Fig. 2. Colocalisation of Lucifer-Yellow-fluorescence and anti-GABA-immunofluorescence in a nonspiking local interneuron in the metathoracic ganglion, demonstrated by confocal microscopy (all images are from whole mounts of ganglia, viewed ventrally). (A) A single cell body (arrow) in the anterior lateral region of the ganglion shows intense fluorescence for Lucifer Yellow (in green pseudocolour in this and following figures; slight background is due to tissue autofluorescence). (B) The same cell body (arrow) shows intense fluorescence for Cy3 in the same optical section, indicating binding of the GABA antibody (Cy3/GABA in red pseudocolour). A number of other small cell bodies are also GABA-immunopositive. A and B each show a composite of two confocal planes separated by 3.1 µm. (C) A merging of the two images in A and B results in a single yellow cell body, indicating colocalisation of the two fluorophores in the interneuron. The surrounding large cell bodies belong to glutamatergic motor neurons (five are indicated with asterisks) that showed no immunoreactivity with the GABA antibody. (D) The morphology of the neuron as revealed by the Lucifer Yellow dye (branches reconstructed from 29 confocal planes, each separated by 5.4 µm; cell body from a single confocal plane 56 µm more ventral than the ventralmost plane used in the reconstruction of the branches). A single primary neurite (arrow) emerges from the cell body to give rise to a profusion of fine branches in the neuropil. The ganglion is viewed ventrally with anterior to the top. Scale bars, 100 µm.

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