Enhanced contextual fear memory in central serotonin-deficient mice

Abstract

Central serotonin (5-HT) dysregulation contributes to the susceptibility for mental disorders, including depression, anxiety, and posttraumatic stress disorder, and learning and memory deficits. We report that the formation of hippocampus-dependent spatial memory is compromised, but the acquisition and retrieval of contextual fear memory are enhanced, in central 5-HT-deficient mice. Genetic deletion of serotonin in the brain was achieved by inactivating Lmx1b selectively in the raphe nuclei of the brainstem, resulting in a near-complete loss of 5-HT throughout the brain. These 5-HT-deficient mice exhibited no gross abnormality in brain structures and had normal locomotor activity. Spatial learning in the Morris water maze was unaffected, but the retrieval of spatial memory was impaired. In contrast, contextual fear learning and memory induced by foot-shock conditioning was markedly enhanced, but this enhancement could be prevented by intracerebroventricular administration of 5-HT. Foot shock impaired long-term potentiation and facilitated long-term depression in hippocampal slices in WT mice but had no effect in 5-HT-deficient mice. Furthermore, bath application of 5-HT in 5-HT-deficient mice restored foot shock-induced alterations of hippocampal synaptic plasticity. Thus, central 5-HT regulates hippocampus-dependent contextual fear memory, and 5-HT modulation of hippocampal synaptic plasticity may be the underlying mechanism. The enhanced fear memory in 5-HT-deficient mice supports the notion that 5-HT deficiency confers susceptibility to posttraumatic stress disorder in humans.

Fig. 1.

Retrieval of spatial memory is impaired and anxiety-like behavior is reduced in CKO mice. (A–D) Normal spatial learning but impaired spatial memory is seen in CKO mice. (A) WT and CKO mice had comparable mean escape latencies throughout the 9-d Morris water maze spatial learning trials (WT vs. CKO; F_8,288 = 0.813, P = 0.591). (B) One day and 10 d after training, WT mice spent significantly more time in the target quadrant than CKO mice (1-d comparison, F_1,32 = 4.442, *, P = 0.046; 10-d comparison, F_1,15 = 5.050, *, P = 0.044). (C) WT mice also made more frequent crossings onto the target site than CKO mice (1-d comparison, F_1,32 = 4.825, *, P = 0.037; 10-d comparison, F_1,15 = 16.946, **, P < 0.001). (D) WT mice exhibited shorter mean latency for the first crossing of the location of the platform than CKO mice (1-d comparison, F_1,32 = 7.073, *, P = 0.012; 10-d comparison, F_1,15 = 16.567, **, P < 0.001). (E) These CKO mice spent more time in the open arm than WT mice in the elevated-plus maze test (F_1,28 = 5.336, *, P = 0.028). (F) These CKO mice exhibited shorter mean feeding latency after exposure to a novel context than WT mice (F_1,15 = 26.782, **, P < 0.001).

Fig. 2.

Intensity and persistence of foot shock-induced fear memory are enhanced in 5-HT-deficient mice, but this enhancement is reversed to the level of WT mice after intracerebroventricular administration of 5-HT. (A) Foot shock provokes a significant increase of freezing behavior in CKO mice after the fourth and fifth shock compared with WT mice (after fourth shock, *, P = 0.036; after fifth shock, *, P = 0.025). (B) Thirty minutes, 1 d, and 10 d after fear conditioning, exposure to the conditioned environment evokes much stronger freezing behavior in CKO mice than in WT mice (30 min, *, P = 0.033; 1 d, *, P = 0.002; 10 d, **, P = 0.001). (C) Enhanced contextual fear memory 1 d after conditioning in CKO mice (CKO 5-HT vs. CKO saline, *, P = 0.032) is converted to the level of WT mice after intracerebroventricular administration of 5-HT (CKO 5-HT vs. WT saline, P = 0.468; CKO 5-HT vs. WT 5-HT, ^#, P = 0.045). Enhanced contextual fear memory in CKO mice is reduced by i.p. injection of DPAT (*, P = 0.008; t test), but it does not reach the level of WT mice (CKO DPAT vs. WT saline; ^§P = 0.030; CKO DPAT vs. WT DPAT, ^#, P = 0.025). Administration of 5-HT but not DPAT in WT mice also reduces contextual fear memory compared with WT mice treated with saline solution (*, P = 0.002). (D) Enhanced contextual fear memory in CKO mice 10 d after conditioning is also converted by intracerebroventricular administration of 5-HT (CKO 5-HT vs. CKO saline, **, P < 0.001) to a level similar to WT mice (CKO 5-HT vs. WT saline, P = 0.250; CKO 5-HT vs. WT 5-HT, P = 0.385). DPAT administration via i.p. route also reduced contextual fear memory in CKO mice (n = 7 for each; CKO DPAT vs. CKO saline, *, P = 0.040), but freezing time was still higher than in WT mice (CKO DPAT vs. WT saline, §, P = 0.008; CKO DPAT vs. WT DPAT, ^#, P = 0.040). Administration of either 5-HT or DPAT in WT mice did not reduce freezing behavior compared with WT mice treated with saline solution at this time point.

Fig. 3.

Foot shock impairs hippocampal CA1 LTP measured by fEPSP amplitude (% baseline) at the striatum radiatum in WT, but has no effect in CKO mice. However, this inability is restored after bath application of 5-HT and DPAT (an agonist of 5HT1A receptor) in CKO slices. (A, B, and E) Foot shock (Fear) impairs HFS-evoked LTP in hippocampal slices from WT mice (F_1,24 = 8.440, *, P = 0.008), but has no effect on LTP in CKO mice (F_1,15 = 0.358, P = 0.558). (B, C, and F) When 5-HT is applied in the hippocampal slices of CKO mice, foot shock is able to impair LTP (Fear CKO 5-HT vs. Fear CKO, F_1,13 = 10.472, *, P = 0.006), as it does in WT mice. (B, D, and F) Foot shock-induced impairment of LTP is partially restored after bath application of DPAT in CKO mice (Fear CKO DPAT vs. Fear CKO, F_1,18 = 5.443, *, P = 0.031).

Fig. 4.

Foot shock facilitates hippocampal CA1 LTD measured by fEPSP amplitude (% baseline) at the striatum radiatum in WT mice but has no effect in CKO mice. However, this inability is restored after bath application of 5-HT. (A, B, and E) LFS of the CA1 of hippocampal slice from WT mice induces small LTP, but evokes neither LTP nor LTD in CKO mice (naïve CKO vs. naïve WT, F_1,12 = 17.775, *, P = 0.002). Foot shock (Fear) converts LFS-induced LTP to LTD in WT mice (Fear WT vs. naïve WT, F_1,11 = 33.562, ^##, P < 0.001; one-way ANOVA), whereas foot shock has no effect in CKO mice (Fear CKO vs. naïve CKO, F_1,10 = 0.225, P = 0.645). (C, D, and F) The failure of LFS-induced LTD in CKO mice is rescued by bath application of 5-HT (Fear CKO 5-HT vs. Fear CKO, F_1,10 = 3.825, *, P = 0.039), but not DPAT (Fear CKO DPAT vs. Fear CKO, F_1,10 = 0.183, P = 0.678).