title: Intrinsic myofilament alterations underlying the decreased contractility of stunned myocardium. A consequence of Ca2+-dependent proteolysis?
abstract: We investigated the mechanism of the decreased myofilament Ca2+ responsiveness in stunned myocardium. The steady state force-[Ca2+] relationship was measured before and after skinning in thin ventricular trabeculae from control or stunned (20 minutes of ischemia, 20 minutes of reperfusion) rat hearts.[Ca2+]i was determined using microinjected fura 2 salt in intact muscles, whereas the myofilaments of chemically skinned trabeculae were activated directly with solutions of varied [Ca2+]. Maximal Ca2+- activated force (F max) before and after skinning was identical within either the control or stunned groups but was markedly depressed in both groups of stunned trabeculae (P < .001)). After ischemia and reperfusion, the [Ca2+] required for 50% of maximal activation (Ca50) was increased in both intact (control, 0.60 +/- 0.09 micromol/L; stunned, 0.85 +/- 0.09 micromol/L;P < .001) and skinned (control, 1.13 +/- 0.24 micromol/L; stunned 1.39 +/- 0.21 micromol/L; P = .0025) trabeculae. These data indicate that the decreased Ca2+ responsiveness of stunned myocardium is due to intrinsic alterations of the myofilaments. Therefore, we tested the hypothesis that activation of proteases by reperfusion-induced Ca2+ overload decreases the Ca2+ responsiveness of the cardiac myofilaments. Force-[Ca2+] relations were compared before and 5 to 30 minutes after direct exposure of skinned trabeculae to calpain I (18 microgram/mL, 20 minutes at [Ca2+]=10.8 micromol/L), a Ca2+-activated protease that is present in myocardium. Calpain I reduced F max from 94.3 +/- 8.3 to 56 +/- 8.5 mN/mm2 while increasing Ca50 from 0.94 +/- 0.11 to 1.36 +/- 0.21 micromol/L (P < .01). Calpastatin, a specific calpain inhibitor prevented the effects of calpain I on skinned trabeculae. The results show that the reduced Ca2+ responsiveness of stunned myocardium reflects alteration of the myofilaments themselves, not of soluble cytosolic factors, which can be faithfully reproduced by exposure to Ca2+-dependent protease.
authors.affiliation: Department of Medicine, Johns Hopkins University School of Medicine, Baltimore, MD, USA.
authors.forename: W D
authors.initials: WD
authors.lastname: Gao
authors.forename: Y
authors.initials: Y
authors.lastname: Liu
authors.forename: R
authors.initials: R
authors.lastname: Mellgren
authors.forename: E
authors.initials: E
authors.lastname: Marban
mesh_terms: D008841:Actin Cytoskeleton / Q000378:metabolism*; D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002154:Calpain / Q000494:pharmacology; D010450:Endopeptidases / Q000378:metabolism; D005260:Female; D006868:Hydrolysis; D066298:In Vitro Techniques; D007700:Kinetics; D008297:Male; D009200:Myocardial Contraction / Q000187:drug effects; D017202:Myocardial Ischemia / Q000378:metabolism* / Q000503:physiopathology; D015428:Myocardial Reperfusion Injury / Q000378:metabolism / Q000503:physiopathology; D051381:Rats; D017207:Rats, Sprague-Dawley
chemical_list: D010450:Endopeptidases; D002154:Calpain; D002118:Calcium