Diastolic dysfunction in hypertrophic cardiomyopathy. Effect on active force generation during systole.

We tested the hypothesis that intracellular Ca++ [( Ca++]i) overload underlies the diastolic dysfunction of patients with hypertrophic cardiomyopathy. Myocardial tissue was obtained at the time of surgery or transplantation from patients with hypertrophic cardiomyopathy and was compared with control myocardium obtained from patients without heart disease. The isometric contractions and electrophysiologic properties of all myocardial specimens were recorded by standard techniques and [Ca++]i was measured with the bioluminescent calcium indicator aequorin. In contrast to the controls, action potentials, Ca++ transients, and isometric contraction and relaxation were markedly prolonged in the hypertrophic myocardium, and the Ca++ transients consisted of two distinct components. At 38 degrees C and 1 Hz pacing frequency, a state of relative Ca++ overload appeared develop, which produced a rise in end-diastolic [Ca++]i, incomplete relaxation, and fusion of twitches with a resultant decrease in active tension development. We also found that drugs with increase [Ca++]i, such as digitalis, exacerbated these abnormalities, whereas drugs that lower [Ca++]i, such as verapamil, or agents that increase cyclic AMP, such as forskolin, prevented them. These results may explain why patients with hypertrophic cardiomyopathy tolerate tachycardia poorly, and may have important implications with regard to the pharmacologic treatment of patients with hypertrophic cardiomyopathy.

Abnormal intracellular calcium handling in myocardium from patients with end-stage heart failure.

Intracellular Ca2+ release and reuptake are essential for contraction and relaxation of normal heart muscle. Intracellular Ca2+ transients were recorded with aequorin during isometric contraction of myocardium from patients with end-stage heart failure. In contrast to controls, contractions and Ca2+ transients of muscles from failing hearts were markedly prolonged, and the Ca2+ transients exhibited 2 distinct components. Muscles from failing hearts showed a diminished capacity to restore low resting Ca2+ levels during diastole. These experiments provide the first direct evidence from actively contracting human myocardium that intracellular Ca2+ handling is abnormal and may cause systolic and diastolic dysfunction in heart failure.

A method for recording isometric tension development by isolated cardiac myocytes: transducer attachment with fibrin glue.

The purpose of this study was to develop a method for attachment of single isolated cardiac myocytes to a transducer for recording isometric tension development. Cardiac myocytes were isolated from the hearts of the toad, Bufo marinus or ferrets by enzymatic digestion with collagenase. The method that we used provided a 60-80% yield of Ca++-tolerant cells. A suspension of cells was placed into a superfusion chamber coated with bovine thrombin. Two glass microtools - each attached to a micromanipulator - were brought into proximity with the ends of a single myocyte; one of the microtools was attached to the element of a low-level force transducer. Human fibrinogen was loaded into a fine-tipped glass micropipette mounted on a micromanipulator. Small amounts of fibrinogen were pressure-ejected from the pipette at each junction between the microtool and the end of the myocyte. The fibrin that formed produced a stable attachment of the ends of the myocyte to the microtools. The myocyte could subsequently be stretched and a length-tension curve recorded. We have used this method to record concentration-dependent tension development in response to the Ca++-ionophore, A23187, and potassium depolarization. Our results indicate that fibrin glue may facilitate the study of the mechanical properties of isolated myocytes.

Deficient production of cyclic AMP: pharmacologic evidence of an important cause of contractile dysfunction in patients with end-stage heart failure.

We studied the effects of different classes of inotropic drugs on human working myocardium in vitro that was isolated from the hearts of patients with end-stage heart failure, and compared the responses to these drugs with those noted in muscles from nonfailing control hearts. Although peak isometric force generated in response to increased extracellular calcium reached control levels in the muscles from patients with heart failure, the time course of contraction and rate of relaxation were greatly prolonged. The inotropic effectiveness of the beta-adrenergic agonist isoproterenol and the phosphodiesterase inhibitors milrinone, caffeine, and isobutylmethylxanthine was markedly reduced in muscles from the patients with heart failure. In contrast, the effectiveness of inotropic stimulation with acetylstrophanthidin and the adenylate cyclase activator forskolin was preserved. After a minimally effective dose of forskolin was given to elevate intracellular cyclic AMP levels, the inotropic responses of muscles from the failing hearts to phosphodiesterase inhibitors were markedly potentiated. These data indicate that an abnormality in cyclic AMP production may be a fundamental defect present in patients with end-stage heart failure that can markedly diminish the effectiveness of agents that depend on generation of this nucleotide for production of a positive inotropic effect.