Palmer, Susan; (1992) The role of troponin C in the modulation of myofibrillar Ca(2+)-sensitivity. Doctoral thesis (Ph.D), UCL (University College London).

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Abstract

It is desirable to treat the failing heart by increasing contractile force without 9 X raising level of intracellular Ca2+. This could be achieved by increasing the sensitivity of myofilaments to Ca2+ but "Ca2+-sensitisers" have unwanted side-effects. In addition, the mechanism of the decrease in Ca2+-sensitivity by low pH, or elevated Pi is unclear. I investigated the role of troponin-C (TnC) in these changes in Ca2+-sensitivity by (i) comparing effects on cardiac and skeletal myofibrils (which have different forms of TnC) and (ii) measuring the Ca2+ affinity of TnC, both isolated and in the myofibrils. The effects of pH, Pi and caffeine on skinned fibres were measured. Isolated skeletal and cardiac TnCs labelled with DANZ and lAANS were used to detect any direct effects of pH, Pi and caffeine on Ca2+-binding to TnC. Neither Pi nor caffeine altered the Ca2+-affinity of the TnC, but acidic pH decreased TnC Ca2+-binding. Thus the negative inotropic effects of low pH in striated muscle are partially explained by a reduction in Ca2+-affinity of the TnC. Pi and caffeine-induced modulation of Ca2+-sensitivity may depend on other proteins, or may directly affect crossbridges. I measured the Ca2+-sensitising effects of novel compounds developed by Ciba-Geigy. The effects were species dependent and most pronounced in porcine muscle. CGP48506, like caffeine, had a greater Ca2+-sensitising action in cardiac than skeletal fibres whereas CGP48508 unexpectedly worked the other way around. Neither compound affected Ca2+-binding to isolated skeletal TnC so it appeared that their effects on skeletal muscle were not solely dependent on the moiety of TnC present. To investigate the effects of CGP48506 on Ca2+-binding to TnC in skinned psoas preparations, native TnC was exchanged for DANZ-labelled skeletal TnC. Results indicated that CGP48506 increased the Ca2+-affinity of TnC. As binding of Ca2+ to isolated TnC was unaffected it appears that sensitisation depended on factor(s) in addition to TnC.

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