lucifer-yellow and thionine

Anatomical study of neurons projecting to the retrocerebral complex of the adult blow fly, Protophormia terraenovae, was done by NiCl2 filling and immunocytochemistry. Retrograde filling through the cardiac-recurrent nerve labeled three groups of neurons in the brain/subesophageal ganglion: (1) paramedial clusters of the pars intercerebralis, (2) neurons in each pars lateralis, and (3) neurons in the subesophageal ganglion. The pars intercerebralis neurons send prominent axons into the median bundle and exit from the brain via the contralateral nervus corporis cardiaci. Based on the projection pattern, two types of the pars lateralis neurons can be distinguished: the most lateral pairs of neurons contralaterally extend through the posterior lateral tract and the remainder ipsilaterally extend through the posterior lateral tract. The neurons in the subesophageal ganglion run through the contralateral nervus corporis cardiaci. The dendritic arborization of the pars intercerebralis and pars lateralis neurons is restricted to the superior protocerebral neuropil and to the anterior neuropil of the subesophageal ganglion where the neurons in the subesophageal ganglion also project. Retrograde filling from the corpus allatum indicated that the pars lateralis neurons and a few pars intercerebralis neurons project to the corpus allatum, but that the neurons in the subesophageal ganglion do not. Orthograde filling from the pars intercerebralis and staining by paraldehyde-thionin/paraldehyde-fuchsin indicated that the pars intercerebralis neurons project primarily to the corpus cardiacum/hypocerebral ganglion complex. Immunostaining with a polyclonal antiserum against diapause hormone, a member of the FXPRLamide family, suggests that some of the subesophageal ganglion neurons contain FXPRLamide-like peptides.

Topics: Age Factors; Animals; Antibodies; Coloring Agents; Corpora Allata; Diptera; Fluorescent Dyes; Ganglia, Invertebrate; Insect Hormones; Isoquinolines; Neural Pathways; Neurons; Neuropeptides; Neurosecretory Systems; Nickel; Paraldehyde; Phenothiazines; Presynaptic Terminals; Rosaniline Dyes

Local collateral projections from the medial superior olivary nucleus in the gerbil auditory brainstem were examined to study the possible communication of this nucleus with periolivary cell groups. The projections were investigated using intracellular and extracellular labeling with Biocytin in the medial superior olive (MSO) in brainstem tissue slices. Collateral axons were found to branch from the main axons of the central cells of the MSO as the latter passed through a dorsally neighboring periolivary nucleus, the superior paraolivary nucleus (SPN), toward the ipsilateral inferior colliculus (IC), their traditionally accepted target. Bouton-like endings and en passant varicosities of these collaterals appeared to contact the somata and proximal dendrites of cells within the SPN. Furthermore, close observation revealed that these collaterals terminate on at least two types of SPN cells. Intracellular labeling of the collateral axons of the MSO neurons combined with retrograde prelabeling of their target cells, however, revealed that the collaterals selectively contact the cells of the SPN that project to the ipsilateral IC. A link between the MSO and SPN has not been reported previously. This connection is of interest since SPN cells themselves project either to the cochlear nuclei (CN) or the IC. The MSO-SPN projection identified here raises the possibility that the latter may serve as an ancillary channel to convey MSO information to the IC.

Topics: Animals; Auditory Pathways; Axons; Coloring Agents; Female; Fluorescent Dyes; Gerbillinae; Isoquinolines; Lysine; Male; Olivary Nucleus; Organ Culture Techniques; Phenothiazines; Sound Localization; Stilbamidines

A technique is described which permits rapid processing of neural tissue for light microscopic analysis of sections of 1-40 microns thickness. This technique was developed as an alternative to paraffin embedding. When compared to paraffin, polyethylene glycol (PEG) offers the following advantages: 10-15 degrees C lower embedding temperatures, net tissue shrinkage of less than 5% vs approximately 50% in paraffin, and approximately one-half the embedding time. Tissue orientation during embedding and sectioning is particularly easy to control, e.g. 500 microns brain slices can be routinely flat-embedded and sectioned at 5 microns to form excellent ribbons. Since PEG is water-soluble, tissue may be dehydrated with a series of aqueous PEG solutions; the embedding matrix is easily removed by washing with a variety of aqueous buffers. These procedures allow subsequent electron microscopic analysis of material with generally well preserved ultrastructure. However, PEG is hygroscopic, thus tissue blocks become soft and difficult to section in high (greater than 90%) relative humidity. PEG was found to be compatible with intracellular staining with Lucifer yellow, horseradish peroxidase enzyme histochemistry, aqueous histofluorescence and immunocytochemical demonstration of neuronal peptides and glial fibrillary acidic protein.

Topics: Animals; Enzymes; Fluorescence; Histocytochemistry; Histological Techniques; Horseradish Peroxidase; Immunologic Techniques; Injections; Intracellular Membranes; Isoquinolines; Microscopy, Electron; Phenothiazines; Polyethylene Glycols; Staining and Labeling