Acute exercise enhances fear extinction through a mechanism involving central mTOR signaling

Abstract

Impaired fear extinction, combined with the likelihood of fear relapse after exposure therapy, contributes to the persistence of many trauma-related disorders such as anxiety and post-traumatic stress disorder. Identifying mechanisms to aid fear extinction and reduce relapse could provide novel strategies for augmentation of exposure therapy. Exercise can enhance learning and memory and augment fear extinction of traumatic memories in humans and rodents. One factor that could contribute to enhanced fear extinction following exercise is the mammalian target of rapamycin (mTOR). mTOR is a translation regulator involved in synaptic plasticity and is sensitive to many exercise signals such as monoamines, growth factors, and cellular metabolism. Further, mTOR signaling is increased after chronic exercise in brain regions involved in learning and emotional behavior. Therefore, mTOR is a compelling potential facilitator of the memory-enhancing and overall beneficial effects of exercise on mental health.The goal of the current study is to test the hypothesis that mTOR signaling is necessary for the enhancement of fear extinction produced by acute, voluntary exercise. We observed that intracerebral-ventricular administration of the mTOR inhibitor rapamycin reduced immunoreactivity of phosphorylated S6, a downstream target of mTOR, in brain regions involved in fear extinction and eliminated the enhancement of fear extinction memory produced by acute exercise, without reducing voluntary exercise behavior or altering fear extinction in sedentary rats. These results suggest that mTOR signaling contributes to exercise-augmentation of fear extinction.

Keywords: Exposure therapy; Fear conditioning; Nucleus accumbens; Rapamycin; S6; Wheel running.

Figure 1.

Experimental Timeline. Cannula surgery targeting the lateral ventricle was performed on all rats. 1 week later, all rats were placed into mobile and locked wheels on alternating nights for 4 nights to equally familiarize rats with each setting. Two days after the end of the wheel familiarization procedure (3 days from the last running opportunity), rats were conditioned to fear an auditory tone in Context A. The following evening, rats were administered either vehicle or rapamycin ICV. All rats were placed into extinction Context B 10 min after ICV injection and exposed to the auditory CS 15 times in the absence of the footshock US. Immediately following fear extinction, rats were placed into either mobile or locked running wheels for 2h. The injection, fear extinction, and running procedures were repeated the following evening, for a total of two nights of injections and fear extinction followed by either mobile or locked conditions. The second fear extinction training session also served as a fear extinction retrieval test. The day after the second fear extinction trial, all rats were placed into a novel Context C to test for fear renewal. 8 days after the fear renewal test, rats were placed into locked or mobile running wheels for 10 min prior to perfusions and removal of brains for pS6 immunohistochemistry (not shown).

Figure 2.

Effects of rapamycin and acute exercise on fear extinction retrieval and relapse. A) Running distances during wheel familiarization. B) Running distance during both days of post-extinction wheel running. C) Levels of freezing during fear conditioning. Fear increased throughout conditioning and was not different between rats subsequently assigned to Veh or RAP injections, or between Sed or Run wheel conditions. D) Levels of freezing during extinction training. Rats were placed into extinction Context B and exposed to 15 CS presentations in the absence of the US. CS presentations are split into 3, 5-tone presentations and classified as early, middle (mid), or late extinction. Freezing in the RAP/Sed, RAP/Run, and Veh/Run groups decreased throughout extinction training trials. Rats in the Veh/Sed group do not show within-session extinction despite subsequent assignment to Sed or Run conditions. E) Levels of freezing during extinction retrieval. When re-exposed to the CS in the extinction context, rats in the Veh/Run group displayed significantly less fear behavior than those in the RAP/Run and Veh/Sed groups. F) Levels of freezing during the fear renewal test. All rats were re-exposed to the auditory CS in a novel Context C, indicative of fear renewal. Each group displayed similar amounts of freezing. Data displayed represent mean ± SEM; (*) P < 0.05; (#) P < 0.01.

Figure 3.

Effects of rapamycin (RAP) on the number of pS6-positive neurons in multiple brain regions following 10 min of sedentary (Sed) or voluntary wheel running (Run) conditions. A) Coronal brain sections modified from Paxinos and Watson (Paxinos 1998) indicating the brain regions where pS6-positive neurons were quantified. B) Representative photomicrographs showing pS6 immunoreactivity in brain regions in which RAP significantly reduced pS6. White scale bar = 200μm; black scale bar = 350μm. C) Number of pS6-positive cells in the dorsomedial striatum (DMS), dorsolateral striatum (DLS), dentate gyrus (DG) cornu ammonis 1, (CA1), central amygdala (CeA), and ventral tegmental area (VTA). Neither RAP nor exercise altered the number of pS6-positive cells in these regions. D) Number of pS6-positive neurons in the infralimbic cortex (IL), prelimbic cortex (PL), cornu ammonis 2 (CA2), cornu ammonis 3 (CA3), and basolateral amygdala (BLA). ICV RAP reduced numbers of pS6-positive cells in each of these regions. E) Number of pS6-positive cells in the nucleus accumbens core (NAcC) and nucleus accumbens shell (NAcS). A single session of exercise significantly increased the number of pS6-positive neurons in these regions, and this effect of exercise was prevented by ICV RAP. Data displayed represent mean ± SEM; (*) indicates significant main effect of drug (p < 0.05); (φ) indicates p < 0.05 relative to all other groups.