Postreactivation glucocorticoids impair recall of established fear memory

Abstract

Pavlovian fear conditioning provides one of the best rodent models of acquired anxiety disorders, including posttraumatic stress disorder. Injection of a variety of drugs after training in fear-conditioning paradigms can impair consolidation of fear memories. Indeed, early clinical trials suggest that immediate administration of such drugs after a traumatic event may decrease the risk of developing posttraumatic stress disorder in humans (Pitman et al., 2002; Vaiva et al., 2003). The use of such a treatment is limited by the difficulty of treating every patient at risk and by the difficulty in predicting which patients will experience chronic adverse consequences. Recent clinical trials suggest that administration of glucocorticoids may have a beneficial effect on established posttraumatic stress disorder (Aerni et al., 2004) and specific phobia (Soravia et al., 2006). Conversely, glucocorticoid administration after training is known to enhance memory consolidation (McGaugh and Roozendaal, 2002; Roozendaal, 2002). From a clinical perspective, enhancement of a fear memory or a reactivated fear memory would not be desirable. We report here that when glucocorticoids are administered immediately after reactivation of a contextual fear memory, subsequent recall is significantly diminished. Additional experiments support the interpretation that glucocorticoids not only decrease fear memory retrieval but, in addition, augment consolidation of fear memory extinction rather than decreasing reconsolidation. These findings provide a rodent model for a potential treatment of established acquired anxiety disorders in humans, as suggested by others (Aerni et al., 2004; Schelling et al., 2004), based on a mechanism of enhanced extinction.

Figure 1.

Postretrieval corticosterone transiently impairs subsequent recall of contextual fear conditioning. a, Percentage of time spent freezing in the training context before training in fear conditioning (Baseline), during subsequent reactivation 48 h hence (48 h from Train), and 24 h after reactivation (72 h from Train). Reactivation was followed by injection of vehicle or corticosterone at the indicated dose (one-way ANOVA for 72 h time point, p < 0.001, F_(3,28) = 10.06; post hoc Tukey's test, p < 0.01 for Cort 3 mg/kg vs vehicle, p < 0.001, for Cort 10 mg/kg vs vehicle, p = 0.65, for Cort 3 mg/kg vs Aniso, p = 0.96, for Cort 10 mg/kg vs Aniso, p < 0.001, for Aniso vs vehicle, p = 0.90, for Cort 3 mg/kg vs Cort 10 mg/kg). b, Subsequent contextual memory recall is blocked by higher doses of corticosterone, and this effect resembles that of anisomycin. A dose–response curve for corticosterone administered immediately after memory reactivation is shown. Bars represent percentage of freezing 72 h after training (24 h after memory reactivation) (one-way ANOVA, p < 0.0001, F_(6,49) = 10.86; post hoc Tukey's test, vehicle vs no injection, p = 0.95, vs Cort 0.3 mg/kg, p = 0.11, vs Cort 1 mg/kg, p = 0.99, vs Cort 3 mg/kg, p < 0.05, vs Cort 10 mg/kg, p < 0.01, vs Aniso, p < 0.001). No Injection was significantly different from Aniso, Cort 3 mg/kg, and Cort 10 mg/kg (p < 0.001 for all) and not different from Vehicle, Cort 0.3 mg/kg, or Cort 1 mg/kg (p = 0.95, 0.15, and 0.92, respectively). Cort 3 mg/kg differed from vehicle, Cort 1 mg/kg, and No Injection (p < 0.05, 0.05, and 0.01, respectively) but did not differ from Cort 0.3 mg/kg or Aniso (p = 0.39 and 0.92, respectively). Similarly, Cort 10 mg/kg differed from vehicle, Cort 1 mg/kg, and No Injection (p < 0.01, 0.01, and 0.001, respectively) but did not differ from Cort 0.3 mg/kg or Aniso (p = 0.11 and 0.99, respectively). ∗p < 0.05 on post hoc Tukey's test versus vehicle and No Injection. c, Effect of corticosterone on subsequent memory recall is transient. Error bars represent freezing 120 h (5 d) after training (72 h after memory reactivation). When tested 72 h after reactivation, the anisomycin effect on subsequent recall remained, whereas that of corticosterone is no longer present (ANOVA main effect of drug, p < 0.01, F_(2,21) = 7.72; post hoc Tukey's test vehicle vs Aniso, ∗p < 0.01, vehicle vs Cort, p = 0.34; Aniso vs Cort, p = 0.058). Although b and c were necessarily performed in separate experiments in different groups on different days and thus normalized to separate control groups, a Student's t test comparison between Cort 3 mg/kg at 24 h from b and Cort 3 mg/kg at 72 h from c revealed a p = 0.068, whereas a similar comparison of Aniso 150 mg/kg at 24 h from b and Aniso 150 mg/kg from c revealed a p = 0.94. n = 8 in all groups. Error bars represent SEM in all figures.

Figure 2.

Reactivation of the memory is necessary for the corticosterone effect on subsequent recall. Mean percentage time spent freezing in contextual memory test 24 h after injection of corticosterone or vehicle is shown (72 h from Train). ANOVA revealed no main effect of drug (p = 0.33; F_(2,33) = 1.16). Because there was no main effect, post hoc Tukey's test revealed no significant differences between vehicle versus Cort 3 mg/kg or Cort 10 mg/kg (p = 0.88 and 0.31, respectively) and no differences between Cort 3 mg/kg versus Cort 10 mg/kg (p = 0.57). n = 12 in all groups.

Figure 3.

a, Reminder shock rescues post-reactivation glucocorticoid effect on subsequent memory and spares that of anisomycin. ANOVA at the 72 h time point after reminder shock reveals a main effect of drug (p < 0.00001; F_(2,33) = 19.52). Post hoc Tukey's test shows significant differences between vehicle and Aniso and between Cort and Aniso (p < 0.001 for both). There was no significant difference between vehicle and Cort with reminder shock (p = 0.91). ∗∗∗p < 0.001 versus corresponding vehicle control. No significant differences were observed in pretrain baseline or 48 h posttrain groups (n = 12 in all groups). b, Comparison of reminder shock (+RS) versus no-reminder shock (no RS) groups on the final day of testing for each drug. ∗∗∗p < 0.001 versus corresponding vehicle control. Two-way ANOVA: main effect of drug, F_(2,66) = 37.25, p < 0.001; main effect of RS/no RS, F_(1,66) = 13.76, p < 0.001; interaction, F_(2,66) = 5.42, p < 0.01. Post hoc Tukey's test indicated significant differences between Cort/no RS, Aniso/RS, and Aniso/no RS versus all other groups and vice versa; p < 0.001 for all post hoc differences. Nonsignificant p values were 0.36 for Cort/no RS versus Aniso/no RS, 0.47 for Aniso/RS versus Aniso/no RS, 0.99 for Aniso/RS versus Cort/no RS, 0.99 for Cort/RS versus Vehicle/no RS, and 0.99 for Vehicle/RS versus Vehicle/no RS (n = 12 in all groups). No significant differences were observed in pretrain baseline or 48 h posttrain groups.

Figure 4.

Corticosterone impairs acute retrieval of contextual fear memory without affecting the stability of the underlying memory trace. a, Injection of corticosterone 30 min before memory test impairs retrieval. Results of “Probe a” (contextual memory test performed 48 h after training; percentage freezing in context 30 min after corticosterone injection) are shown. ANOVA revealed a main effect of drug (p < 0.01, F_(2,33) = 7.49). Post hoc Tukey's test revealed significant differences between vehicle versus Cort 3 mg/kg (p < 0.01) and versus Cort 10 mg/kg (p < 0.01). No significant difference between Cort 3 mg/kg and Cort 10 mg/kg was observed (p = 0.99). ∗∗p < 0.01; n = 12 in all groups. b, Impairment of memory retrieval by corticosterone had no significant effect on subsequent memory. Results of “Probe b” (contextual memory test performed 72 h after training or 24 h after Probe a; percentage freezing in context 24 h) are shown. ANOVA revealed no main effect of drug (p = 0.28; F_(2,33) = 1.33). Because there was no main effect, post hoc Tukey's test revealed no significant differences between vehicle versus Cort 3 mg/kg or Cort 10 mg/kg (p = 0.36 and 0.33, respectively) and no differences between Cort 3 mg/kg versus Cort 10 mg/kg (p = 0.99). When freezing in the Cort groups at 48 h in a (retrieval block by Cort) was compared with that of Cort groups at 72 h in b (memory after retrieval block by Cort), there was no significant difference between Cort groups with retrieval blocked (a) and Cort groups after retrieval blockade (b) (p = 0.26 and 0.23 for 3 mg/kg and 10 mg/kg, respectively).

Figure 5.

Corticosterone augments multiple-trial extinction. a, Corticosterone blockade of retrieval augments extinction. This effect is dependent on multiple reactivation trials (compare with Fig. 4b). Repeated injection of corticosterone 30 min before extinction trials for 3 d impairs both retrieval acutely and subsequent memory recall tested 24 h after the last extinction trial in the absence of corticosterone. ANOVA with repeated measures on days 1–3 indicates a main effect of drug (F_(1,18) = 6.29; p < 0.05). Student's t test of day 4 indicates p < 0.01. Individual t tests for days 1, 2, and 3 indicate p = 0.14, p < 0.05, and p < 0.05, respectively. n = 10 in all groups. b, Multiple trials of post-reactivation corticosterone further augment extinction measured 24 h after the third trial (Trial 4) and prolongs the extinction effect (Probe, measured 72 h after Trial 4). Although a single trial of post-reactivation corticosterone has an effect on subsequent memory, multiple trials seem to enhance this effect (Trial 4). A memory test 3 d hence reveals a longer-lasting effect of post-reactivation corticosterone after multiple reactivation trials (Probe) compared with single-trial reactivation (Fig. 1). Repeated-measures ANOVA reveals main effects of drug (F_(1,22) = 13.88; p < 0.01), day (F_(3,66) = 42.09, p < 0.001), and interaction between drug and day (F_(3,66) = 14.69; p < 0.001). Student's t test of days 1–4 and 7 indicate p = 0.75, p < 0.01, p < 0.001, p < 0.001, and p < 0.001, respectively. c, The effect of corticosterone on extinction with multiple trials requires memory reactivation. Multiple daily injections of corticosterone in the absence of memory reactivation do not affect subsequent memory recall. Student's t test of probe: p = 0.97; n = 12 in all groups.