Fixation of transsynaptically transported WGA-HRP and fluorescent dyes used in combination

The transsynaptic transport of WGA-HRP is a powerful tool for the identific ation of last order interneurones in the spinal cord, but differentiation o f primarily labelled motoneurones from transsynaptically labelled interneur ones can be difficult. We therefore combined the transsynaptic labelling of interneurones with WGA-HRP and the labelling of motoneurones with fluoresc ent dyes (fluorogold, red beads, fast blue, DAPI) by injecting a mixture of the tracers into the nervi mediani of rats. The influence of different fix atives on the simultaneous preservation of WGA-HRP and fluorescence is eval uated. It is shown that, of several perfusion protocols, only perfusion wit h paraformaldehyde in addition to 1.4% lysine and 0.23% periodate allowed s imultaneous visualisation of transsynaptic WGA-HRP and fluorescent dyes. (C ) 2000 Elsevier Science B.V. All rights reserved.