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Prolonged mechanical unloading (UN) of the heart is associated with detrimental changes to the structure and function of cardiomyocytes. The mechanisms underlying these changes are unknown. In this study, we report the influence of UN on excitationcontraction coupling, Ca 2؉ -induced Ca 2؉ release (CICR) in particular, and transverse (t)-tubule structure. UN was induced in male Lewis rat hearts by heterotopic abdominal heart transplantation. Left ventricular cardiomyocytes were isolated from the transplanted hearts after 4 wk and studied using whole-cell patch clamping, confocal microscopy, and scanning ion conductance microscopy (SICM). Recipient hearts were used as control (C). UN reduced the volume of cardiomyocytes by 56.5% compared with C (UN, n‫;09؍‬ C, n‫;95؍‬ P<0.001). The variance of time-to-peak of the Ca 2؉ transients was significantly increased in unloaded cardiomyocytes (UN 227.4؎24.9 ms 2 , n‫24؍‬ vs.

AimsCombined left ventricular assist device (LVAD) and pharmacological therapy has been proposed to favour myocardial recovery in patients with end-stage heart failure (HF). Clenbuterol (Clen), a β2-adrenoceptor (β2-AR) agonist, has been used as a part of this strategy. In this study, we investigated the direct effects of clenbuterol on unloaded myocardium in HF.Methods and resultsLeft coronary artery ligation or sham operation was performed in male Lewis rats. After 4–6 weeks, heterotopic abdominal transplantation of the failing hearts into normal recipients was performed to induce LV unloading (UN). Recipient rats were treated with saline (Sal) or clenbuterol (2 mg/kg/day) via osmotic minipumps (HF + UN + Sal or HF + UN + Clen) for 7 days. Non-transplanted HF animals were treated with Sal (Sham + Sal, HF + Sal) or clenbuterol (HF + Clen). LV myocytes were isolated and studied using optical, fluorescence, and electrophysiological techniques. Clenbuterol treatment improved in vivo LV function measured with echocardiography (LVEF (%): HF 35.9 ± 2 [16], HF + Clen 52.1 ± 1.4 [16]; P < 0.001; mean ± SEM [n]). In combination with unloading, clenbuterol increased sarcomere shortening (amplitude (µm): HF + UN + Clen 0.1 ± 0.01 [50], HF + UN + Sal 0.07 ± 0.01 [38]; P < 0.001) by normalizing the depressed myofilament sensitivity to Ca2+ (slope of the linear relationship between Ca2+ transient and sarcomere shortening hysteresis loop during relaxation (μm/ratio unit): HF + UN + Clen 2.13 ± 0.2 [52], HF + UN + Sal 1.42 ± 0.13 [38]; P < 0.05).ConclusionClenbuterol treatment of failing rat hearts, alone or in combination with mechanical unloading, improves LV function at the whole-heart and cellular levels by affecting cell morphology, excitation–contraction coupling, and myofilament sensitivity to calcium. This study supports the use of this drug in the strategy to enhance recovery in HF patients treated with LVADs and also begins to elucidate some of the possible cellular mechanisms responsible for the improvement in LV function.

. Effects of chronic administration of clenbuterol on function and metabolism of adult rat cardiac muscle. Am J Physiol Heart Circ Physiol 288: H1468 -H1476, 2005. First published November 4, 2004 doi: 10.1152/ajpheart.00624.2004, a ␤2-agonist, is known to produce skeletal and myocardial hypertrophy. This compound has recently been used in combination with left ventricular assist devices for the treatment of end-stage heart failure to reverse or prevent the adverse effects of unloading-induced myocardial atrophy. However, the mechanisms of action of Clen on myocardial cells have not been fully elucidated. In an attempt to clarify this issue, we examined the effects of chronic administration of Clen on Ca 2ϩ handling and substrate preference in cardiac muscle. Rats were treated with either 2 mg ⅐ kg Ϫ1 ⅐ day Ϫ1 Clen or saline (Sal) for 4 wk with the use of osmotic minipumps. Ventricular myocytes were enzymatically dissociated. Cells were field stimulated at 0.5, 1, and 2 Hz, and cytoplasmic Ca 2ϩ transients were monitored with the use of the fluorescent indicator indo-1 acetoxymethyl ester. Two-dimensional surface area and action potentials in current clamp were also measured. We found that in the Clen group there was significant hypertrophy at the organ and cellular levels compared with Sal. In Clen myocytes, the amplitude of the indo-1 ratio transients was significantly increased. Sarcoplasmic reticulum Ca 2ϩ content, estimated by rapid application of 20 mM caffeine, was significantly increased in the Clen group. The action potential was prolonged in the Clen group compared with Sal. Carbohydrate contribution to the tricarboxylic cycle (Krebs cycle) flux was increased several times in the Clen group. This increase was associated with decreased expression of peroxisome proliferator-activated receptor-␣. This study shows that chronic administration of Clen induces cellular hypertrophy and increases oxidative carbohydrate utilization together with an increase in sarcoplasmic reticulum Ca 2ϩ content, which results in increased amplitude of the Ca 2ϩ transients. These effects could be important when Clen is used in conjunction with left ventricular assist devices treatment.sarcoplasmic reticulum Ca 2ϩ content; tricarboxylic cycle; left ventricular assist devices LONG-TERM LEFT VENTRICULAR (LV) assist device (LVAD) support has been shown to improve survival and quality of life in patients with advanced heart failure and contraindications to cardiac transplantation (22). LVAD support is also routinely used as a bridge to transplantation in patients with end-stage heart failure. In a number of patients, with the use of this strategy in association with pharmacological therapy (combination therapy), a significant improvement in myocardial performance has been observed. In some cases the mechanical device could be explanted without resorting to heart transplantation ("bridge to recovery") (11-13, 27, 40).LVAD support results in profound and complex changes in the structure and function of the myocardium, which include "...

Abstract-The 4.1 proteins are a family of multifunctional adaptor proteins. They promote the mechanical stability of plasma membranes by interaction with the cytoskeletal proteins spectrin and actin and are required for the cell surface expression of a number of transmembrane proteins. Protein 4.1R is expressed in heart and upregulated in deteriorating human heart failure, but its functional role in myocardium is unknown. To investigate the role of protein 4.1R on myocardial contractility and electrophysiology, we studied 4.1R-deficient (knockout) mice (4.1R KO). ECG analysis revealed reduced heart rate with prolonged Q-T interval in 4.1R KO. No changes in ejection fraction and fractional shortening, assessed by echocardiography, were found. The action potential duration in isolated ventricular myocytes was prolonged in 4.1R KO. Ca 2ϩ transients were larger and slower to decay in 4.1R KO. The sarcoplasmic reticulum Ca 2ϩ content and Ca 2ϩ sparks frequency were increased. The Na ϩ /Ca 2ϩ exchanger current density was reduced in 4.1R KO. The transient inward current inactivation was faster and the persistent Na ϩ current density was increased in the 4.1R KO group, with possible effects on action potential duration. Although no major morphological changes were noted, 4.1R KO hearts showed reduced expression of NaV1.5␣ and increased expression of protein 4.1G. Our data indicate an unexpected and novel role for the cytoskeletal protein 4.1R in modulating the functional properties of several cardiac ion transporters with consequences on cardiac electrophysiology and with possible significant roles during normal cardiac function and disease. Key Words: cardiac cytoskeleton Ⅲ ion transporter regulation Ⅲ EC coupling T he cardiac cytoskeleton is important in conferring stability to the myocardium, in sensing the mechanical stretch, and in coordinating the assembly of cellular structures and intercellular signaling. 1 One group of cytoskeletal proteins, the spectrin-and ankyrin-associated system, is involved in the complex interplay between actin, spectrin, and various ion transporters in relation to the regulation of intracellular [Ca 2ϩ ]. 2 Beta II spectrin, muscle LIM-only protein, and ankyrin B and G have all been associated with cardiac function and regulation of the electrophysiological properties of the myocardium. 3 The 4.1 protein family is also part of the spectrin-associated cytoskeleton. It promotes the interaction between spectrin and F-actin and, thus, membrane stability. 2 In mammals, the 4 genes, EPB41, EPB41L1, EPB41L2, and EPB41L3, encode proteins 4.1R, 4.1G, 4.1N, and 4.1B. mRNA transcripts from all 4 genes are found in mouse myocardium. 4 All 4.1 proteins have a FERM membranebinding domain, a spectrin-actin binding (SAB) domain, and a C-terminal domain. 2 A FERM-adjacent domain regulates the activities of both FERM and SAB domains. 5 The FERM and C-terminal domains bind to membrane proteins, 6 whereas the SAB domain binds the spectrin-actin cytoskeleton. 7 Multiple ion channels, pumps, and exc...

Significant progress has been made toward revealing molecular changes associated with myocardial recovery from heart failure. These studies also offer new insight into the pathogenesis of heart failure and point to novel therapeutic strategies.

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