Relation between steady-state force and intracellular [Ca2+] in intact human myocardium. Index of myofibrillar responsiveness to Ca2+.
A novel approach was developed allowing the measurement of steady-state force and intracellular calcium concentration ([Ca2+]i) in tetanized human ventricular trabeculae carneae without pharmacological intervention. We compared and contrasted three methods of assessing calcium sensitivity of the myofilaments: 1) force-[Ca2+] relations in skinned fiber preparations, 2) peak twitch force-peak [Ca2+]i relations, and 3) steady-state force-[Ca2+]i relations in intact muscles. Steady-state contractile activation was achieved rapidly by tetanizing intact human trabeculae, loaded with aequorin, a Ca2(+)-sensitive bioluminescent protein, at a stimulation frequency of 15-20 Hz. Steady-state force and [Ca2+]i were measured during tetani, and the force versus [Ca2+]i relation was obtained by varying the extracellular calcium concentration ([Ca2+]o). Force-[Ca2+]i relations obtained from control and myopathic hearts were fitted to the Hill equation: %Force = [Ca2+]inh/([Ca2+]inh50% + [Ca2+]inh), where nh is the Hill coefficient, and [Ca2+]50% is the [Ca2+] required for 50% activation. The curves of tetani had Hill coefficients of 5.21 +/- 0.20 (n = 6) and 5.61 +/- 0.60 (n = 10) and [Ca2+]50% of 0.56 +/- 0.05 microM (n = 6) and 0.54 +/- 0.09 microM (n = 10) in control and myopathic muscles, respectively. We also constructed peak force-peak [Ca2+]i relations using isometric twitches from the same muscles. These curves were shifted toward higher [Ca2+]i compared with the steady-state force-[Ca2+]i curve derived from tetani. Ryanodine (1 microM), which increased the time course of the Ca2+ and force transients, shifted the peak force-peak [Ca2+]i relation to the left, without affecting the steady-state force-[Ca2+]i relation. Exposure to 10 mM caffeine shifted the steady-state force-[Ca2+]i relation to the left, whereas exposure to 3 microM isoproterenol shifted this relation to the right. Experiments using skinned fiber preparations were performed in parallel with experiments on intact muscles from the same hearts. The force-pCa (-log[Ca2+]) relations in saponin-skinned trabeculae from control and myopathic tissue were superimposable. Ryanodine (1 microM) had no effect on the force-pCa relation in skinned fibers. Maximal tension was evoked by the posttetanic twitch, which was larger than the tetanus. This potentiation was abolished in the presence of ryanodine, a sarcoplasmic reticulum inhibitor. We propose that the changes in the steady-state force-[Ca2+]i relations are correlated with alterations in the sensitivity of the myofilaments to Ca2+, whereas changes in the peak force-peak [Ca2+]i relations represent temporal changes in the twitch transient.(ABSTRACT TRUNCATED AT 400 WORDS)
Gwathmey
J K
JK
Department of Cellular and Molecular Physiology, Harvard Medical School, Boston, MA.
Hajjar
R J
RJ
D002110:Caffeine / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism* / Q000502:physiology; D006652:Histological Techniques; D006706:Homeostasis; D006801:Humans; D007425:Intracellular Membranes / Q000378:metabolism*; D007477:Ions; D007545:Isoproterenol / Q000494:pharmacology; D009200:Myocardial Contraction / Q000187:drug effects; D009206:Myocardium / Q000378:metabolism*; D009210:Myofibrils / Q000502:physiology*; D009994:Osmolar Concentration; D010879:Piperazines / Q000494:pharmacology; D012433:Ryanodine / Q000494:pharmacology
D007477:Ions; D010879:Piperazines; D012433:Ryanodine; D002110:Caffeine; C046261:DPI 201-106; D007545:Isoproterenol; D002118:Calcium