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In vitro biocytin injection into perinatal mouse brain: a method for tract tracing in developing tissue
S L Chang1, J J LoTurco, L K Nisenbaum
1Department of Physiology and Neurobiology, University of Connecticut, 3107 Horsebarn Hill Rd., U-156, Storrs, CT 06269, USA.
Journal of Neuroscience Methods
|April 20, 2000
Summary
This study presents an improved biocytin injection method for axonal tracing in developing mouse brains. Explanted brain hemispheres allow for easier access and faster labeling of neural projections in perinatal animals.
Area of Science:
- Neuroscience
- Developmental Biology
- Histology
Background:
- Biocytin injection is effective for axonal projection labeling.
- In vivo application in fetal/postnatal animals faces challenges like limited access and long survival times.
Purpose of the Study:
- To adapt extracellular biocytin injection for explanted brain hemispheres in developing mice.
- To optimize conditions for efficient axonal labeling in perinatal brains.
Main Methods:
- Explanted brain hemispheres (E16-P9) were used.
- Visually guided biocytin (2%) injection into the prelimbic cortex.
- Iontophoresis with patch clamp electrodes and incubation in artificial cerebrospinal fluid (aCSF).
Main Results:
- Optimal labeling achieved with 10-min iontophoresis using 8-12 megaohm electrodes in E16-P9 animals.
- Incubation of 4+ hours sufficient for labeling fine collaterals and long-distance axon bundles (>3 mm).
Conclusions:
- In vitro biocytin injection into explanted hemispheres is a rapid and accessible method for tract tracing in developing brains.
- This technique overcomes limitations of in vivo methods for perinatal studies.