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Title: Relaxation processes in cardiac muscle
Author: Simnett, Sarah Jacqueline
ISNI:       0000 0001 3412 1706
Awarding Body: University of Oxford
Current Institution: University of Oxford
Date of Award: 1993
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The technique of laser flash photolysis was used in conjunction with diazo-2 to investigate the relaxation kinetics of skinned trabeculae from the guinea-pig. Diazo-2 is a caged Ca2+ chelator, based on BAPTA, which undergoes a rapid photochemical conversion (> 2000s-1), when exposed to ultraviolet light, from a Ca2+ chelator with low affinity (Kd=2.2μM) to one with a higher affinity (Kd=0.073μM). The resulting rapid Ca2+ uptake produces a rapid decline in the free Ca2+ concentration (within 2-3msec). Subsequent computer simulations have shown that this rate of Ca 2+ decline is independent of the Ca2+ off rate from the Ca2+ specific binding site on troponin C. The use of diazo-2 overcomes problems of diffusional delays of Ca2+ equilibration which were inherent in previous studies of muscle kinetics. The half time of the relaxation of skinned trabeculae following the photolysis of diazo-2 was 72.3 ± 7.5msec (mean ± SD) at 12°C, and decreased to 53.4±5.2msec at 20°C, both several times faster than the relaxation rate of electrically stimulated trabeculae at the same temperature. The relaxation transients were closely fitted to two exponentials, suggesting that relaxation is governed by two rate limiting processes. Varying the degree of relaxation from 100% relaxation to 30% relaxation had no effect on the half time of the decline in force, indicating that, unlike activation, the processes of relaxation are not cooperative. The effects of the metabolites H+, ADP and Pi (at concentrations similar to those reached during cardiac ischaemia and hypoxia) on the rate of relaxation were investigated. Pi increased the relaxation rate, in contrast with its effect in skeletal muscle, where a slowing of relaxation was seen; a discrepancy suggested to be due to differences in the muscle types. Both H+ and ADP decreased the relaxation rate; however, their effects on the relaxation transients were distinct, indicating that they altered the kinetics of relaxation by different mechanisms. EMD 57033, a thiadiazinone derivative, is a novel cardiotonic agent which acts by increasing the apparent Ca2+ sensitivity of the myofibrils. Maximal Ca2+ activated force (Pmax) is increased by 10 μM EMD to 138±4.23% of control. EMD 57033 produces a slight increase in the rate of relaxation initiated by the photolysis of diazo-2, unlike the endogenous potentiator ADP. In experiments using caged Ca2+ (nitr-5), EMD 57033 markedly increases the rate of activation. This suggests that the Ca2+ sensitising effect of EMD 57033 is, at least partially, mediated by an increase in the rate at which crossbridges reach the force-generating state. In contrast, although ADP also increases Pmax it decreases the rate of transition out of the force-generating state.
Supervisor: Ashley, Christopher C. Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available
Keywords: Myocardium ; Heart ; Metabolism ; Calcium ions ; Physiological effect ; Contractility (Biology)