Traumatic Stress Produces Distinct Activations of GABAergic and Glutamatergic Neurons in Amygdala

Abstract

Posttraumatic stress disorder (PTSD) is an anxiety disorder characterized by intrusive recollections of a severe traumatic event and hyperarousal following exposure to the event. Human and animal studies have shown that the change of amygdala activity after traumatic stress may contribute to occurrences of some symptoms or behaviors of the patients or animals with PTSD. However, it is still unknown how the neuronal activation of different sub-regions in amygdala changes during the development of PTSD. In the present study, we used single prolonged stress (SPS) procedure to obtain the animal model of PTSD, and found that 1 day after SPS, there were normal anxiety behavior and extinction of fear memory in rats which were accompanied by a reduced proportion of activated glutamatergic neurons and increased proportion of activated GABAergic neurons in basolateral amygdala (BLA). About 10 days after SPS, we observed enhanced anxiety and impaired extinction of fear memory with increased activated both glutamatergic and GABAergic neurons in BLA and increased activated GABAergic neurons in central amygdala (CeA). These results indicate that during early and late phase after traumatic stress, distinct patterns of activation of glutamatergic neurons and GABAergic neurons are displayed in amygdala, which may be implicated in the development of PTSD.

Keywords: basolateral amygdala (BLA); central amygdala (CeA); neuronal activations; posttraumatic stress disorder (PTSD); single prolonged stress (SPS).

FIGURE 1

Effect of single prolonged stress (SPS) on anxiety-like behaviors and fear extinction on day 1 or day 10 after stress. (A) Experimental procedures. We treated rats with SPS procedure. About 1 day or 10 days later, we examined the anxiety-like behaviors with open field and elevated plus maze, and examined fear extinction with contextual fear conditioning. (B,C) Time spent in the central area (B) and the total locomotor distance (C) in different experimental conditions in open field test. (D,E) The entries into open arms (D) and time spent in open arms (E) in different experimental conditions in elevated plus maze test. (F,G) Formation and extinction of fear conditioning (F) and memory retention of fear extinction (G) in different experimental conditions. n = 8–10 per experimental condition. Data are mean ± SEM². ^∗Different from all other groups, one-way ANOVA, p < 0.05.

FIGURE 2

Effect of single prolonged stress (SPS) on activation of glutamatergic neurons in BLA on day 1 or day 10 after stress. (A) Experimental procedures. We treated rats with SPS procedure, and performed perfusion and brain dissection 1 day or 10 days later. (B) Representative images showing green (Fos protein), red (CaMKII protein), and merged channels of double-label neurons in BLA in different experimental conditions. Scale bars represent 100 μm. (C,D) Effect of the experimental manipulations on Fos protein (C) and CaMKII expression (D) in BLA. (E) Percentage of glutamatergic neurons in activated cells of BLA in different experimental manipulations, as calculated by number of overlap (Fos + CaMKII protein-IR)/Fos-IR. (F) Percentage of activated cells in glutamatergic neurons of BLA in different experimental manipulations, as calculated by number of overlap (Fos + CaMKII protein-IR)/CaMKII-IR. n = 6–8 per group. Data are mean ± SEM. ^∗Different from no SPS group at each post-stress day. ^#Different from SPS-1 day group, one-way ANOVA, p < 0.05.

FIGURE 3

Effect of single prolonged stress (SPS) on activation of GABAergic neurons in BLA on day 1 or day 10 after stress. (A) Representative images showing green (Fos protein), red GAD67 protein), and merged channels of double-label neurons in BLA in different experimental conditions. Scale bars represent 100 μm. (B,C) Effect of the experimental manipulations on Fos protein (B) and GAD67 expression (C) in BLA. (D) Percentage of GABAergic neurons in activated cells of BLA in different experimental manipulations, as calculated by number of overlap (Fos + GAD67 protein-IR)/Fos-IR. (E) Percentage of activated cells in GABAergic neurons in different experimental manipulations, as calculated by number of overlap (Fos + GAD67 protein-IR)/GAD67-IR. n = 6–7 per group. Data are mean ± SEM. ^∗Different from no SPS group at each post-stress day. ^#Different from SPS-1 day group, one-way ANOVA, p < 0.05.

FIGURE 4

Effect of single prolonged stress (SPS) on activation of GABAergic neurons in CeA on day 1 or day 10 after stress. (A) Representative images showing green (Fos protein), red GAD67 protein), and merged channels of double-label neurons in CeA in different experimental conditions. Scale bars represent 100 μm. (B,C) Effect of the experimental manipulations on Fos protein (B) and GAD67 expression (C) in CeA. (D) Percentage of GABAergic neurons in activated cells of CeA in different experimental manipulations, as calculated by number of overlap (Fos + GAD67 protein-IR)/Fos-IR. (E) Percentage of activated cells in GABAergic neurons of CeA in different experimental manipulations, as calculated by number of overlap (Fos + GAD67 protein-IR)/GAD67-IR. n = 7–8 per group. Data are mean ± SEM. ^∗Different from no SPS group at each post-stress day. ^#Different from SPS-1 day group, one-way ANOVA, p < 0.05.