Three-dimensional spatial relationship of neuropeptides and receptors in the rat dorsal vagal complex

Retrograde tracing, multi-label fluorescence immunohistochemistry, confocal microscopy and three-dimensional (3-D) reconstruction techniques were combined to examine the spatial relationship of immunoreactive nerve terminals containing either calcitonin gene-related polypeptide (CGRP) or substance P (SP) to identified gastric efferent neurons in the dorsal motor nucleus of the vagus (DMV) in the brainstem of the rat. The availability of an antibody to the receptor for SP (NK-1r) permitted observation of the association between peptide and receptor. Although both SP-IR and CGRP-IR nerve fibres came in close spatial proximity to identified gastric efferent neurons, few discrete contacts between these fibres and the neuronal membrane were observed. In addition, NK-1r-IR was localized to the somatic and dendritic membranes of a subpopulation of neurons within the DMV, with the majority of receptor labelling not in close spatial proximity to SP-IR nerve fibres. The methodology described in this study permitted the simultaneous observation of the spatial relationship between neuropeptide and an identified neuron (and the corresponding receptor in the case of SP) in 3-D, which is something that cannot be achieved using conventional microscopic techniques