TY - JOUR
T1 - Rotation of actin monomers during isometric contraction of skeletal muscle
AU - Borejdo, Julian
AU - Muthu, Priya
AU - Talent, John
AU - Akopova, Irina
AU - Burghardt, Thomas P.
N1 - Funding Information:
We thank Drs. Z. and I. Gryczynski for helpful suggestions and comments on a manuscript. Supported by R21CA9732 and R01AR048622 (NIAMS) (JB) and by R01AR049277 (NIAMS) and the Mayo Foundation (TPB).
PY - 2007/1
Y1 - 2007/1
N2 - Cyclic interactions of myosin and actin are responsible for contraction of muscle. It is not self-evident, however, that the mechanical cycle occurs during steady-state isometric contraction where no work is produced. Studying cross-bridge dynamics during isometric steady-state contraction requires an equilibrium time-resolved method (not involving application of a transient). This work introduces such a method, which analyzes fluctuations of anisotropy of a few actin molecules in muscle. Fluorescence anisotropy, indicating orientation of an actin protomer, is collected from a volume of a few attoliters (10-18 L) by confocal total internal reflection (CTIR) microscopy. In this method, the detection volume is made shallow by TIR illumination, and narrow by confocal aperture inserted in the conjugate image plane. The signal is contributed by approximately 12 labeled actin molecules. Shortening of a myofibril during contraction is prevented by light cross-linking with 1-ethyl-3-[3-dimethylamino)-propyl]-carbodiimide. The root mean-squared anisotropy fluctuations are greater in isometrically contracting than in rigor myofibrils. The results support the view that during isometric contraction, cross-bridges undergo a mechanical cycle.
AB - Cyclic interactions of myosin and actin are responsible for contraction of muscle. It is not self-evident, however, that the mechanical cycle occurs during steady-state isometric contraction where no work is produced. Studying cross-bridge dynamics during isometric steady-state contraction requires an equilibrium time-resolved method (not involving application of a transient). This work introduces such a method, which analyzes fluctuations of anisotropy of a few actin molecules in muscle. Fluorescence anisotropy, indicating orientation of an actin protomer, is collected from a volume of a few attoliters (10-18 L) by confocal total internal reflection (CTIR) microscopy. In this method, the detection volume is made shallow by TIR illumination, and narrow by confocal aperture inserted in the conjugate image plane. The signal is contributed by approximately 12 labeled actin molecules. Shortening of a myofibril during contraction is prevented by light cross-linking with 1-ethyl-3-[3-dimethylamino)-propyl]-carbodiimide. The root mean-squared anisotropy fluctuations are greater in isometrically contracting than in rigor myofibrils. The results support the view that during isometric contraction, cross-bridges undergo a mechanical cycle.
KW - Actin
KW - Isometric contraction
KW - Microscopy
KW - Muscle
KW - Total internal reflection
UR - http://www.scopus.com/inward/record.url?scp=34247648050&partnerID=8YFLogxK
U2 - 10.1117/1.2697286
DO - 10.1117/1.2697286
M3 - Article
C2 - 17343488
AN - SCOPUS:34247648050
SN - 1083-3668
VL - 12
JO - Journal of Biomedical Optics
JF - Journal of Biomedical Optics
IS - 1
M1 - 014013
ER -