Ultrastructural studies on peptides in the dorsal horn of the rat spinal cord--III. Effects of peripheral axotomy with special reference to galanin.

Abstract

In this study co-localization of galanin- with calcitonin gene-related peptide (CGRP)-like immunoreactivity was examined in dorsal root ganglion neurons 14 days after sciatic nerve cut using a laser scanning confocal microscope. CGRP- and galanin-like immunoreactivities were also analysed in the dorsal horn of the spinal cord of these animals with immunofluorescence microscopy. The ultrastructural changes in galanin-immunoreactive, presumably primary afferent terminals in the superficial dorsal horn, were studied as well as the relationship between galanin-, substance P- and CGRP-like immunoreactivities in primary afferent terminals. Local galanin-positive neurons in lamina II were also analysed after peripheral axotomy. Under the confocal microscope, CGRP-like immunoreactivity was located in the perinuclear region, probably the Golgi complex, and in dot-like structures, probably representing large dense-core vesicles, in normal dorsal root ganglion neurons. However, after peripheral axotomy CGRP was mainly detected in dot-like structures. Only a slight decrease in percentage of CGRP neurons in dorsal root ganglion was seen after axotomy, and about 84% of the galanin-positive neurons contained CGRP. The field of galanin-positive nerve fibres in the superficial lumbar (L)4 and L5 dorsal horn expanded and the intensity of staining for CGRP was reduced in these regions 14 days after sciatic nerve cut. Using pre-embedding immunoelectron microscopy, several morphological changes were observed in galanin-positive terminals in laminae I and II ipsilateral to the lesion. Most importantly, the most frequently occurring type of galanin-positive terminals (type 1) showed distinct changes with a granular matrix, many immunoreactive, peripherally located large dense-core vesicles, empty large vesicles and synaptic vesicles which were displaced from the presynaptic zone. Other galanin-positive terminals underwent even more pronounced morphological changes, including extensive vesiculolysis, also of large dense-core vesicles, filamentous degeneration or formation of axonal labyrinths. An increased number of galanin-positive nerve terminals was observed in lamina III of the ipsilateral dorsal horn after axotomy. They did not form glomeruli and contained few large dense-core vesicles. Post-embedding immunocytochemistry combined with quantitative analysis revealed that significant changes occurred in a proportion of terminals also with regard to peptide content in large dense-core vesicles after axotomy. Thus, the percentage of galanin-positive large dense-core vesicles increased in several cases and that of substance P- and CGRP-immunoreactive ones decreased.(ABSTRACT TRUNCATED AT 400 WORDS)