טכניון מכון טכנולוגי לישראל
הטכניון מכון טכנולוגי לישראל - בית הספר ללימודי מוסמכים  
M.Sc Thesis
M.Sc StudentHarary Tamar
SubjectQuantification of Cross-bridge Cycling between the Different
Physicochemical Conformations by Optical Means
DepartmentDepartment of Biomedical Engineering
Supervisor Professor Amir Landesberg
Full Thesis textFull thesis text - English Version


Abstract

Background: In the past thirty years different experimental techniques have been explored in attempt to elucidate the structural dynamic changes of the contractile proteins inside the sarcomere during muscle contraction. We hypothesized that the temporal transitions of the cross-bridge (XB) motors between the different conformations during muscle contraction can be further characterized by analyzing the changes in the transmitted light polarization. Methods: Thin trabeculae (n=6) were isolated from rat right ventricle (K-H solution, 25°C). Sarcomere length (SL) was measured by laser diffraction technique and controlled by a fast servomotor. The elliptical polarization changes were quantified by utilizing a linear polarized incident light that was aligned at an angle of 45° relative to the longitudinal fiber axis.   The transmitted light passed though linear polarizer towards the photo detector. The changes in the transmitted light polarization were measured at all the polarizer angles during rest, muscle and sarcomere isometric contraction (1.95[?m]), and at different extracellular calcium concentrations (0.75, 1.5, 4.5 [mM]). Results: Passive length oscillations had mild effects on the transmitted light polarization (1.70?1.70). However, large changes in the polarization were observed during active contraction. Controlled sarcomere isometric contractions had larger effects on the transmitted elliptical polarization (17.00?10.60, n=6, p<0.01) compared to muscle isometric contraction (10.80?6.40). The kinetics of the polarization changes differed from the kinetics of force development. The changes in the polarization preceded the force development at the beginning of the contraction (51.7?18.9 [msec]) and lagged behind force relaxation at the end of the contraction (50?41 [msec]). The rate of changes in the polarization depended on the extracellular calcium level. The increase in the extracellular calcium (from 0.75 to 1.5 and 4.5 [mM]) was associated with an increase in the force (from 36?14.5 to 106.2?51.6 and 152.7?42.2 [], respectively) and a larger shift in the polarization at the same SL (, and , n=3, p<0.01). Conclusions: The plausible explanation to the observations is that the movement of myosin heads toward actin filament and XB attachment is the dominant mechanism that modifies the transmitted light polarization under physiological conditions. Significance: optical measurements provide additional information about XB dynamics that differs from the observed dynamics of force or stiffness measurements, and can be used for quantifying cardiac muscle activation and XB cycling.