ALTERATION OF THE FRANK-STARLING LAW IN HEART FAILURE RELATES TO TITIN Olivier Cazorla, Alain Lacampagne, Harvard Viko, Claire Bony, CCline Cantos, J-Yves Leguennec & Guy Vassort. INSERM U-390, IFR3, CHU AdV, Montpellier, FRANCE

Alterations of the Frank-Starling relationship under pathological conditions is still a matter of controversy. We investigated this property in perfused whole hearts and in single cells isolated from normal and post-myocardium infarcted (PMI) rats. A balloon connected to a pressure sensor was implanted in the left ventricle, and the intraventricular pressure was monitored following changes in volume. A first group of rats with a very low level of remodeling had a preserved Frank-Starling relationship. The second group, in heart failure, was characterized by tremendous fibrosis and ventricle dilatation, and presented an alteration of the Frank-Starling relationship. ‘Ihe passive and contractile properties of single chemically skinned ventricular cells were measured as well as their length-dependent Ca2’ sensitivity. Cells from failing hearts exhibited a lower stiffness and abnormal tension-pCa properties characterized by a decrease (from 0.4 pCa Unit in control to below 0.2) in the shift of pCa50 induced by a stretch from 1.9 pm to 2.3 nm sarcomere length. Biochemical analysis of contractile proteins was performed. Polyacrylamide gel electrophoresis (2.5-12%) revealed an altered form of titin in the pathological conditions. Based on a previous report showing that the degradation of titin by a mild trypsin digestion resulted in an alteration of the contractility of normal cells (Cazorla et al. 1999), we conclude that the inability of failing hearts to use the Frank-Starling relationship is, in part, attributable to titin alteration.

A COMPARISON BETWEEN NMR AND GCMS %- ISOTOPOMER ANALYSIS IN CARDIAC METABOLISM. John Chatham, Bertrand Bouchard (L Christlne Des Roslers. NMR Res 8 Dev, U. Alabama at Birmingham; Dept. Nutrition & CHUM Res. Ctr, Univ. Montreal.

Carbon-13 labeled substrates combined with isotopomer analysis by NMR spectroscopy or gas chromatography-mass spectrometry (GCMS) has been used to study cardiac metabolism. It is not known if the data generated by these methods agree or if the combination of methods may yield insights into metabolism that cannot be obtained by either method alone. Therefore, NMR and GCMS isotopomer analyses were performed on extracts from hearts perfused with physiological concentrations of 13C-labe&d lactate, pyruvate and palmitate, and unlabeled glucose. The absolute mass isotope distribution (MID) of 2-ketoglutarate (2KG) measured directly by GCMS was compared with that calculated using ‘3C-NMR glutamate isotopomer analysis. There was a linear correlation between the two techniques (P=O.817, pcO.001); although the slope was less than one (0.79rtO.04). A the contribution from unlabeled 2KG was removed (i.e. relative MID) the correlation coefficient improved (&0.975) and the slope increased to 0.90~0.02. Thus, both NMR and GCMS provide comparable ‘c isotopomer information: however, compared to GCMS, NMR underestimates the unlabeled fraction of 2KG by 30-40%. This may reflect the fact that in contrast to GCMS, NMR does not measure the unlabeled component directly but rather calculates it by difference. Alternatively, these results may point to a potential compartmentation of citric acid cycle intermediates, which would not have been apparent with either technique alone.

FACTORS MODULATING SARCOPLASMIC RETICULUM GENE EXPRESSION IN THE ISOLATED RAT HEART Donald Chapman, Rana M. Temsah & Naranjan S. Dhalla. Institute of Cardiovascular Sciences, University of Manitoba, Faculty of Medicine, Winnipeg, Manitoba, Canada.

In view of the deleterious effects of ischemia-reperfusion (IR) on the sarcoplasmic reticulum (SR) function, we investigated the effects of IR on SR gene expression in the isolated rat heart. Globally ischemic hearts showed down-regulation in mRNA levels for the SR Ca’+-pump, Cazt-release channels and phospholatnban proteins; reperfusion further reduced the abundance of these genes. Treatment with a mixture of superoxide dismutase plus catalase prevented these IR-induced alterations. Hearts treated with xanthine plus xanthine oxidase or HzOz induced changes similar to those observed in the IR hearts. Moreover, Ca*’ overloaded hearts (achieved by Ca*‘- depletion/repletion) induced drastic decrease in the SR mRNA levels. In view of the occurrence of hypoxia and glucose deprivation in the ischemic heart, we found that neither of these factors above had any effect on the SR genes whereas hypoxia plus glucose deprivation depressed SR tnRNA levels in the reperfused hearts. These data suggest that: 1) IR has a deleterious effect on SR gene expression, 2) oxidative stress and Ca”-overload modulate SR gene expression, and 3) hypoxia plus glucose deprivation may be possible ischetnic factors that affect SR genes in the IR heart. (Supported by CIHR Group in Experimental Cardiology)

CARDlAC CARBOHYDRATE METABOLISM IN HEARTS FROM NORMAL AND TYPE-2 DIABETlC RATS John Chatham & AnneMarie Seymour. Center for NMR Res. 8 Dev., Univ Alabama at Birmingham; Dept. Biological Sciences, University of Hull, England.

Studies on the regulation of cardiac metabolism in diabetes have focused on the interaction between glucose and fatty acids in hearts from Type 1 diabetic animals. However, -90% of diabetic population have Type 2 rather than Type 1 diabetes. Furthermore, in vivo, lactate and pyruvate can also be used for energy production. Therefore, we determined a) the relative contributions of glucose, lactate and pyruvate to oxidative energy production and b) the impact of Type-2 diabetes on carbohydrate utilization. Hearts from control rats were perfused with Krebs-Henseleit buffer containing 5mM glucose, 1mM lactate, O.lmM pyruvate, 0.4mM palmitate, 5OuWml insulin. Cardiac function as rate pressure product was similar in control and diabetic groups. Lactate contributed 6423% of total pyruvate oxidation; pyruvate and glucose provided 24*3% and 1 l*l% respectively. In hearts from Type 2 diabetic rats the contribution of glucose to pyruvate oxidation was decreased by -35% (~~0.03). The contribution of glucose to tissue lactate and alanine formation was also depressed. Flux through PDH provided 9il% TCA cycle flux in controls and was decreased by -30% in the diabetic group (&l%; p=O.OsS). In the diabetic group the fraction of actiie PDH was decreased compared to controls (0.26*0.05 vs. 0.51*0.06; pcO.025). These results demonstrate that under normal conditions glucose contributes -10% of total carbohydrate oxidation. Type 2 diabetes leads to a decrease in the provision of glucose to tissue pyruvate and a decrease in total pyruvate oxidation,

Al9