Coexistence of corticotropin releasing factor and enkephalin in cerebellar afferent systems

In this study, we tested the hypothesis that corticotropin-releasing factor (CRF) and enkephalin (ENK) coexist within restricted brainstem-cerebellar circuits. Antisera for CRF and ENK were applied simultaneously or to serial sections of adult opossum brain and were processed for fluorescence or peroxidase, antiperoxidase immunohistochemistry, respectively. CRF and ENK were colocalized in 1) climbing fibers within the flocculus and in bilateral foci in the cerebellar vermis; 2) fibers with morphological characteristics of simple mossy fiber terminals or climbing fiber glomeruli that were located within the granule cell layer of the vermis underlying the foci of CRF/ENK-IR climbing fibers; 3) mossy fiber terminals within the flocculus; 4) neuronal perikarya in subnucleus A and C of the medial accessory olive and in the dorsal cap of Kooy, nuclei known to project as climbing fibers; and 5) cell bodies in nucleus prepositus, the subtrigeminal reticular nucleus, and the reticular formation, likely origins of CRF/ENK colocalized mossy fibers. The demonstration that single climbing and mossy fibers contain two peptides extends previous studies that have described chemical heterogeneity within cerebellar afferent pathways. Furthermore, these results support suggestions that this heterogeneity may provide a substrate for differential regulation of signal transduction by chemically coded cerebellar afferents.