School of Medicine and Health Sciences Poster Presentations
Activation of Angiotensin type 2 Receptor (AT2R) Contributes to Fear Memory
Poster Number
291
Document Type
Poster
Status
Postdoc
Publication Date
Spring 2018
Abstract
Background: Previous clinical studies identify the renin-angiotensin system (RAS) as a potential therapeutic target in post-traumatic stress disorder (PTSD), however the mechanism(s) are unknown. We propose that brain angiotensin receptors modulate inhibitory and excitatory fear circuits critical to the consolidation and extinction of fear memory in PTSD.
Methods: Using a transgenic brain angiotensin type 2-receptor (AT2R) eGFP-BAC reporter mouse combined with pharmacological and behavioral approaches, the aim of the study was to examine the role of the brain AT2R in fear memory.
Results: We first assessed AT2R-eGFP+ immunoreactive cell types in the mouse brains, which were highly colocalized with the neuronal specific marker NeuN. There were no detectable levels of co-localization in astrocytes (GFAP as a marker) or microglia (Iba-1 as a marker). We then quantified the number of AT2R-eGFP+ expressing neurons in the major subnuclei of the amygdala, an important structure in the consolidation and extinction of fear memory. AT2R-eGFP+ neurons were predominately observed in the medial amygdala (MeA) (203.8 ± 39.4 cells/mm2 ) and the medial division of central amygdala (CeM) (221.2 ± 15.3 cells/mm2 ), while the basolateral amygdala (BLA) (11.7 ± 1.8 cells/mm2 ) contained very few AT2R-eGFP+ neurons. Moreover, within the CeM, 96% of the AT2R-eGFP+ neurons expressed the GABAergic marker GAD1, while approximately 40% GABAergic cells expressed interneuron markers calbindin and nNOS. To examine the behavioral role of AT2R in fear memory, we used classic Pavlovian fear conditioning, pairing auditory cues with footshocks. Following the acquisition of fear, AT2R mRNA expression was significantly elevated (t(22) = 2.5; p<0.05) within the CeA but not following extinction learning. In separate studies, a bilateral intra-CeA injection of the AT2R agonist C21 (0.06ug/ul) was administered prior to fear acquisition testing and freezing behavior was quantified. Compared to vehicle controls, mice receiving C21 into the CeA displayed enhanced extinction by a decreased percentage of freezing (61.7% ± 5.4 vehicle v.s. 42.1% ± 5.7 C21 group, p<0.05, n=10-12) during repeated conditioned stimuli (30 second auditory tones). These behavioral results were independent of general anxiety-like measures as C21 did not affect locomotor activity or center entries in the open field test.
Conclusions: Together, these data provide anatomical, functional and behavioral evidence for activation brain AT2R in the consolidation and extinction of conditioned fear. Further studies are required to determine the neurobiological mechanism(s), which may involve changes in cerebral vascular blood flow and/or modulation of neuronal excitability and plasticity.
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Open Access
1
Activation of Angiotensin type 2 Receptor (AT2R) Contributes to Fear Memory
Background: Previous clinical studies identify the renin-angiotensin system (RAS) as a potential therapeutic target in post-traumatic stress disorder (PTSD), however the mechanism(s) are unknown. We propose that brain angiotensin receptors modulate inhibitory and excitatory fear circuits critical to the consolidation and extinction of fear memory in PTSD.
Methods: Using a transgenic brain angiotensin type 2-receptor (AT2R) eGFP-BAC reporter mouse combined with pharmacological and behavioral approaches, the aim of the study was to examine the role of the brain AT2R in fear memory.
Results: We first assessed AT2R-eGFP+ immunoreactive cell types in the mouse brains, which were highly colocalized with the neuronal specific marker NeuN. There were no detectable levels of co-localization in astrocytes (GFAP as a marker) or microglia (Iba-1 as a marker). We then quantified the number of AT2R-eGFP+ expressing neurons in the major subnuclei of the amygdala, an important structure in the consolidation and extinction of fear memory. AT2R-eGFP+ neurons were predominately observed in the medial amygdala (MeA) (203.8 ± 39.4 cells/mm2 ) and the medial division of central amygdala (CeM) (221.2 ± 15.3 cells/mm2 ), while the basolateral amygdala (BLA) (11.7 ± 1.8 cells/mm2 ) contained very few AT2R-eGFP+ neurons. Moreover, within the CeM, 96% of the AT2R-eGFP+ neurons expressed the GABAergic marker GAD1, while approximately 40% GABAergic cells expressed interneuron markers calbindin and nNOS. To examine the behavioral role of AT2R in fear memory, we used classic Pavlovian fear conditioning, pairing auditory cues with footshocks. Following the acquisition of fear, AT2R mRNA expression was significantly elevated (t(22) = 2.5; p<0.05) within the CeA but not following extinction learning. In separate studies, a bilateral intra-CeA injection of the AT2R agonist C21 (0.06ug/ul) was administered prior to fear acquisition testing and freezing behavior was quantified. Compared to vehicle controls, mice receiving C21 into the CeA displayed enhanced extinction by a decreased percentage of freezing (61.7% ± 5.4 vehicle v.s. 42.1% ± 5.7 C21 group, p<0.05, n=10-12) during repeated conditioned stimuli (30 second auditory tones). These behavioral results were independent of general anxiety-like measures as C21 did not affect locomotor activity or center entries in the open field test.
Conclusions: Together, these data provide anatomical, functional and behavioral evidence for activation brain AT2R in the consolidation and extinction of conditioned fear. Further studies are required to determine the neurobiological mechanism(s), which may involve changes in cerebral vascular blood flow and/or modulation of neuronal excitability and plasticity.