Impaired extinction of learned fear in rats selectively bred for high anxiety--evidence of altered neuronal processing in prefrontal-amygdala pathways

Abstract

The impaired extinction of acquired fear is a core symptom of anxiety disorders, such as post-traumatic stress disorder, phobias or panic disorder, and is known to be particularly resistant to existing pharmacotherapy. We provide here evidence that a similar relationship between trait anxiety and resistance to extinction of fear memory can be mimicked in a psychopathologic animal model. Wistar rat lines selectively bred for high (HAB) or low (LAB) anxiety-related behaviour were tested in a classical cued fear conditioning task utilizing freezing responses as a measure of fear. Fear acquisition was similar in both lines. In the extinction trial, however, HAB rats showed a marked deficit in the attenuation of freezing responses to repeated auditory conditioned stimulus presentations as compared with LAB rats, which exhibited rapid extinction. To gain information concerning the putatively altered neuronal processing associated with the differential behavioural response between HAB and LAB rats, c-Fos expression was investigated in the main prefrontal-amygdala pathways important for cued fear extinction. HAB compared to LAB rats showed an attenuated c-Fos response to repeated conditioned stimulus presentations in infralimbic and cingulate cortices, as well as in the lateral amygdala, but facilitated the c-Fos response in the medial part of the central amygdala. In conclusion, the present results support the notion that impaired extinction in high anxiety rats is accompanied by an aberrant activation profile in extinction-relevant prefrontal-amygdala circuits. Thus, HAB rats may represent a clinically relevant model to study the mechanisms and potential targets to accelerate delayed extinction processes in subjects with enhanced trait anxiety.

Fig. 2

Schematic diagrams adapted from the atlas of Paxinos & Watson (1998) showing subregions of the amygdala and mPFC in which a differential c-Fos response after repeated conditioned auditory stimulus presentation in the extinction trial was found in HAB vs. LAB rats [C(+)CS groups]. Acetylcholinesterase staining was performed to aid identification of CeA subdivisions. Grey boxes indicate reduced c-Fos expression in HAB vs. LAB rats and black box indicates increased c-Fos expression in HAB vs. LAB rats. CeC, capsular part of the central nucleus of the amygdala; CeL, lateral part of the central nucleus of the amygdala; opt, optic nerve.

Fig. 1

(A) Experiment 1 [C(+)CS groups only]. Freezing responses during the fear conditioning trial did not differ between HAB and LAB rats. After repeated conditioned auditory stimulus presentations during the extinction trial, LAB rats showed a progressive decline in freezing behaviour, whereas freezing in HAB rats remained elevated until the 29th CS presentation. Note that no freezing was observed in all groups of cued-conditioned rats upon exposure to context alone (trial 0). HAB, n= 9; LAB, n= 10. During extinction recall, HAB rats showed elevated levels of freezing compared with LAB rats. (B) Experiment 2 (c-Fos study). The freezing responses in C(+)CS HAB and LAB rats were similar to those observed in Experiment 1. Non-conditioned animals [C(−)CS groups] did not freeze during the conditioning. Animals of both control groups [C(−)CS and C(+)no-CS groups] did not freeze during the extinction trial. Trials in conditioning (left image), number of CS/US pairings; trials in extinction (right image), number of CS presentation. *P<0.05, **P<0.01, ***P<0.001, HAB C(+)CS vs. LAB C(+)CS group; ^#P<0.01, HAB C(+)CS vs. HAB C(−)CS and HAB C(+)no-CS groups; ^•P<0.01, LAB C(+) vs. LAB C(−) group; ^ψP>0.01, HAB C(+) vs. HAB C(−) group; ⁺P>0.01, LAB C(+)CS vs. LAB C(−)CS group; ^†P>0.01, LAB C(+)CS vs. LAB C(+)no-CS group. Extinction groups: C(+)CS HAB (n= 7) and LAB (n= 9); Control groups: C(+)no-CS HAB and LAB (n= 5, respectively), and C(−)CS HAB and LAB (n= 3, respectively).

Fig. 3

Bright-field photomicrographs of representative sections showing significant differences in c-Fos expression within the medial part of the CeM and LA, and the IL in HAB and LAB rats after the extinction trial [C(+)CS groups only]. High-power photomicrograph showing examples of c-Fos-positive cells in the CeM. The prelimbic cortex (PL) is shown as an example of similar c-Fos responses in both lines. The selected areas are depicted in the schematic drawings (adapted from Paxinos & Watson, 1998). Scale bar, 200 μm.