Immunohistochemistry as a tool for topographical semi-quantification of neurotransmitters in the brain.
Immunohistochemistry is a powerful tool to detect neurotransmitter (NT) presence in different brain structures with a high spatial resolution. However, it is only scarcely used in quantitative approach due to lack of reproducibility and sensitivity. We developed a protocol of NT detection based on immunohistochemistry and image analysis to show that this approach could also be useful to evaluate NT content variations. We focused our study on the GABAergic system in the cerebellum and measured different accurate parameters, namely the optical density (O.D.), the stained area and the number of immunoreactive cells in each cerebellar cell layer. In order to modify the GABA content, we used gamma-vinyl-GABA (GVG), an inhibitor of GABA-transaminase, known to dramatically increase GABA concentration in the central nervous system (CNS) and especially in the cerebellum. We observed a significant increase in the three parameters measured in the molecular and the granular layers of the cerebellum after treatment with GVG, reflecting the well-established increase in GABA content after such a treatment. Therefore, our technical approach allows not only a precise determination of the effects in particular cell layers but also a semi-quantification of GABA content variations. This technique could be suitable for monitoring NT variations following any treatment.
Mausset-Bonnefont
Anne Laure
AL
INSERM U336, Développement, Plasticité et Vieillissement du Système Nerveux, Université Montpellier II, Place Eugène Bataillon, B.P. 106, 34095 Montpellier Cedex 5, France. bonnefont@ch1.fgg.eur.nl
de Sèze
René
R
Privat
Alain
A
D000612:4-Aminobutyrate Transaminase / Q000037:antagonists & inhibitors; D000818:Animals; D001923:Brain Chemistry / Q000187:drug effects / Q000502:physiology*; D001931:Brain Mapping / Q000379:methods*; D002531:Cerebellum / Q000187:drug effects / Q000201:enzymology; D004396:Coloring Agents; D004305:Dose-Response Relationship, Drug; D004791:Enzyme Inhibitors / Q000494:pharmacology; D007091:Image Processing, Computer-Assisted; D007150:Immunohistochemistry / Q000379:methods*; D007202:Indicators and Reagents; D008297:Male; D018377:Neurotransmitter Agents / Q000032:analysis / Q000502:physiology*; D051381:Rats; D020888:Vigabatrin / Q000494:pharmacology; D005680:gamma-Aminobutyric Acid / Q000378:metabolism
D004396:Coloring Agents; D004791:Enzyme Inhibitors; D007202:Indicators and Reagents; D018377:Neurotransmitter Agents; D005680:gamma-Aminobutyric Acid; D000612:4-Aminobutyrate Transaminase; D020888:Vigabatrin