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Adeno‐Associated Virus Labeling of Catecholaminergic neurons in Nucleus of Solitary Tract (NTS) of Rats
Author(s) -
Bathina Chandra,
Mifflin Steve
Publication year - 2012
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.26.1_supplement.706.7
Subject(s) - green fluorescent protein , tyrosine hydroxylase , catecholaminergic cell groups , catecholaminergic , immunohistochemistry , biology , neuron , solitary tract , forebrain , microbiology and biotechnology , nucleus , axoplasmic transport , neuroscience , central nervous system , catecholamine , immunology , gene , genetics
A2 catecholaminergic neurons located within caudal and sub‐postremal NTS play a critical role in modulating responses to physiological stress and project to multiple brain stem and forebrain hypothalamic regions. A2 neurons by definition express tyrosine hydroxylase, the rate‐limiting enzyme in catecholamine synthesis. In vitro analyses of A2 neurons in the rat have been limited as identification of A2 neurons requires labeling of neuron and post‐mortem immunohistochemistry. As a result, one does not know the phenotype of the neuron at the time of study. To overcome this challenge we have used commercially available adeno ‐ associated virus (AAV) vector mediated delivery of green fluorescent protein (GFP) labeled TH promoter (AAV‐GFP‐TH), which will incorporate into the TH genome and express GFP with the TH expression. To verify that GFP expression was restricted to catecholaminergic neurons, conventional immunohistochemistry for TH was performed on sections from AAV‐GFP‐TH injected animals. We found that, 78.4 ± 3.07% TH immunoreactive neurons to be GFP labeled and 95 ± 2.40% (n=3 sections) of the GFP expressing neurons of caudal NTS to be TH‐immunoreactive, making our technique a reliable and accurate way to label A2 neurons prior to in vitro study. Brain slices from these rats can be used for direct visualization of TH‐immunoreactive neurons without any need for further processing.