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We examined the effects of pharmacological alteration of Ca2+sources on mechanical and energetic properties of paired-pulse (“bigeminic”) contractions. The fraction of heat release that is related to pressure development and pressure-independent heat release were measured during isovolumic contractions in arterially perfused rat ventricles. The heat released by regular and bigeminic contractions showed two brief pressure-independent components (H1 and H2) and a pressure-dependent component (H3). We used the ratio of active heat (H′a) to pressure-time integral (PtI) and the ratio of H3 to PtI to estimate the energetic cost of muscle contraction (overall economy) and pressure maintenance (contractile economy), respectively. Neither of these ratios was affected by stimulation pattern. Caffeine (an inhibitor of sarcoplasmic reticulum function) significantly decreased mechanical responses and increased the energetic cost of contraction (Δ = 101 ± 12.6%). Verapamil (an L-type Ca2+channel blocker) decreased pressure maintenance of extrasystolic (Δ = 43.4 ± 3.7%) and postextrasystolic (Δ = 37.5 ± 3.5%) contractions without affecting postextrasystolic potentiation, suggesting that a verapamil-insensitive fraction is responsible for potentiation. The verapamil-insensitive fraction was further studied in the presence of lithium (45 mM) and KB-R7943 (5 μM), inhibitors of the Na+/Ca2+exchanger. Both agents decreased all mechanical responses, including postextrasystolic potentiation (Δ = 67.3 ± 3.3%), without altering overall or contractile economies, suggesting an association of the verapamil-insensitive Ca2+fraction to the sarcolemmal Na+/Ca2+exchanger. The effect of the inhibitors of the Na+/Ca2+exchanger on potentiation suggests an increased participation of extracellular Ca2+(and, thus, a redistribution of the relative participation of the Ca2+pools) during bigeminic contractions in rat myocardium.

Chagasic (Ch) and nonchagasic (NCh) IgG fraction (20 μg/ml) effects on cardiac performance of adult Wistar rat ventricles were studied with a novel approach applying a microcalorimetric technique. Resting heat ( Hr) was significantly decreased by Ch antibodies (Δ HrCh = 4.8 ± 0.9 mW/g). Although the Hr decrease can be associated with diminished activity of the Na+/K+ pump, the magnitude of the effect (25% of control Hr) indicates that additional processes may also be affected. Ch antibodies induced an initial increase in developed pressure (P), which was associated with a decreased contractile economy. However, after 30 min of Ch antibody perfusion, P reached a significantly lower level (ΔPCh = 3.8 ± 1.2 mN/mm2) without changes in active heat per beat ( Ha). Consequently, Ha/P ratio increased, indicating that the energetic cost per unit of P was higher. In contrast, P and Ha were both significantly and reversibly decreased by NCh antibodies (ΔPNCh = 4.4 ± 1.2 mN/mm2; Δ HaNCh = 9.7 ± 2.2 mJ/g), but Ha/P remained unaffected. According to these data, normal hearts exposed to Ch antibodies present a biphasic mechanical response: 1) an initial period of increased contractility (and decreased global muscle economy) consistent with antibodies with β1-adrenergic activity, such as those used in the present study, and 2) a decrease in P at 30 min of Ch antibody perfusion, which suggests that another Ca2+-related mechanism is compromised. These data contribute to redefine the role of antibody-mediated responses in the pathophysiology of chronic chagasic cardiomyopathy as agents of myocardial failure.

The consequences of an extrasystole (ES) on cardiac muscle’s energetics and Ca2+homeostasis were investigated in the beating heart. The fraction of heat release related to pressure development (pressure dependent) and pressure-independent heat release were measured during isovolumic contractions in arterially perfused rat ventricle. The heat release by a contraction showed two pressure-independent components (H1 and H2) of short evolution and a pressure-dependent component (H3). The additional heat released by ES was decomposed into one pressure-independent ([Formula: see text]) and one pressure-dependent ([Formula: see text]) component with time courses similar to those of control components H2 and H3. ES also induced the potentiation of pressure development (P) and heat release during the postextrasystolic (PES) beat. The slope of the linear relationship between pressure-dependent heat and pressure maintenance was similar in control, ES, and PES contractions (0.08 ± 0.01, 0.10 ± 0.02, and 0.08 ± 0.01 mJ ⋅ g−1⋅ mmHg−1⋅ s−1, respectively). The potentiation of H2 (heat component related with Ca2+removal processes) in PES was equal to [Formula: see text] at 0.3, 0.5, 1, and 2 mM Ca2+, suggesting that the extra amount of Ca2+mobilized during ES was recycled in PES. Pretreatment with 1 mM caffeine to deplete sarcoplasmic reticulum Ca2+content inhibited both the mechanical and energetic potentiation of PES. However, the heat released and the pressure developed during ES were not changed by sarcoplasmic reticulum depletion. The results suggest that 1) the source of Ca2+for ES would be entirely extracellular, 2) the Ca2+entered during ES is accumulated in the sarcoplasmic reticulum, and 3) the Ca2+stored by the sarcoplasmic reticulum during ES induces an increased contribution of this organelle during PES compared with the normal contraction.

Tension-dependent (TDH) and tension-independent heat (TIH) release were measured during single isovolumetric contractions in the arterially perfused rat ventricle. Under perfusion with 7 mM K-0.5 mM Ca, TDH showed only one component (H3), whereas TIH could be divided into two components (H1and H2) of short evolution (similar to the classically identified activation heat) and one component (H4) of long duration (dependent on mitochondrial respiration). Under 25 mM K, TIH components (i.e., H1, H2, and H4) increased with the increase in extracellular Ca concentration ([Ca]o) from 0.5 to 4 mM, and H3correlated with pressure at all [Ca]o, with regression parameters similar to those observed under 7 mM K. Under 25 mM K-2 mM Ca, peak pressure development (P), H1, H2, and H3, plotted against the number of beats under 0.4 μM verapamil, exponentially decreased, but H4decreased to 5.5 ± 2.9% in the first contraction and remained constant thereafter. Under hypoxia, P, H1, H2, and H3progressively decreased for about six contractions, but H4was not detectable from the second contraction. The results suggest that increasing extracellular K concentration decreases contractile economy mainly by increasing energy expenditure related to a Ca-dependent (verapamil-sensitive) mitochondrial activity that is not related to force generation.

A new calorimetry method has been developed to measure heat production from heart cell suspensions under continuous perfusion. The method is technically independent of the temperature at which the measurements are made, allows full control of the perfusion media, and is suitable for various biological preparations such as cells from diverse tissues, membrane vesicles, or skinned cells. The resting heat rate (Hr) measured at 18.5 degrees C in three different species (19.2 +/- 0.43, 12.8 +/- 0.56, and 9.4 +/- 0.52 mW/g dry wt for rat, guinea pig, and rabbit ventricular myocytes, respectively) agrees with that obtained with other methodologies such as oxygen consumption, thermopiles, and whole heart calorimetry. The Hr measurements showed an excellent correlation with the percentage of rod-shaped cells, indicating that rounded cells are metabolically inactive. Although the time course of the effect of increasing extracellular [K] was dependent on the species, the new steady level of Hr observed under higher extracellular [K] was significantly higher in all three species (+8.3 +/- 1.2, +9.5 +/- 4.0, and +9.3 +/- 2.7 mW/g dry wt for rat, guinea pig, and rabbit ventricular cells, respectively). This indicates that the commonly used "arrested-heart" preparation (with high extracellular [K]) for evaluation of basal metabolism most probably overestimates the real resting values. The present results also show that the wide range of resting metabolism reported in whole tissue is not due to cellular heterogeneity nor to myocyte interaction and supports the idea of an inverse relationship between resting metabolism and body weight or animal size across species.

The mechanical and energetic effects of verapamil (VER) and reduction of extracellular Ca concentration ([Ca]o) were studied in the interventricular rabbit septa and the dog papillary muscle. Even though the negative inotropic effects of VER [i.e., decrease in developed tension (T), maximal rates of contraction (+T) and relaxation (-T), and tension time integral] qualitatively resemble [Ca]o reduction, VER also elicited an anti-relaxant effect (decrease in -T/T and prolongation of the last phase of relaxation) that was not found with [Ca]o reduction. Resting heat production was similar in both preparations and remained unaffected either by changes in [Ca]o or by the presence of VER. The ratio between T and active heat production per beat (H'a) under constant fiber length decreased with VER, and this decreased economy of contraction was more marked with the increase in contraction frequency. Conversely, the T/H'a remained unaltered with changes in [Ca]o. Tension-independent heat decreased in the presence of VER and, although muscle economy can be improved by increasing muscle length in a VER-treated muscle, it is not possible to achieve either the maximal T or the maximal contraction economy that can be obtained by stretching a nontreated muscle. It may be concluded that at constant fiber length and frequency of contraction VER decreases myocardial contractile force, impairs relaxation, and decreases contraction economy. Neither the mechanical nor the energetic effects of VER can be explained solely on the basis of a reduced extracellular Ca availability, so that either the density of the Ca that enters through the channel is different from that of other sources of Ca or VER has an effect on the cross-bridge cycling mechanism.

A method has been developed to measure myocardial heat production simultaneously with mechanical (developed tension, rate of contraction and relaxation) and metabolic parameters in the arterially perfused interventricular septum of the rabbit. The system allows control of rest tension, frequency of contraction, temperature, and composition of the perfusate. The technique is based on the differential measurement of the heat flux from the muscle to the calorimetric bath. This technique is able to resolve changes in heat production as small as 0.06 mW. The resting heat production measured with the present calorimeter (1.62 +/- 0.1 mW/g wet tissue) agrees with that obtained with thermopiles and with that calculated from measurements of oxygen consumption. The heat per contraction (6.5 +/- 1.4 mJ/g wet tissue) also agreed with that measured with thermopiles in rabbit papillary muscles. The heat production measured at 22 degrees C under severe hypoxia can be fully explained by the addition of the expected change of enthalpy due to the reaction 0.5 glucose-lactate, calculated on the basis of the lactate measured in the perfusate, and the expected change of enthalpy of oxygen consumption [assuming that all remaining O2 in perfusate (0.09 vol%) is used for combustion of glucose]. These results clearly demonstrated the feasibility of this method for the correlation of changes in energy turnover, through the measurement of the heat production, with mechanical and metabolic processes on-line in arterially perfused septum.