Characterization of secophalloidin-induced force loss in cardiac myofibrils.

Secophalloidin (SPH) is known to cause in cardiac myofibrils force without Ca(2+) (half-maximal effect approximately 2 mM) followed by irreversible loss of Ca(2+)-activated force. At maximal Ca(2+) activation, SPH increases force (half-maximal effect < 0.1 mM). We found that SPH at low concentration (0.5 mM) did not cause either force activation or force loss at pCa 8.7, but both of these effects did occur when force was activated by Ca(2+). The force loss was prevented when SPH was applied during rigor or in the presence of 2,3-butanedione monoxime (85 mM). Furthermore, studying muscle in which the force was previously reduced by SPH (up to 50%) did not reveal significant changes in Ca(2+) sensitivity and cooperativity of Ca(2+) activation or qualitative alterations in SPH-induced changes in Ca(2+)-activated contraction. Data suggest that the force loss is mediated by cycling cross-bridges, and might reflect a reduction in force generated by individual cross-bridges.

Mechano-chemical effects of Ca(2+) in cross-linked troponin-C films.

Changes in troponin C (TnC) conformation upon Ca(2+) binding forms the basis for regulatory and structural functions of TnC molecules. In the present study, Ca(2+)-induced conformational changes in TnC were observed by mechanical measurements. TnC films were prepared by drying or electrospraying TnC solutions, cross-linked with glutaraldehyde, and isometric tension and stiffness measured as a function of pCa. An increase in Ca(2+) from a pCa of 9 to 4 induced large-scale mechanical changes in the TnC films causing several percent shrinkage of the unloaded films. This shrinkage could be partially assigned to Ca(2+) binding to the Ca(2+)/Mg(2+) sites of TnC.

Calcium-independent activation of skinned cardiac muscle by secophalloidin.

Thin filament regulation of muscle contraction is believed to be mediated by both Ca2+ and strongly bound myosin cross-bridges. We found that secophalloidin (SPH, 5-8 mM) activates cross-bridge cycling without Ca2+ causing isometric force comparable to that induced by Ca2+. At saturated [SPH], Ca2+ further increased force by 20%. SPH-induced force was reversible upon washing with a relaxing solution. However, there was more than 30% irreversible loss in subsequent Ca2+-activated force. We hypothesize that SPH activates muscle via strongly bound cross-bridges. SPH-activated contraction provides a new model for studying the role of Ca2+ and cross-bridges in muscle regulation.

Detection of progressive myocardial tissue injury by ultrasonic integrated backscatter immediately after coronary reperfusion.

Myocardial reperfusion following ischemia may paradoxically cause additional injury, including microvascular damage and edema. These structural alterations augment tissue echogenicity, which is measurable by ultrasonic integrated backscatter (IB). We sought to characterize alterations in myocardial IB in an ischemic and reperfused region of the rat heart. Myocardial IB of the regions of interest in 12 adult male Sprague-Dawley rats was studied at baseline, during ischemia, and chronologically after coronary reopening, using an ultrasound frequency of 8 MHz. IB did not significantly change between baseline and ischemia. However, within 1 min of reperfusion, IB significantly increased and continued to increase until 10 min of reperfusion, when a plateau was reached. Areas of high echogenicity were comparable to infarcted areas on gross pathologic slices and had edema with extravasation of red blood cells. Myocardial reperfusion following ischemia significantly augments tissue echogenicity. A continuing increase of IB suggests a rapid progression of reperfusion injury.

The left ventricle responds to acute graded elevation of right ventricular afterload by augmentation of twist magnitude and untwist rate.

BACKGROUND: The right and left ventricles share the interventricular septum, which mechanically transmits pressure gradients. The aim of this study was to investigate how acute mild or moderate right ventricular (RV) afterload affects left ventricular (LV) function. METHODS: In 14 open-chest pigs (mean weight, 43 ± 4 kg) with preserved pericardium, acute mild (>35 and ≤50 mm Hg) and moderate (>50 and ≤60 mm Hg) RV pressure loading conditions were induced by constriction of the pulmonary artery. Hemodynamic parameters and LV twist and untwist were evaluated under each condition. RESULTS: From baseline to mild and moderate RV afterload, the mean RV systolic pressure increased from 31.0 ± 4.3 to 41.1 ± 2.7 and 52.7 ± 3.4 mm Hg (P < .001), while LV twist magnitudes increased from 15.4 ± 5.1° to 18.5 ± 3.1° and 19.8 ± 5.0° (P = .004), respectively. Absolute values of LV untwist rate increased from -116.9 ± 64.9°/sec to -160.0 ± 53.3°/sec and -169.1 ± 47.0°/sec, respectively (P = .001). After adjusting for all variables, only the ratio of the early and atrial components of mitral inflow and RV outflow tract acceleration time was significantly associated with the LV twist magnitude and LV untwist rate. CONCLUSIONS: In an acute setting, the left ventricle responds to suddenly elevated RV afterload and decreased RV stroke volume by promptly increasing its twist magnitude and untwist rate.

Secophalloidin as a novel activator of skinned cardiac muscle.

Secophalloidin (SPH) is known to activate skinned cardiac muscle in the absence of Ca(2+). We hypothesized that SPH-induced changes in cross-bridge properties underlie muscle activation. We found that force responsiveness to orthovanadate was drastically reduced in SPH activated muscles compared to Ca(2+)-activated contraction. Moreover, SPH caused approximately 30% increase in Ca(2+)-independent force in muscles where Ca(2+) sensitivity was totally destroyed by troponin I extraction with 10mM vanadate. Thus, SPH and Ca(2+) activation differ in both properties of the cross-bridge cycle and protein requirements for thin filament regulation. In addition, we tested the relationship between the activating effects SPH and EMD 57033, a Ca(2+) sensitizer that increases resting force in cardiac muscle. After maximal activation by either SPH or EMD 57033, the other compound was found to further increase force, indicating that SPH activates muscle via a novel mechanism.