|
|
| TR |
TITLE |
AUTHORS |
KEYWORDS |
MATERIALS & METHODS
|
MICROSCOPIC
TECHNIQUES |
SPECIES |
MORPHOLOGY |
CELL
LINE |
| 584 |
Characterization
of the myosin-based source for second-harmonic
generation from muscle sarcomeres. |
Sergey
V. Plotnikov, Andrew C. Millard, Paul J. Campagnola, and William A.
Mohler |
second-harmonic,
generation (SHG), sarcomere, myosin, striated muscle, SHG harmonophore |
An aliquot (0.5 ml) of isolated
scallop myofibril suspension was placed into a glass bottom Petri dish (MatTek Corp., Ashland,
MA) and the dish was centrifuged at 4000g for 10 min.
|
nonlinear
optical microscopy |
chicken |
|
cardiac
and skeletal myocytes |
| Abstract |
| |
Several biologically important protein structures
give rise to strong second-harmonic generation (SHG) in their native context. In addition to
high-contrast optical sections of cells and tissues, SHG imaging can provide detailed structural
information based on the physical constraints of the optical effect. In this study we characterize,
by biochemical and optical analysis, the critical structures underlying SHG from the complex
muscle sarcomere. SHG emission arises from domains of the sarcomere containing thick filaments,
even within nascent sarcomeres of differentiating myocytes. SHG from isolated myofibrils is
abolished by extraction of myosin, but is unaffected by removal or addition of actin filaments.
Furthermore, the polarization-dependence of sarcomeric SHG is not affected by either the proportion
of myosin head domains or the orientation of myosin heads. By fitting SHG polarization anisotropy
readings to theoretical response curves, we find an orientation for the elemental harmonophore
that corresponds well to the pitch of the myosin rod a-helix along the thick filament axis.
Taken together, these data indicate that myosin rod domains are the key structures giving SHG
from striated muscle. This study should guide the interpretation of SHG contrast in images of
cardiac and skeletal muscle tissue for a variety of biomedical applications. |
|
|
|
|
