Fropofol decreases force development in cardiac muscle

Abstract

Supranormal contractile properties are frequently associated with cardiac diseases. Anesthetic agents, including propofol, can depress myocardial contraction. We tested the hypothesis that fropofol, a propofol derivative, reduces force development in cardiac muscles via inhibition of cross-bridge cycling and may therefore have therapeutic potential. Force and intracellular Ca²⁺ concentration ([Ca²⁺]_i) transients of rat trabecular muscles were determined. Myofilament ATPase, actin-activated myosin ATPase, and velocity of actin filaments propelled by myosin were also measured. Fropofol dose dependently decreased force without altering [Ca²⁺]_i in normal and pressure-induced hypertrophied-hypercontractile muscles. Similarly, fropofol depressed maximum Ca²⁺-activated force ( F_max) and increased the [Ca²⁺]_i required for 50% of F_max (Ca₅₀) at steady state without affecting the Hill coefficient in both intact and skinned cardiac fibers. The drug also depressed cardiac myofibrillar and actin-activated myosin ATPase activity. In vitro actin sliding velocity was significantly reduced when fropofol was introduced during rigor binding of cross-bridges. The data suggest that the depressing effects of fropofol on cardiac contractility are likely to be related to direct targeting of actomyosin interactions. From a clinical standpoint, these findings are particularly significant, given that fropofol is a nonanesthetic small molecule that decreases myocardial contractility specifically and thus may be useful in the treatment of hypercontractile cardiac disorders.-Ren, X., Schmidt, W., Huang, Y., Lu, H., Liu, W., Bu, W., Eckenhoff, R., Cammarato, A., Gao, W. D. Fropofol decreases force development in cardiac muscle.

Keywords: excitation contraction coupling; fropofol; in vitro motility; intracellular calcium; myofilament protein.

Figure 1

Fropofol decreases twitch force development in normal cardiac muscle. A) Raw recordings of force development (left) and corresponding intracellular Ca²⁺ transients (right) from a normal trabecular muscle in the presence of 0, 50, and 200 µM fropofol. Fropofol dose dependently reduced force development, but intracellular Ca²⁺ remained unchanged. B) Pooled data of trabecular force development and intracellular Ca²⁺ transient amplitudes in the presence of different doses of fropofol. Force decreased in a dose-dependent manner as concentrations of fropofol increased. Force became significantly less than that at baseline at doses >20 μM. *P < 0.01, vs. baseline, by paired Student’s t test. Maximal depression of force (∼28%) was achieved at 200 µM. Intracellular Ca²⁺ transient amplitudes remained unchanged at all doses of fropofol. C) Effect of fropofol on diastolic force and intracellular Ca²⁺ levels. Fropofol had no effect on diastolic force and intracellular Ca²⁺. D) Effect of fropofol on time to peak force and time from peak to half relaxation. Fropofol did not affect force dynamics. E) Effect of fropofol on the time course of intracellular Ca²⁺ transients. Both the time to peak and the time to half relaxation were not affected. Temperature, 22°C; external Ca²⁺, 1.0 mM; stimulation rate, 0.5 Hz (n = 8–11).

Figure 2

MCT injection increases cardiac muscle contractility. Changes in RV wall fractional shortening after intra-abdominal MCT injection. RV free-wall fractional shortening were significantly elevated at 2.5 wk after MCT injection (n = 4–7 animals examined by transthoracic echocardiography at each time point). *P < 0.05, increase vs. baseline.

Figure 3

Fropofol decreases force development in hypercontractile muscles. A) Raw recordings of force development (left) and corresponding intracellular Ca²⁺ transients (right) of a trabecular muscle from the RV of MCT-injected rats in the presence of 0, 50, and 200 µM fropofol. Fropofol decreased force in a dose-dependent manner, but intracellular Ca²⁺ remained unchanged. B) Pooled data of force development and amplitudes of intracellular Ca²⁺ transient in the presence of different doses of fropofol. Force became significantly lower than that at baseline at doses >20 μM. *P < 0.01 vs. baseline, paired Student’s t test. Maximum depression of force (∼35%) was achieved at 200 µM. Intracellular Ca²⁺ transient amplitudes remained unchanged at all doses of fropofol. C) Effect of fropofol on diastolic force and intracellular Ca²⁺ levels. Significant decreases in diastolic force were seen at fropofol doses higher than 20 µM. *P < 0.05 vs. baseline, paired Student’s t test. There were no changes in diastolic Ca²⁺ levels. D) Effect of fropofol on time to peak force and time from peak to half relaxation. Fropofol accelerated relaxation of force. E) Effect of fropofol on time course of intracellular Ca²⁺ transients. Both the time to peak and the time to half relaxation were not affected. Temperature, 22°C; external Ca²⁺, 1.0 mM; stimulation rate, 0.5 Hz (n = 9–11).

Figure 4

Relationship between steady-state force and intracellular Ca²⁺ in the presence and absence of fropofol (100 µM) in intact and skinned cardiac trabecular muscles. The steady-state forces were plotted against corresponding intracellular Ca²⁺ transients. A) In normal intact trabeculae, fropofol significantly reduced F_max and increased Ca₅₀ (n = 10). B) The same muscles in which steady-state relations were first obtained were chemically skinned and activated with various Ca²⁺ concentrations in the absence and presence of fropofol. Note that the effect of fropofol on F_max and Ca₅₀ persisted in these skinned muscles (n = 9). C) In intact trabeculae from MCT-injected rats, fropofol significantly reduced F_max and increased Ca₅₀ (n = 10). D) The same muscles in which steady-state relations were first obtained were chemically skinned and activated with various Ca²⁺ concentrations in the absence and presence of fropofol. The effect of fropofol on F_max and Ca₅₀ persisted in these skinned muscles (n = 9). Temperature, 22°C.

Figure 5

Effect of fropofol on myofibrillar Mg²⁺-ATPase activity, actin sliding velocity, and myosin ATPase activity of rat myocardium. A) Fropofol significantly depressed the myofibrillar Mg²⁺-ATPase activity–Ca²⁺ relationship, as evidenced by a decrease in maximum Ca²⁺-activated ATPase activity and a rightward shift of the curve (n = 5). Temperature, 22°C. P < 0.05, multivariate ANOVA. B) Fropofol slowed actin sliding speed by 9 and 12% at 100 and 300 µM, respectively, with significance reached at 300 µM. Inset: decrease in absolute velocity of actin sliding over myosin at 300 µM fropofol (n = 24 filaments). **P < 0.01 vs. 0 fropofol, unpaired Student’s t test. C) Myosin ATPase activity was depressed at increasing doses of fropofol. At doses above 100 µM, fropofol significantly depressed actin-activated myosin ATPase activity (actin, 0.4 mg/ml), but had no effect on myosin ATPase activity in the absence of actin (n = 5). *P < 0.03, multivariate ANOVA. D) The actin-myosin ATPase relationship was also depressed by 200 µM fropofol (n = 3–5). *P < 0.05, multivariate ANOVA.