In vivo knockdown of GAD67 in the amygdala disrupts fear extinction and the anxiolytic-like effect of diazepam in mice

• Type of Material: Article
• Authors: S.A. Heldt, University of Tennessee; L. Mou, Emory University; Kerry Ressler, Emory University
• Language: English
• Date: 2012-11-01
• Publisher: NATURE PUBLISHING GROUP
• Publication Version: Final Published Version
• Copyright Statement: © 2012 Macmillan Publishers Limited All rights reserved.
• License: Creative Commons Attribution-NonCommerical-NoDerivs 3.0 Unported
• Final Published Version (URL): http://dx.doi.org/10.1038/tp.2012.101
• Title of Journal or Parent Work: TRANSLATIONAL PSYCHIATRY
• Volume: 2
• Issue: 11
• Start Page: e181
• End Page: e181
• Grant/Funding Information: This study was supported by NIH (DA019624, P30 NS055077, F32 MH073389), NARSAD, the Burroughs Wellcome Fund, and Center for Behavioral Neuroscience STC Center: NSF Agmt #IBN-9876754, National Primate Research Center base Grant #RR-00165, Animal Resource Program at NIH.
• Keywords: ACTIVITY-DEPENDENT EXPRESSION; 2 DISTINCT MECHANISMS; ELEVATED PLUS-MAZE; BENZODIAZEPINE-RECEPTOR-BINDING; BASOLATERAL AMYGDALA; GABA; GABA PLASMA-LEVELS; Science & Technology; Psychiatry; amygdala; LONG-TERM POTENTIATION; GLUTAMATE-DECARBOXYLASE GAD; GAD67; fear; Life Sciences & Biomedicine; extinction; GAMMA-AMINOBUTYRIC-ACID; PARVALBUMIN-CONTAINING INTERNEURONS; lentivirus
• Research Categories: Psychology, Behavioral; Biology, Neuroscience

Abstract

In mammals, γ-aminobutyric acid (GABA) transmission in the amygdala is particularly important for controlling levels of fear and anxiety. Most GABA synthesis in the brain is catalyzed in inhibitory neurons from L-glutamic acid by the enzyme glutamic acid decarboxylase 67 (GAD67). In the current study, we sought to examine the acquisition and extinction of conditioned fear in mice with knocked down expression of the GABA synthesizing enzyme GAD67 in the amygdala using a lentiviral-based (LV) RNA interference strategy to locally induce loss-of-function. In vitro experiments revealed that our LV-siRNA-GAD67 construct diminished the expression of GAD67 as determined with western blot and fluorescent immunocytochemical analyses. In vivo experiments, in which male C57BL6J mice received bilateral amygdala microinjections, revealed that LV-siRNA-GAD67 injections produce significant inhibition of endogenous GAD67 when compared with control injections. In contrast, no significant changes in GAD65 expression were detected in the amygdala, validating the specificity of LV knockdown. Behavioral experiments showed that LV knockdown of GAD67 results in a deficit in the extinction, but not the acquisition or retention, of fear as measured by conditioned freezing. GAD67 knockdown did not affect baseline locomotion or basal measures of anxiety as measured in open field apparatus. However, diminished GAD67 in the amygdala blunted the anxiolytic-like effect of diazepam (1.5 mg kg -1) as measured in the elevated plus maze. Together, these studies suggest that of GABAergic transmission in amygdala mediates the inhibition of conditioned fear and the anxiolytic-like effect of diazepam in adult mice.