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J Vis Exp. 2017 Jun 5;(124). doi: 10.3791/55893.

Quadruple Immunostaining of the Olfactory Bulb for Visualization of Olfactory Sensory Axon Molecular Identity Codes.

Author information

1
Laboratory of Chemical Pharmacology, Graduate School of Pharmaceutical Sciences, The University of Tokyo.
2
Laboratory of Chemical Pharmacology, Graduate School of Pharmaceutical Sciences, The University of Tokyo; Center for Information and Neural Networks, National Institute of Information and Communications Technology.
3
Laboratory of Chemical Pharmacology, Graduate School of Pharmaceutical Sciences, The University of Tokyo; PRESTO, Japan Science and Technology Agency (JST); haruki-t@mol.f.u-tokyo.ac.jp.

Abstract

The mouse olfactory system is often used to study mechanisms of neural circuit formation because of its simple anatomical structure. An Olfactory Sensory Neuron (OSN) is a bipolar cell with a single dendrite and a single unbranched axon. An OSN expresses only one Olfactory Receptor (OR) gene, OSNs expressing a given type of OR converge their axons to a few sets of invariant glomeruli in the Olfactory Bulb (OB). A remarkable feature of OSN projection is that the expressed ORs play instructive roles in axonal projection. ORs regulate the expression of multiple axon-sorting molecules and generate the combinatorial molecular code of axon-sorting molecules at the OSN axon termini. Thus, to understand the molecular mechanisms of OR-specific axon guidance mechanisms, it is vital to characterize their expression profiles at the OSN axon termini within the same glomerulus. The aim of this article was to introduce methods for collecting as many glomeruli as possible on a single OB section and for performing immunostaining using multiple antibodies. This would allow the comparison and analysis of the expression patterns of axon-sorting molecules without staining variation between OB sections.

PMID:
28605383
DOI:
10.3791/55893
[Indexed for MEDLINE]

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