridil (25 microM) were without effect on the rate of dissociation of Ca2+ from the high affinity Ca2+/Mg2+ sites. Oxmetidine caused 24% decrease in the rate of Ca2+ dissociation from these sites. 6. The rate of dissociation of Ca2+ from the regulatory site on the complex of troponin-tropomyosin (TnTm) was measured. Sulmazole and pimobendan (50 microM) were without effect on the rate of dissociation of Ca2+ from the regulatory site in the protein complex, and isomazole (50 microM) caused only a slight reduction (23%). Perhexiline (50 microM) or bepridil (10 microM) reduced the rate of Ca2 dissociation by about 50%. The rate of dissociation of Ca2+ from the high affinity Ca2 +/Mg2 + sites was not altered by sulmazole, isomazole, or pimobendan (50 microM), but was decreased - 35% by perhexiline (50 microM) or bepridil (10 microM).
摘要
某些药物会改变心肌纤维产生力量的钙敏感性。已对三种此类化合物（匹莫苯丹、舒马唑、异马唑）在肌原纤维内的作用位点进行了研究。还研究了钙调蛋白拮抗剂哌克昔林和苄普地尔，它们已被证明可改变肌原纤维ATP酶活性的钙依赖性，以及H2受体拮抗剂奥美替丁，它可与钙调蛋白结合。2. 在停流荧光计中测量了钙从牛离体心肌肌钙蛋白C（cTnC）的调节位点和高亲和力位点解离的速率。发现解离速率分别为136.5±16 s-1和1.3±0.20 s-1（平均值±标准误平均值，n = 11次测定；条件：100 mM KCl、10 mM MOPS、3 mM MgCl2、0.1 mM二硫苏糖醇、pH 7.0、15℃）。舒马唑、异马唑和哌克昔林（终浓度50μM）对Ca2+从调节性Ca2+位点的解离速率没有影响，表明这些化合物不直接作用于cTnC。3. 匹莫苯丹（50和100μM）使Ca2+从调节位点的解离速率略有降低（约20%），而奥美替丁（100μM时为28%）使其有所增加。4. 苄普地尔（25μM）使解离速率降低了50%，表明苄普地尔对TnC有直接作用。5. 舒马唑、异马唑、哌克昔林、匹莫苯丹（50μM）和苄普地尔（25μM）对Ca2+从高亲和力Ca2+/Mg2+位点的解离速率没有影响。奥美替丁使Ca2+从这些位点的解离速率降低了24%。6. 测量了钙从肌钙蛋白-原肌球蛋白（TnTm）复合物调节位点的解离速率。舒马唑和匹莫苯丹（50μM）对蛋白质复合物中调节位点的Ca2+解离速率没有影响，而异马唑（50μM）仅使其略有降低（23%）。哌克昔林（50μM）或苄普地尔（10μM）使Ca2+解离速率降低了约50%。舒马唑、异马唑或匹莫苯丹（50μM）对Ca2+从高亲和力Ca2 +/Mg2 +位点的解离速率没有改变，但哌克昔林（50μM）或苄普地尔（10μM）使其降低了35%。
