Stimulation Induced Changes in Frog Neuromuscular Junctions: A Quantitative Ultrastructural Comparison of Rapid-Frozen and Chemically Fixed Nerve Terminals
Abstract
This study was conducted to determine the presynaptic morphological changes due to neural activity in rapidly stimulated neuromuscular junctions preserved byrapid-freezing and freeze-substitution and to ascertain what, if any, additional changes are produced by chemical fixation. Cutaneous pectoris nerve-muscle preparations from juvenile Rana pipiens were rapid-frozen against a copper block cooled to 8 deg K and freeze-substituted with OsO4 in acetone. This frozen material was compared to material which had been chemically fixed by immersion in glutaraldehyde and post-fixed in OsO4/ferrocyanide. The nerves of experimental preparations were stimulated at 15 Hz for 15 minutes with a suction electrode and then either fixed or frozen one minute after cessation of the stimulus. Rested preparations were fixed or frozen 60 minutes after cessation of the stimulating current. Control preparations were unstimulated and preserved either by chemical fixation or rapid-freezing. This study provides evidence that most of the previously reported effects of stimulation are real; that is, they are present in rapid-frozen fresh (unfixed) tissue. However, there were significant differences in the reactions of the tissue to the two types of preservation. Rapid-freezing confirmed that rapid stimulation causes a reduction in the number of synaptic vesicles and an increase in the number of cisternal profiles, changes reported by previous investigators in chemically fixed tissue. The rapid-frozen preparations in the present study showed, in addition, that rapid stimulation produces an increase in synaptic vesicle diameter, a change in the shape of mitochondria without any detectable change in their volume and a decrease in the number of microtubules. Terminals preserved by either method exhibited recovery toward control morphology with 60 minutes of rest after stimulation.
Document Details
- Document Type
- Technical Report
- Publication Date
- Mar 06, 1984
- Accession Number
- AD1010665
Entities
People
- Patricia A. Brewer
Organizations
- Uniformed Services University of the Health Sciences