Clinical and Experimental Pharmacology and Physiology (2009) 36, 141–145 doi: 10.1111/j.1440-1681.2008.05034.x Blackwell Publishing Asia Creatine kinase and cardiac function J Ren et al. CREATINE KINASE INHIBITOR IODOACETAMIDE ANTAGONIZES CALCIUM-STIMULATED INOTROPY IN CARDIOMYOCYTES Jun Ren,* § Amy J Davidoff †§ and Joanne S Ingwall ‡ *Center for Cardiovascular Research and Alternative Medicine, University of Wyoming, Laramie, Wyoming, † Department of Pharmacology, University of New England College of Osteopathic Medicine, Biddeford, Maine, ‡ Division of Cardiovascular Medicine, Department of Medicine, Brigham and Women’s Hospital, Harvard Medical School, Boston, Massachusetts and § Program in Cellular and Molecular Cardiology, Department of Internal Medicine, Wayne State University School of Medicine, Detroit, Michigan, USA SUMMARY 1. Inhibition of creatine kinase is known to suppress cardiac contractile reserve in intact hearts, although the underlying mechanism has not been elucidated. 2. The present study was designed to examine whether cardiac depression induced by creatine kinase inhibition was due to action at the level of the essential contractile element, namely cardiomyocytes. Adult rat cardiomyocytes were perfused with the creatine kinase inhibitor iodoacetamide (90 mol/L) for 90 min. Mechanical and intracellular Ca 2+ properties were evaluated using edge-detection and fluorescence microscopy, respectively. Myocytes were superfused with normal (1.3 mmol/L) or high (3.3 mmol/L) extracellular Ca 2+ contractile buffer. Mechanical function was examined, including peak shortening (PS), maximal velocity of shortening/relengthening (±dL/dt), time to 90% PS (TPS 90 ), time to 90% relengthening (TR 90 ) and integration of shortening/relengthening (normalized to PS). Intracellular Ca 2+ transients were evaluated using the following indices: resting and rise of fura-2 fluorescence intensity (FFI) and intracellular Ca 2+ decay time constant. 3. The results indicate that elevated extracellular Ca 2+ stimulated cardiomyocyte positive inotrope, manifested as increased PS, ±dL/dt, area of shortening, resting FFI and FFI associated with a shortened TR 90 and intracellular Ca 2+ decay time constant. High extracellular Ca 2+ did not affect TPS 90 and area of relengthening. Iodoacetamide ablated high Ca 2+ -induced increases in PS, ±dL/dt, area of shortening, resting FFI, FFI and shortened TR 90 and intracellular Ca 2+ decay time constant. Iodoacetamide itself significantly enhanced the area of relengthen- ing and TR 90 without affecting other indices. 4. Collectively, these data demonstrate that inhibition of cre- atine kinase blunts high extracellular Ca 2+ -induced increases in cardiomyocyte contractile response (i.e. cardiac contractile reserve). Key words: cardiomyocytes, contraction, creatine kinase, intracellular Ca 2+ , iodoacetamide. INTRODUCTION The heart requires chemical energy in the form of ATP to maintain its physiological pumping function, which is supplied via the creatine kinase reaction, glycolysis and oxidative phosphorylation. The rate of ATP turnover by creatine kinase is at least one order of magnitude higher than that combined by glycolysis and oxidative phosphorylation. 1 Creatine kinase catalyses the reversible reaction between ADP and phosphocreatine to form ATP and creatine, thus maintaining a high level of chemical energy released from ATP hydrolysis, which is used to support cardiac performance at rest and exercise. 2,3 Despite the critical role of creatine kinase in cardiac energy metabolism, its precise impact on cardiac pumping function has been controversial, especially under pathophysiological conditions. Creatine kinase substrate depletion with phosphocreatine analogues and inhibition of creatine kinase have been demonstrated to suppress creatine kinase reaction kinetics without overtly affecting cardiac performance. 4 This is consistent with the observation that mice lacking the muscle-specific isoenzymes of creatine kinase display impaired cardiac energetics and higher energetic cost for increasing workload is associated with normal cardiac contractile function. 5 Lygate et al. 6 and Zweier et al. 7 have independently shown reduced contractile reserve in hearts with a disturbed creatine kinase system or, vice versa, alterations in the creatine kinase system during heart failure. Data from our group also reveal that inhibition of the creatine kinase reaction reduces cardiac contractile reserve (i.e. the potential to sustain a higher workload upon inotropic stimulation or exercise in intact hearts). 8,9 Reduced cardiac reserve (or stress intolerance) is one of the most prominent defects in the elderly and in patients with congestive heart failure. 10,11 Diminished cardiac reserve is commonly shown as compromised systolic function, prolonged contraction duration and reduced wall compliance, which makes it difficult to support increased cardiac workload under exercise. 12 The correlation between creatine kinase reaction and cardiac function also received compelling support from the find- ing that reduced creatine kinase reaction is often associated with Correspondence: Dr Jun Ren, Center for Cardiovascular Research and Alternative Medicine, University of Wyoming, Laramie, WY 82071, USA. Email: jren@uwyo.edu Received 7 April 2008; revision 18 June 2008; accepted 23 June 2008. © 2008 The Authors Journal compilation © 2008 Blackwell Publishing Asia Pty Ltd