Voluntary exercise delays heart failure onset in rats with pulmonary artery hypertension.

Increased physical activity is recommended for the general population and to patients of many diseases because of its health benefits but can be contraindicated if it is thought a risk for serious cardiovascular events. One such condition is pulmonary artery hypertension (PAH). PAH and right ventricular failure was induced in rats by a single injection of monocrotaline (MCT). MCT rats with voluntary access to a running wheel ran on average 2km per day. The time for half the animals to develop heart failure signs (median survival time) was 28 days (exercise failure (EF) group), significantly longer than sedentary animals (sedentary failure (SF) group), 23 days). The contractility of single failing myocytes in response to increasing demand (stimulation frequency) was significantly impaired compared with both sedentary control (SC) and exercising control (EC) myocytes. However, myocytes from exercising MCT rats, tested at 23 days (EM group) showed responses intermediate to the control (SC, EC) and failing (SF, EF) groups. We conclude that voluntary exercise is beneficial to rats with heart failure induced by PAH and this is evidence to support the consideration of appropriate exercise regimes for potentially vulnerable groups

Cellular Hypertrophy and Increased Susceptibility to Spontaneous Calcium-Release of Rat Left Atrial Myocytes Due to Elevated Afterload

Atrial remodeling due to elevated arterial pressure predisposes the heart to atrial fibrillation (AF). Although abnormal sarcoplasmic reticulum (SR) function has been associated with AF, there is little information on the effects of elevated afterload on atrial Ca2+-handling. We investigated the effects of ascending aortic banding (AoB) on Ca2+-handling in rat isolated atrial myocytes in comparison to age-matched sham-operated animals (Sham). Myocytes were either labelled for ryanodine receptor (RyR) or loaded with fluo-3-AM and imaged by confocal microscopy. AoB myocytes were hypertrophied in comparison to Sham controls (P<0.0001). RyR labeling was localized to the z-lines and to the cell edge. There were no differences between AoB and Sham in the intensity or pattern of RyR-staining. In both AoB and Sham, electrical stimulation evoked robust SR Ca2+-release at the cell edge whereas Ca2+ transients at the cell center were much smaller. Western blotting showed a decreased L-type Ca channel expression but no significant changes in RyR or RyR phosphorylation or in expression of Na+/Ca2+ exchanger, SR Ca2+ ATPase or phospholamban. Mathematical modeling indicated that [Ca2+]i transients at the cell center were accounted for by simple centripetal diffusion of Ca2+ released at the cell edge. In contrast, caffeine (10 mM) induced Ca2+ release was uniform across the cell. The caffeine-induced transient was smaller in AoB than in Sham, suggesting a reduced SR Ca2+-load in hypertrophied cells. There were no significant differences between AoB and Sham cells in the rate of Ca2+ extrusion during recovery of electrically-stimulated or caffeine-induced transients. The incidence and frequency of spontaneous Ca2+-transients following rapid-pacing (4 Hz) was greater in AoB than in Sham myocytes. In conclusion, elevated afterload causes cellular hypertrophy and remodeling of atrial SR Ca2+-release

Caveolae in Rabbit Ventricular Myocytes: Distribution and Dynamic Diminution after Cell Isolation

Caveolae are signal transduction centers, yet their subcellular distribution and preservation in cardiac myocytes after cell isolation are not well documented. Here, we quantify caveolae located within 100 nm of the outer cell surface membrane in rabbit single-ventricular cardiomyocytes over 8 h post-isolation and relate this to the presence of caveolae in intact tissue. Hearts from New Zealand white rabbits were either chemically fixed by coronary perfusion or enzymatically digested to isolate ventricular myocytes, which were subsequently fixed at 0, 3, and 8 h post-isolation. In live cells, the patch-clamp technique was used to measure whole-cell plasma membrane capacitance, and in fixed cells, caveolae were quantified by transmission electron microscopy. Changes in cell-surface topology were assessed using scanning electron microscopy. In fixed ventricular myocardium, dual-axis electron tomography was used for three-dimensional reconstruction and analysis of caveolae in situ. The presence and distribution of surface-sarcolemmal caveolae in freshly isolated cells matches that of intact myocardium. With time, the number of surface-sarcolemmal caveolae decreases in isolated cardiomyocytes. This is associated with a gradual increase in whole-cell membrane capacitance. Concurrently, there is a significant increase in area, diameter, and circularity of sub-sarcolemmal mitochondria, indicative of swelling. In addition, electron tomography data from intact heart illustrate the regular presence of caveolae not only at the surface sarcolemma, but also on transverse-tubular membranes in ventricular myocardium. Thus, caveolae are dynamic structures, present both at surface-sarcolemmal and transverse-tubular membranes. After cell isolation, the number of surface-sarcolemmal caveolae decreases significantly within a time frame relevant for single-cell research. The concurrent increase in cell capacitance suggests that membrane incorporation of surface-sarcolemmal caveolae underlies this, but internalization and/or micro-vesicle loss to the extracellular space may also contribute. Given that much of the research into cardiac caveolae-dependent signaling utilizes isolated cells, and since caveolae-dependent pathways matter for a wide range of other study targets, analysis of isolated cell data should take the time post-isolation into account

Caveolin contributes to the modulation of basal and β-adrenoceptor stimulated function of the adult rat ventricular myocyte by simvastatin: A novel pleiotropic effect

The number of people taking statins is increasing across the globe, highlighting the Importance of fully understanding statins effects on the cardiovascular system. The beneficial impact of statins extends well beyond regression of atherosclerosis to include direct effects on tissues of the cardiovascular system (pleiotropic effects). Pleiotropic effects on the cardiac myocyte are often overlooked. Here we consider the contribution of the caveolin protein, whose expression and cellular distribution is dependent on cholesterol, to statin effects on the cardiac myocyte. Caveolin is a structural and regulatory component of caveolae, and is a key regulator of cardiac contractile function and adrenergic responsiveness. We employed an experimental model in which inhibition of myocyte HMG CoA reductase could be studied in the absence of paracrine influences from non-myocyte cells. Adult rat ventricular myocytes were treated with 10 μM simvastatin for 2 days. Simvastatin treatment reduced myocyte cholesterol, caveolin 3 and caveolar density. Negative inotropic and positive lusitropic effects (with corresponding changes in [Ca2]¡) were seen in statin-treated cells. Simvastatin significantly potentiated the inotropic response to β2-, but not β1-, adrenoceptor stimulation. Under conditions of β2-adrenoceptor stimulation, phosphorylation of phospholamban at Ser16and troponin I at Ser23/24was enhanced with statin treatment. Simvastatin increased NO production without significant effects on eNOS expression or phosphorylation (Ser1177), consistent with the reduced expression of caveolin 3, its constitutive Inhibitor. In conclusion, statin treatment can reduce caveolin 3 expression, with functional consequences consistent with the known role of caveolae in the cardiac cell. These data are likely to be of significance, particularly during the early phases of statin treatment, and in patients with heart failure who have altered ß-adrenoceptor signalling. In addition, as caveolin is ubiquitously expressed and has myriad tissue-specific functions, the impact of statin-dependent changes in caveolin is likely to have many other functional sequelae

The cellular basis for enhanced volume-modulated cardiac output in fish hearts

During vertebrate evolution there has been a shift in the way in which the heart varies cardiac output (the product of heart rate and stroke volume). While mammals, birds, and amphibians increase cardiac output through large increases in heart rate and only modest increases (∼30%) in stroke volume, fish and some reptiles use modest increases in heart rate and very large increases in stroke volume (up to 300%). The cellular mechanisms underlying these fundamentally different approaches to cardiac output modulation are unknown. We hypothesized that the divergence between volume modulation and frequency modulation lies in the response of different vertebrate myocardium to stretch. We tested this by progressively stretching individual cardiac myocytes from the fish heart while measuring sarcomere length (SL), developed tension, and intracellular Ca(2+) ([Ca(2+)](i)) transients. We show that in fish cardiac myocytes, active tension increases at SLs greater than those previously demonstrated for intact mammalian myocytes, representing a twofold increase in the functional ascending limb of the length–tension relationship. The mechanism of action is a length-dependent increase in myofilament Ca(2+) sensitivity, rather than changes in the [Ca(2+)](i) transient or actin filament length in the fish cell. The capacity for greater sarcomere extension in fish myocardium may be linked to the low resting tension that is developed during stretch. These adaptations allow the fish heart to volume modulate and thus underpin the fundamental difference between the way fish and higher vertebrates vary cardiac output

Effects of cholesterol depletion on compartmentalized cAMP responses in adult cardiac myocytes

β1-Adrenergic receptors (β1ARs) and E-type prostaglandin receptors (EPRs) both produce compartmentalized cAMP responses in cardiac myocytes. The role of cholesterol-dependent lipid rafts in producing these compartmentalized responses was investigated in adult rat ventricular myocytes. β1ARs were found in lipid raft and non-lipid raft containing membrane fractions, while EPRs were only found in non-lipid raft fractions. Furthermore, β1AR activation enhanced the L-type Ca2+ current, intracellular Ca2+ transient, and myocyte shortening, while EPR activation had no effect, consistent with the idea that these functional responses are regulated by cAMP produced by receptors found in lipid raft domains. Using methyl-β-cyclodextrin to disrupt lipid rafts by depleting membrane cholesterol did not eliminate compartmentalized behavior, but it did selectively alter specific receptor-mediated responses. Cholesterol depletion enhanced the sensitivity of functional responses produced by β1ARs without having any effect on EPR activation. Changes in cAMP activity were also measured in intact cells using two different FRET-based biosensors: a type II PKA-based probe to monitor cAMP in subcellular compartments that include microdomains associated with caveolar lipid rafts and a freely diffusible Epac2-based probe to monitor total cytosolic cAMP. β1AR and EPR activation elicited responses detected by both FRET probes. However, cholesterol depletion only affected β1AR responses detected by the PKA probe. These results indicate that lipid rafts alone are not sufficient to explain the difference between β1AR and EPR responses. They also suggest that β1AR regulation of myocyte contraction involves the local production of cAMP by a subpopulation of receptors associated with caveolar lipid rafts