Fluorescence mapping of afferent topography in three dimensions (original) (raw)

Abstract

Neural circuits are organized into complex topographic maps. Although several neuroanatomical and genetic tools are available for studying circuit architecture, a limited number of methods exist for reliably revealing the global patterning of multiple topographic projections. Here we used wheat germ agglutinin (WGA) conjugated to Alexa 555 and 488 for dual color fluorescent mapping of parasagittal spinocerebellar topography in three dimensions. Using tissue section and wholemount imaging we show that WGA-Alexa tracers have three main characteristics that make them ideal tools for analyses of neural projection topography. First, the intense brightness of Alexa fluorophores allows multi-color imaging of patterned afferent projections in wholemount preparations. Second, WGA-Alexa tracers robustly label the entire trajectory of developing and adult projections. Third, long tracts such as the adult spinocerebellar tract can be traced in less than 6 h. Moreover, using WGA-Alexa tracers we resolved a level of complexity in the compartmentalized topography of the spinocerebellar projection map that has never before been appreciated. In summary, we introduce versatile tracers for rapidly labeling multiple topographic projections in three dimensions and uncover wiring complexities in the spinocerebellar map.

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Acknowledgments

This work was supported by new investigator start-up funds from Albert Einstein College of Medicine of Yeshiva University to RVS.

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Authors and Affiliations

1. Dominick P. Purpura Department of Neuroscience, Albert Einstein College of Medicine, Yeshiva University, Bronx, NY, 10461, USA
   Stacey L. Reeber, Samrawit A. Gebre & Roy V. Sillitoe

Authors

1. Stacey L. Reeber
2. Samrawit A. Gebre
3. Roy V. Sillitoe

Corresponding author

Correspondence toRoy V. Sillitoe.

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Supplementary Fig. 1 (download TIF )

Spinal cord injections of WGA-Alexa 555 or WGA-Alexa 488 consistently label bands of mossy fiber terminals. WGA-Alexa 555 and 488 are transported into afferent terminals in the AZ (a, b) and PZ (d, e). The insets in a and b show images of the injections sites after delivery of WGA-Alexa 555 and 488. c, f. The dotted lines in the sagittal schematics indicate the level from which the coronal sections were taken, and correspond to tissue sections shown in a, b, d, and e, respectively. g. Anterogradely transported WGA-Alexa 555 accumulates as punctate deposits in terminals. g’. VGLUT2 labels mossy fiber terminals in the cerebellum. g”. High power image of mossy fiber terminals shows that WGA-Alexa 555 co-labels with a subpopulation of VGLUT2 expressing mossy fibers. h, h’, h”. High magnification images of a mossy fiber glomerulus double labeled with VGLUT2 and WGA-Alexa Fluor 555. Scale bars: e, 500 μm (for a–d); g”, 100 μm (for g, g’); h”, 12.5 μm (for h, h’). Supplementary material 1 (TIFF 9418 kb)

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Reeber, S.L., Gebre, S.A. & Sillitoe, R.V. Fluorescence mapping of afferent topography in three dimensions.Brain Struct Funct 216, 159–169 (2011). https://doi.org/10.1007/s00429-011-0304-2

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• Received: 10 January 2011
• Accepted: 22 February 2011
• Published: 09 March 2011
• Issue date: September 2011
• DOI: https://doi.org/10.1007/s00429-011-0304-2

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