Side bridge geometry after quick-freezing of stimulated and unstimulated frog skeletal muscle fibers

Abstract

Quick-freezing allows the structural analysis of timed perturbations of morphology. We are presenting preliminary results concerning the feasibility of studying directly the side bridge geometry of actin-myosin interactions within the time course of a twitch in single intact frog skeletal muscle fibers, both by freeze-substitution and freeze-fracture after quick-freezing, and following various time intervals between stimulation and impact of the fibers on a liquid He-cooled copper block.Materials and Methods. The quick-freezing device was a "Slammer"(Polaron) for which the electronics had been redesigned; they are capable, in combination with a Grass S48 stimulator, of any stimulation interval between 0 and 1 sec prior to freezing, including tetanus. The actual elapsed time between stimulation and freezing is recorded with a digital clock. Single intact tendonto- tendon frog skeletal muscle fibers (semitendinosus of r. temporaria) or toe muscle bundles (r.pipiens) were isolated by sharp dissection and placed between coextensive Pt stimulation wires on blackened 2% agarose, the height of which on the specimen holder was adjusted appropriately with respect to a spacer ring both, to calibrate the impact time and to prevent smashing of the fibers.