Synchronous in situ ATPase activity, mechanics, and Ca2+ sensitivity of human and porcine myocardium.
Flash-frozen myocardium samples provide a valuable means of correlating clinical cardiomyopathies with abnormalities in sarcomeric contractile and biochemical parameters. We examined flash-frozen left-ventricle human cardiomyocyte bundles from healthy donors to determine control parameters for isometric tension (P(o)) development and Ca(2+) sensitivity, while simultaneously measuring actomyosin ATPase activity in situ by a fluorimetric technique. P(o) was 17 kN m(-2) and pCa(50%) was 5.99 (28 degrees C, I = 130 mM). ATPase activity increased linearly with tension to 132 muM s(-1). To determine the influence of flash-freezing, we compared the same parameters in both glycerinated and flash-frozen porcine left-ventricle trabeculae. P(o) in glycerinated porcine myocardium was 25 kN m(-2), and maximum ATPase activity was 183 microM s(-1). In flash-frozen porcine myocardium, P(o) was 16 kN m(-2) and maximum ATPase activity was 207 microM s(-1). pCa(50%) was 5.77 in the glycerinated and 5.83 in the flash-frozen sample. Both passive and active stiffness of flash-frozen porcine myocardium were lower than for glycerinated tissue and similar to the human samples. Although lower stiffness and isometric tension development may indicate flash-freezing impairment of axial force transmission, we cannot exclude variability between samples as the cause. ATPase activity and pCa(50%) were unaffected by flash-freezing. The lower ATPase activity measured in human tissue suggests a slower actomyosin turnover by the contractile proteins.
Griffiths
P J
PJ
Department of Physiology, Anatomy and Genetics, University of Oxford, Oxford, United Kingdom. pjg@dpag.ox.ac.uk
Isackson
H
H
Pelc
R
R
Redwood
C S
CS
Funari
S S
SS
Watkins
H
H
Ashley
C C
CC
D000205:Actomyosin / Q000737:chemistry; D000251:Adenosine Triphosphatases / Q000737:chemistry*; D000255:Adenosine Triphosphate / Q000737:chemistry; D000818:Animals; D001703:Biophysics / Q000379:methods; D002118:Calcium / Q000737:chemistry* / Q000378:metabolism; D005470:Fluorometry / Q000379:methods; D005990:Glycerol / Q000737:chemistry; D006801:Humans; D006863:Hydrogen-Ion Concentration; D018625:Microscopy, Atomic Force / Q000379:methods; D009200:Myocardial Contraction; D009206:Myocardium / Q000378:metabolism / Q000473:pathology*; D009218:Myosins / Q000737:chemistry; D013552:Swine
D000255:Adenosine Triphosphate; D000205:Actomyosin; D000251:Adenosine Triphosphatases; D009218:Myosins; D005990:Glycerol; D002118:Calcium