Abstract
The plant lectin Phaseolus vulgaris leukoagglutinin (PHA-L) has unique properties that have made it particularly useful for tracing the axonal projections of neurons (Gerfen and Sawchenko, 1984, 1985). When PHA-L is injected by iontophoresis into the brain and subsequently localized by standard im-munohistochemical techniques, neurons at the injection site are labeled in their entirety, revealing the morphology of dendritic arborizations and both local and distant axonal projections. PHA-L appears to label only those neurons that incorporate tracer at the injection site; i.e., there is negligible uptake of the tracer by fibers of passage and negligible retrograde axonal transport of the tracer. In addition, neurons and their axonal projections are labeled with the PHA-L method in a manner that rivals the best Golgi impregnation techniques. With this method the amplification of the signal is generated by the immunohistochemical procedure and both the fluorescent and peroxidase-coupled diaminobenzidine labels provide an accurate visualization of cellular and axonal morphology. Furthermore, because PHA-L is localized with standard immunohistochemical techniques, it may be combined with other neuroanatomical and neurohistochemical methods. Such combined methods generate information not only concerning the interrelations between separate populations of neurons but also about the neurochemical characteristics of such neurons.
This is a preview of subscription content, log in via an institution to check access.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
Preview
Unable to display preview. Download preview PDF.
References
Chu, G., Hayakawa, H., and Berg, P., 1987, Electroporation for the efficient transfection of mammalian cells with DNA, Nucleic Acids Res. 15:1311–1318.
Cowan, W. M., Gottleib, D. L., Hendrickson, A. E., Price, J. L., and Woolsey, T. A., 1972, The autoradiographic demonstration of axonal connections in the central nervous system, Brain Res. 37:21–51.
Cunningham, E. T., Jr., and Sawchenko, P. E., 1987, Anatomical specificity of noradrenergic inputs to the paraventricular and supraoptic nuclei of the rat hypothalamus, J. Comp. Neurol 274:60–76.
Gerfen, C. R., 1984, The neostriatal mosaic: Compartmentalization of corticostriatal input and striatonigral output systems, Nature 311:461–464.
Gerfen, C. R., 1985, The neostriatal mosaic: I. Compartmental organization of projections from the striatum to the substantia nigra in the rat, J. Comp. Neurol. 236:454–476.
Gerfen, C. R., and Sawchenko, P. E., 1984, An anterograde neuroanatomical tracing method that shows the detailed morphology of neurons, their axons and terminals: Immunohisto-chemical localization of an axonally transported plant lectin, Phaseolus vulgaris-leucoagglu-tinin (PHA-L), Brain Res. 290:219–238.
Gerfen, C. R., and Sawchenko, P. E., 1985, A method for anterograde axonal tracing of chemically specified circuits in the central nervous system: Combined Phaseolus vulgaris-leucoag-glutinin (PHA-L) tract tracing and immunohistochemistry, Brain Res. 343:144–150.
Gerfen, C. R., Baimbridge, K. G., and Miller, J. J., 1985, The neostriatal mosaic: Compartmental distribution of calcium binding protein and parvalbumin in the basal ganglia of the rat and monkey, Proc. Natl. Acad. Sci. U.S.A. 82:8780–8784.
Gerfen, C. R., Herkenham, M., and Thibault, J., 1987, The neostriatal mosaic. II. Compartmental organization of dopaminergic and non-dopaminergic mesostriatal systems, J. Neu-rosci. 7:3915–3934.
Gonatas, N. K., Haper, C., Mizutani, T., and Gonatas, J. O., 1979, Superior sensitivity of conjugates of horseradish peroxidase with wheat germ agglutinin for studies of retrograde transport, J. Histochem. Cytochem. 27:728–734.
Heimer, L., Zaborsky, L., Zahm, D. S., and Alheid, G. F., 1987, The ventral striatopallidothal-amic projection: I. The striatopallidal link originating in the striatal parts of the olfactory tubercle, J. Comp. Neurol. 255:571–591.
Herkenham, M., and Pert, C. B., 1982, Light microscopic localization of brain opiate receptors: A general autoradiographic method which preserves tissue quality, J. Neurosci. 2:1129–1149.
Hökfelt, T., Skirboll, L., Rehfeld, J. F., Goldstein, M., Markey, K., and Dann, O., 1980, A subpopulation of mesencephalic dopamine neurons projecting to limbic areas contains a cholescystokinin-like peptide: evidence from immunohistochemistry combined with retrograde tracing, Neuroscience 5:2093–2124.
Hsu, S. M. L., Raine, L., and Fanger, H., 1981, The use of avidin—biotin peroxidase complex (ABC) in immunoperoxidase techniques: A comparison between ABC and unlabeled antibody (PAP) procedures, J. Histochem. Cytochem. 29:577–580.
Itoh, K., Konishi, A., Nomura, S., Mizuno, N., Nakamura, Y., and Sugimoto, I., 1979, Application of coupled oxidation reaction to electron microscopic demonstration of horseradish peroxidase: Cobalt-glucose oxidase, Brain Res. 175:341–346.
McLean, I. W., and Nakane, P. K., 1974, Periodate-lysine-paraformaldehyde fixative. A new fixative for immunoelectron microscopy, J. Histochem. Cytochem. 22:1077–1083.
Mesulam, M. M., 1978, Tetramethyl benzidine for horseradish peroxidase neurochemistry: A non-carcinogenic blue reaction product with superior sensitivity for visualizing neural afferents and efferents, J. Histochem. Cytochem. 26:106–117.
Sawchenko, P. E., and Swanson, L. W., 1981, A method for tracing biochemically defined pathways in the central nervous system using combined fluorescence retrograde transport and immunohistochemical techniques, Brain Res. 210:31–51.
Somogyi, P., and Takagi, H., 1982, A note on the use of picric acid-paraformaldehyde-glutar-aldehyde fixative for light and electron microscopic immunocytochemistry, Neuroscience 7:1779–1783.
Staines, W. A., Kimura, H., Fibiger, H. C., and McGeer, E. G., 1980, Peroxidase-labeled lectin as a neuroanatomical tracer: Evaluation in a CNS pathway, Brain Res. 197:485–490.
van der Kooy, D., Coscina, D. V., and Hattori, T., 1981, Is there a non-dopaminergic nigro-striatal pathway? Neuroscience 6:345–357.
Wouterloud, F. G., and Groenewegen, H. J., 1985, Neuroanatomical tracing by use of Phaseolus vulgaris-leucoagglutinin (PHA-L): Electron microscopy of PHA-L-filled neuronal somata, dendrites, and axon terminals, Brain Res. 326:188–191.
Zaborszky, L., and Cullinan, W. E., 1989, Hypothalamic axons terminate on forebrain cholinergic neurons: An ultrastructural double-labeling study using PHA-L tracing and ChAT immunocytochemistry, Brain Res. 479:177–184.
Zahm, D. S., and Heimer, L., 1985, Synaptic contacts of ventral striatal cells in the olfactory tubercle of the rat: Correlated light and electron microscopy of anterogradely transported Phaseolus vulgaris-leucoagglutinin, Neurosci. Lett. 60:169–175.
Author information
Authors and Affiliations
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 1989 Springer Science+Business Media New York
About this chapter
Cite this chapter
Gerfen, C.R., Sawchenko, P.E., Carlsen, J. (1989). The PHA-L Anterograde Axonal Tracing Method. In: Heimer, L., Záborszky, L. (eds) Neuroanatomical Tract-Tracing Methods 2. Springer, Boston, MA. https://doi.org/10.1007/978-1-4757-2055-6_3
Download citation
DOI: https://doi.org/10.1007/978-1-4757-2055-6_3
Publisher Name: Springer, Boston, MA
Print ISBN: 978-1-4757-2057-0
Online ISBN: 978-1-4757-2055-6
eBook Packages: Springer Book Archive