Radioautographs of (45)Ca-labeled frog skeletal muscles have been prepared using freeze-dry and vapor fixation techniques to avoid displacement of the isotope during the preparation of the radioautographs. (45)Ca has been localized in resting muscles exposed to (45)Ca Ringer's for 5 min or 5 hr and in isotopically labeled muscles recovering from tetanic stimulation at room temperature or at 4 degrees C. In muscles soaked at rest for 5 min (45)Ca was present almost exclusively in the terminal cisternae. In all other muscles there were three sites at which the isotope was concentrated: (a) the terminal cisternae, (b) the intermediate cisternae and the longitudinal tubules, and (c) the A band portion of the myofibrils. The terminal cisternae were labeled more rapidly than the myofibrils, but both exchanges were accelerated by electrical stimulation. The amount of (45)Ca in the longitudinal tubules and the intermediate cisternae decreased with time after a tetanus as the amount in the terminal cisternae increased. It is proposed that electrical stimulation releases calcium from the terminal cisternae and that relaxation occurs from the binding of the released calcium by the longitudinal tubules and the intermediate cisternae. Complete recovery from mechanical activity involves the transport of this bound calcium into the reticulum and its subsequent binding by the terminal cisternae. Resting exchange of calcium occurs primarily between the terminal cisternae and the transverse tubules.