Adjunct treatment with ketamine enhances the therapeutic effects of extinction learning after chronic unpredictable stress

Abstract

Post-traumatic stress disorder (PTSD) is a debilitating illness characterized by dysfunction in the medial prefrontal cortex (mPFC). Although both pharmacological and cognitive behavioral interventions have shown some promise at alleviating symptoms, high attrition and persistence of treatment-resistant symptoms pose significant challenges that remain unresolved. Specifically, prolonged exposure therapy, a gold standard intervention to treat PTSD, has high dropout rates resulting in many patients receiving less than a fully effective course of treatment. Administering pharmacological treatments together with behavioral psychotherapies like prolonged exposure may offer an important avenue for enhancing therapeutic efficacy sooner, thus reducing the duration of treatment and mitigating the impact of attrition. In this study, using extinction learning as a rat model of exposure therapy, we hypothesized that administering ketamine as an adjunct treatment together with extinction will enhance the efficacy of extinction in reversing stress-induced deficits in set shifting, a measure of cognitive flexibility. Results showed that combining a sub-effective dose of ketamine with a shortened, sub-effective extinction protocol fully reversed stress-induced cognitive set-shifting deficits in both male and female rats. These effects may be due to shared molecular mechanisms between extinction and ketamine, such as increased neuronal plasticity in common circuitry (e.g., hippocampus-mPFC), or increased BDNF signaling. This work suggests that fast-acting drugs, such as ketamine, can be effectively used in combination with behavioral interventions to reduce treatment duration and potentially mitigate the impact of attrition. Future work is needed to delineate other pharmacotherapies that may complement the effects of extinction via shared or independent mechanisms.

Keywords: Adjunct treatment; Chronic stress; Fear extinction; Ketamine; Set shifting.

Fig. 1

8-Tone extinction produces less retention compared to 16-tone extinction. (A) Fear conditioning curves showing % time freezing during tone presentation for 16-tone and 8-tone extinction groups. (B) Fear extinction curves during 16-tone and 8-tone extinction. Data include both males (6–10/group) and females (6–8/group). (C) Extinction retention is worse after 8-tone extinction compared to 16-tone extinction (#p < 0.05). Middle and right graphs show male and female data, respectively. Bars represent SEM.

Fig. 2

8-Tone extinction does not reverse stress-induced deficits in MDT-evoked response in the mPFC. (A) Experimental timeline for male and female rats. (B) Fear extinction curves for 16-tone and 8-tone extinction were not different over the first 8 tones. (C) Effects of 16- and 8-tone extinction on stress-attenuated MDT-evoked responses in the mPFC of males and females. The effect of stress (CUS-tones vs NS Ext-16, *p < 0.0001) was reversed by 16-tone extinction (⁺p<0.0001). There was a partial effect of 8-tone extinction (+p < 0.05), in that it was also significantly different from that of Ext-16 (#p < 0.05). In males (bottom left), there was an effect of stress (*p < 0.05) that was reversed by 16-tone extinction (⁺p<0.0001), and partially reversed by 8-tone extinction (⁺p<0.001), which also differed from 16-tone extinction (^#p < 0.01). In females, there was an effect of stress (*p < 0.05) that was reversed by 16-tone extinction (⁺p<0.001). The effect of 8-tone extinction was not significant in females. However, in females, 8-tone extinction also did not differ from 16-tone extinction. (D) Recording electrode placements in the mPFC (top) and stimulating electrode placements in the MDT (bottom). Graphics are from the atlas of Paxinos and Watson (Paxinos and Watson, 2006). (E) CUS induced a deficit in set shifting compared to nonstress controls (*p < 0.0001). The CUS-induced deficit in set-shifting was reversed by 16-tone extinction (⁺p<0.05), but not 8-tone extinction; the full stress effect was still evident in the CUS 8-tone extinction group (*p < 0.0001 vs nonstress controls). In both males (5–11/group) and females (5–6/group), there was an effect of stress (*p < 0.05). The stress effect was attenuated by 16-tone extinction, similar to the therapeutic effect seen in the combined groups, but pairwise comparisons in the sexes analyzed separately were not significant. By contrast, the stress effect was unchanged by 8-tone extinction in either sex (*p < 0.05). Bars represent SEM.

Fig. 3

Dose-dependent effects of ketamine on stress-induced deficits in MDT-evoked mPFC response and cognitive set shifting. (A) Experimental timeline for male and female rats. (B) In stressed rats, 10.0 mg/kg, 3.0 mg/kg and 1.0 mg/kg restored mPFC responsivity to MDT stimulation (⁺p<0.05). 0.3 mg/kg was not different from saline. However, 1.0 mg/kg was also different from 10.0 mg/kg (^#p < 0.05), indicating a partial effect. In males (left inset; 5–10/group), 1.0 mg/kg, 3.0 m/kg and 10.0 mg/kg all restored stress-compromised mPFC response compared to saline (⁺p<0.001). 1.0 mg/kg was also different from 10.0 mg/kg (^#p < 0.05). In females (right inset, 5–11/group), only 10.0 mg/kg differed from saline (⁺p<0.0001). (C) CUS induced a deficit in set shifting (*p < 0.001), that was reversed by ketamine at 3.0 mg/kg and 10.0 mg/kg (⁺p<0.002), but not 0.3 mg/kg or 1.0 mg/kg. Similarly, in males (top inset, 4–9/group), there was a stress effect (*p < 0.05), that was reversed only by 3.0 mg/kg and 10.0 mg/kg ketamine (⁺p<0.01). CUS induced a set-shifting deficit in females (4–6/group, bottom inset, *p < 0.05), that was also reversed by 3.0 mg/kg and 10 mg/kg (⁺p<0.05), but not 0.3 mg/kg or 1.0 mg/kg. Bars represent SEM.

Fig. 4

Adjunct treatment with 8-tone extinction plus 1.0 mg/kg ketamine reverses stress-induced set shifting deficits. (A) Extinction curves did not differ between groups. Ketamine injection was given immediately after extinction. (B) Combined treatment with 8-tone extinction plus 1.0 mg/kg ketamine significantly improved set-shifting in stressed rats (⁺p<0.001). Neither 8-tone extinction alone, nor 1.0 mg/kg ketamine alone had any effect on set-shifting in stressed rats. Similarly, in stressed males (upper inset, 4–5/group), only combined treatment with Ext-8 + 1.0 mg/kg ketamine significantly improved set-shifting in stressed rats (p < 0.0001). Likewise, in females (lower inset, n = 5–7/group), only combined treatment with Ext-8 + 1.0 mg/kg ketamine differed from CUS 1.0 mg/kg (p < 0.0001). (C) Ketamine does not enhance the effects of 8-tone extinction on evoked LFPs in the mPFC. CUS-tones + saline is different from both CUS 8-tones + saline (⁺p<0.005), and CUS 8-tones + 1.0 mg/kg ketamine (⁺p<0.005). Ketamine did not enhance the effects of 8-tone extinction (CUS 8-tones saline vs. CUS 8-tones 1.0 mg/kg ketamine, p = 0.6674). In males (middle, 3–5/group), no differences were detected between groups. In females (right, 3–5/group), CUS-tones saline is different from both CUS 8-tones saline and CUS 8-tones ketamine (⁺p<0.001). Bars represent SEM.