(Supported by NIH NS22189 to J.R. Wolpaw) Operant conditioning of the triceps surae (TS) H-reflex in monkeys affects the F-type and C-type terminals on TS motorneurons (Feng-Chen and Wolpaw, (1996) PNAS, 93 9206-9211). It is not clear whether conditioning in primates also affects Ia afferent terminals on TS motorneurons and/or their presynaptic (P) terminals. We are developing methods for a new rat model, in which the soleus H-reflex is conditioned (rats can be trained to acquire learning skills). The crucial requirement is a method for efficiently identifying the relatively infrequent Ia afferent terminals on TS motorneurons and their P terminals, measuring their size, and determining the frequency per unit of motorneuron membrane. We are developing a triple-labeling method. Soleus motorneurons are retrogradely labeled with cholera toxin-horseradish peroxidase injected into the muscle, Ia afferent terminals are labeled with biocytin injected into the dorsal ro ots, and P terminals are labeled with GABA antiserum. Thin sections routinely prepared for electron microscopy are recorded, and the negatives digitized. Then Sterecon is used to trace the membranes and quantitate the immunogold label. A set of test measurements was made on one digitized image. Sterecon enabled the user to accurately measure membrane lengths, as well as to quantitate the number of gold particles. The statistics of the test data were acceptable, so the work proceeded. Eight more images were digitized, traced and quantitated.
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