Targeted Memory Reactivation in REM But Not SWS Selectively Reduces Arousal Responses

Contact: Audrey Hu audrey.hu@wecistanche.com

A growing body of evidence suggests that sleep can help to decouple the memory of emotional experiences from their associated effective charge. This process is thought to rely on the spontaneous reactivation of emotional memories during sleep, though it is still unclear which sleep stage is optimal for such reactivation. We examined this question by explicitly manipulating memory reactivation in both rapid-eye-movement sleep(REM)and slow-wave sleep(SWS)using targeted memory reactivation (TMR)and testing the impact of this manipulation on the habituation of subjective arousal responses across a night. Our results show that TMR during REM, but not SWS significantly decreased subjective arousal, and this effect is driven by the more negative stimuli. These results support one aspect of the sleep to forget, sleep to remember(SFSR)hypothesis which proposes that emotional memory reactivation during REM sleep underlies sleep-dependent habituation.

effects of cistanche: improve memory

Targeted memory reactivation (TMR) is a technique in which memory reactivation is intentionally triggered in sleep through the re-presentation of cues that were linked to the memory in the wake, and is commonly achieved using sounds or smells, see14 for a review. While early studies reported a benefit of TMR in REM sleep on overnight memory consolidation15,16, a growing number of reports suggest that TMR is beneficial when presented during non-REM17−21, but not during REM17,19,22 see23 for meta-analysis. In terms of how TMR may influence emotional arousal, one recent study showed an impact of non-REM TMR on pleasantness and arousal ratings, but this only emerged in socially anxious participants and after a week of consolidation24. Very few studies have investigated the impact of REM TMR on emotional material. Thus, inducing reactivation of fear memories during REM was shown to result in increased generalization25, and TMR of a Pavlovian conditioning task during REM lead to increased habituation compared to TMR of the same task during stage 2 of non-REM26.

Building on this literature, we set out to test the SFSR prediction that replaying emotionally arousing memories during REM, but not SWS, would be associated with a reduced arousal rating for this material the next day. We did this by manipulating the reactivation of emotional memories during sleep using TMR. Our participants rated emotionally negative and neutral picturesound pairs for arousal both before and after a night of sleep. During sleep, we cued half of the negative and half of the neutral stimuli for reactivation by softly replaying the associated sounds. We then examined the impact of this TMR upon overnight habituation of the arousal response. We carefully controlled the sleep stage in which TMR was applied, cueing participants either in the REM (REM Group) or SWS (SWS Group), see Fig. 1. Based on the SFSR hypothesis, we predicted that TMR would lead to greater habituation when applied during REM, but not SWS.

effects of cistanche: improve memory

Results

At baseline, arousal ratings were higher for negative than for neutral items, showing that participants rated the stimuli in keeping with expectations. This was confirmed by a 2×2x2 ANOVA on pre-sleep arousal with the factors Group, Cueing, and Emotion(Fus)=337.93,p<0.001; Paired t testp<0.001in both SWS and REM groups). Ratings did not differ between Cued and Un-cued stimuli prior to sleep(paired t-tests p>0.2 in both cases), showing that the baseline was well balanced, and no participants reported awareness that sounds were played during their sleep.

We investigated how overnight habituation of arousal was modulated by TMR cueing using a 2×2×2 ANOVA with factors Group, Cueing, and Emotion, see Table 1. Overnight habituation was calculated as(second pre-sleep arousal rating - post-sleep arousal rating)/ first pre-sleep arousal rating. This revealed a significant interaction between Group and Cueing (F(1,32) = 5.341, p = 0.027) suggesting that the impact of cueing on this measure of habituation depends on the cognitive state during which TMR is applied. The 2 × 2 × 2 ANOVA gave no other significant result. To determine which sleep stage drove the Group x Cueing interaction, we then conducted separate 2 × 2 ANOVAs with the factors Cueing and Emotion in SWS and REM groups, respectively. This showed a main effect of cueing in REM (F(1,14) = 7.48, p < 0.02) but not SWS (F(1,18) = 0.086, p = 0.8), see Fig. 2. Closer examination of the REM group showed that the effect of Cueing was driven by the negative items (paired test t = 3.21; p = 0.006), with neutral items showing a trend towards the same effect (t = 1.6; p = 0.102). Interestingly, there was a main effect of emotion in the SWS group (F(1,18) = 6.28, p = 0.022) but not in the REM group. Closer examination showed that this was driven by greater habituation of neutral items compared to negative items in the Un-cued condition (t = 2.25, p = 0.037), with a trend in the Cued condition (t = 1.45, p = 0.163).

Memory for the sound-image pairs was near the ceiling during the pre-sleep test. On average, participants responded correctly on >97% of trials. The mean proportion of incorrect trials was as follows [REM group: 1.45%(±1.98)trials(pre-sleep);1.55%(±1.83) trials (post-sleep);SWS group:2.05%(±2.77)trials (pre-sleep);2.62%(±1.08)trials (post-sleep)]. To test for differences in pre-sleep learning of sound-image which could have biased the results, we conducted ANOVAs on pre-sleep proportion correct memory trials with the factors cued/not cued and negative/neutral for the REM and SWS groups, respectively. This revealed no main effects or interactions in the either REM group or the SWS group(p>0.1 in all cases). To test for impacts of TMR or emotion on overnight changes in memory, we performed the same two ANOVAs, now with the overnight change in the proportion of correct memory trials as the dependent variable. This revealed no significant effects for either SWS (p>0.5)in all cases or REM(p >0.8)in all cases.

To ensure that reaction times did not differ markedly between cued and un-cued items during the pre-sleep arousal test, we performed a2×2 ANOVA with the factors valence (neutral, negative)and cueing (un-cued, cued)within each group of participants. This revealed a main effect of emotion (F=29.9,p< 0.001)in the SWS group, and a trend towards the same effect of emotion(F=3.984, p=0.066)in the REM group. Because response times are often modulated by both TMR and emotion, we examined the effects of cueing and valence on overnight change in reaction times for REM and SWS groups. We used a pair of 2×2 ANOVA with the factors valence (neutral, negative) and cueing(un-cued, cued). This revealed no main effect or interaction in either REM or SWS groups (p>0.15)in all cases, see Supplementary Information Table S1 for means.

Sleep stage data for each group are reported in Table 2. Note that REM time, SWS time, N2 time, and NI time did not differ significantly between groups (p>0.1 in all cases). However, total sleep time did differ significantly(p=0.01), being shorter on average in the REM than in the SWS group. To determine whether there was a relationship between habituation and time spent in REM or SWS or spectral power in slow-wave, delta, theta, and gamma bands we conducted a series of Pearson correlations with data across both REM and SWS groups; these revealed several marginal correlations, but none of these survived correction for our four multiple comparisons (p>0.05 in all cases).

effects of cistanche: improve memory

Discussion

Our data support a role for REM sleep in overnight habituation of arousal responses to picture-sound combinations by demonstrating that TMR in REM but not SWS leads to increased overnight habituation. This finding is in line with a previous observation that REM TMR of sounds used in pre-sleep Pavlovian conditioning resulted in greater habituation towards these sounds the next day than TMR of the same sounds in stage 2 sleep-26. These two studies combine to support the one aspect of the sleep to forget, sleep to remember hypothesis27, e.g., the idea that memory reactivation in REM is specifically associated with reductions in subsequent arousal.

A recent study by Lehmann et al22. used a similar design to investigate the role of TMR in REM and SWS in emotional memory consolidation and found no effect of cueing on subjective habituation in any group22. One possible reason for the difference between these results and our own may stem from the fact that our participants rated pictures for arousal immediately before and after sleep, while participants in Lehmann 2016 performed learning and retrieval tasks after they rated the stimuli but before sleep. Repeated viewing of stimuli can lead to habituation, so Lehmann’s design may have allowed pre-sleep habituation to drown out any effects of TMR.

A number of studies suggest that SWS may play a role in habituation8, for instance, the observation that aspects of autonomic habituation are predicted by SWS28, that SWS percentage predicts overnight emotional attenuation29, and that blocking the release of noradrenaline during SWS diminished this habituation12. Interestingly, we observed significantly stronger habituation of neutral as compared to negative items in the SWS group irrespective of TMR cueing, possibly because the neutral items are less strongly arousing, to begin with, and it is, therefore, easier for participants to alter the way these items are rated than it is for strongly arousing negative. Alternatively, the arousal states of neutral items could be more open to interpretation than the arousal status of negative items, for instance, responses to negative pictures may be influenced by top-down processes that identify them as conceptually negative and increase the likelihood of a strong arousal rating. Although difficult to interpret, the marked difference between this pattern of consolidation in the SWS group and the pattern of habituation observed in the Uncued REM group, where negative and neutral items habituated to the same extent, could suggest that REM TMR disrupts a natural process of habituation across NREM sleep which works better for comparatively neutral stimuli than for those which are more arousing.

While our findings appear to support a unique role for REM sleep in modulating emotional arousal, we found no correlation between REM time or theta power and habituation. As REM occurs later in the night than SWS we cannot rule out the possibility that the dissociation we observed between these sleep stages was caused by this difference in timing. Notably, however, in Rihm and Rasch26 cue-related habituation was more pronounced when cues were presented during REM than stage 2 sleep within the same period of early morning sleep, supporting the idea that this difference depends on the sleep stage in which TMR is performed rather than the time of night at which it is performed.

Only female participants were included in this study. This was due to previous studies reporting a greater self-reported response towards negative stimuli in female vs. male individuals. Future work should extend these investigations to males. Finally, it is worth noting that, throughout the study, participants rated the sound-picture pair, rather than just pictures or sounds. Our results, therefore, relate to this multimodal pair rather than sounds or images alone.

effects of cistanche: improve memory

References

1. Rasch, B. & Born, J. About sleep’s role in memory. Physiol. Rev. 93, 681–766 (2013).

2. Hu, P., Stylos-Allan, M. & Walker, M. P. Sleep facilitates consolidation of emotional declarative memory. Psychol. Sci. 17, 891–898 (2006).

3. Payne, J. D., Chambers, A. M. & Kensinger, E. A. Sleep promotes lasting changes in selective memory for emotional scenes. Front. Integer. Neurosci. 6, 108 (2012).

4. Wagner, U., Hallschmid, M., Rasch, B. & Born, J. Brief sleep after learning keeps emotional memories alive for years. Biol. Psychiatry 60, 788–790 (2006).

5. Wagner, U., Kashyap, N., Diekelmann, S. & Born, J. The impact of post-learning sleep vs. wakefulness on recognition memory for faces with different facial expressions. Neurobiol. Learn. Mem. 87, 679–687 (2007).

6. Dolcos, F., LaBar, K. S. & Cabeza, R. Remembering one year later: role of the amygdala and the medial temporal lobe memory system in retrieving emotional memories. Proc. Natl Acad. Sci. USA 102, 2626–2631 (2005).

7. Goldstein, A. N. & Walker, M. P. The role of sleep in emotional brain function. Annu. Rev. Clin. Psychol. 10, 679–708 (2014).

8. van der Helm, E. et al. REM sleep depotentiates amygdala activity to previous emotional experiences. Curr. Biol 21, 2029–2032 (2011).

9. Gujar, N., McDonald, S. A., Nishida, M. & Walker, M. P. A role for rem sleep in recalibrating the sensitivity of the human brain to specific emotions. Cereb. Cortex 21, 115–123 (2011).

10. Baran, B., Pace-Schott, E. F., Ericson, C. & Spencer, R. M. C. Processing of emotional reactivity and emotional memory oversleep. J. Neurosci. 32, 1035–1042 (2012).

11. Groch S., Wilhelm I., Diekelmann S. & Born, J. The role of REM sleep in the processing of emotional memories: Evidence from behavior and event-related potentials. Neurobiol. Learn. Mem. 99, 1–9 (2013).

12. Groch, S. et al. Contribution of norepinephrine to emotional memory consolidation during sleep. Psychoneuroendocrinology 36, 1342–1350 (2011).

13. Wagner, U., Fischer, S. & Born, J. Changes in emotional responses to aversive pictures across periods rich in slow-wave sleep versus rapid eye movement sleep. Psychosom. Med. 64, 627–634 (2002).

14. Cellini, N. & Cappuzo, A. Shaping memory consolidation via targeted memory. Ann. NY Acad. Sci. 1426, 52–71 (2018).

15. Guerrini, A., Dujardin, K., Mandal, O., Sockeel, P. & Leconte, P. Enhancement of memory by auditory stimulation during post-learning REM sleep in humans. Physiol. Behav. 45, 947–950 (1989).

16. Tilley, A. J. Sleep learning during stage 2 and REM sleep. Biol. Psychol. 9, 155–161 (1979).

17. Cordi, M. J., Diekelmann, S., Born, J. & Rasch, B. No effect of odor-induced memory reactivation during REM sleep on declarative memory stability. Front. Syst. Neurosci. 8, 157 (2014).

18. Fuentemilla, L. et al. Hippocampus-dependent strengthening of targeted memories via reactivation during sleep in humans. Curr. Biol. 23, 1769–1775 (2013).

19. Rasch, B., Buchel, C., Gais, S. & Born, J. Odor cues during slow-wave sleep prompt declarative memory consolidation. Science 315, 1426–1429 (2007).

20. Rudoy, J. D., Voss, J. L., Westerberg, C. E. & Paller, K. A. Strengthening individual memories by reactivating them during sleep. Science 326, 1079 (2009).

21. Schreiner, T. & Rasch, B. Boosting vocabulary learning by verbal cueing during sleep. Cereb. Cortex https://doi.org/10.1093/cercor/bhu139 (2015).

22. Lehmann, M., Schreiner, T., Seifritz, E. & Rasch, B. Emotional arousal modulates oscillatory correlates of targeted memory reactivation during NREM, but not REM sleep.

23. Hu, X., Cheng, L. Y., Chiu, M. H. & Paller, K. A. Promoting memory consolidation during sleep: A meta-analysis of targeted memory reactivation. Psychol. Bull. 146, 218–244 (2020).

24. Groch, S. et al. Targeted reactivation during sleep differentially affects negative memories in socially anxious and healthy children and adolescents. J. Neurosci. 37, 2425–2434 (2017).

25. Sterpenich, V. et al. Memory reactivation during rapid eye movement sleep promotes its generalization and integration in cortical stores. Sleep 37, 1061–1075 (2014).

26. Rihm, J. S. & Rasch, B. Replay of conditioned stimuli during late REM and stage N2 sleep influences affective tone rather than emotional memory strength. Neurobiol. Learn. Mem. 122, 142–151 (2015).

27. Walker, M. P., van der, H. E. & van der Helm, E. Overnight therapy? The role of sleep in emotional brain processing. Psychol. Bull. 135, 731–748 (2009).

28. Pace-Schott, E. F. et al. Napping promotes inter-session habituation to emotional stimuli. Neurobiol. Learn. Mem. 95, 508–523 (2012).

29. Talamini, L. M., Bringmann, L. F., de Boer, M. & Hofman, W. F. Sleeping worries away or worrying away sleep? Physiological evidence on sleep-emotion interactions.

30. Horne, J. A. & Ostberg, O. A self-assessment questionnaire to determine morningness-eveningness in human circadian rhythms. Int. J. Chronobiol. 4, 97–110 (1976).

31. Lang, P. J., Bradley, M. M. & Cuthbert, B. N. International Affective Picture System (IAPS): Affective Ratings of Pictures and Instruction Manual. Technical Report No. A-8 (University of Florida, 2008).

32. Willenbockel, V. et al. Controlling low-level image properties: the SHINE toolbox. Behav. Res. Methods 42, 671–684 (2010).

33. Bradley, M. M. & Lang, P. J. International Affective Digitized Sounds (IADS): Stimuli, Instruction Manual and Affective Ratings. Technical Report No. B-2 (1999).

34. Breiter, H. C. et al. Response and habituation of the human amygdala during visual processing of a facial expression. Neuron 17, 875–887 (1996).

35. Büchel, C., Dolan, R. J., Armony, J. L. & Friston, K. J. Amygdala-hippocampal involvement in human aversive trace conditioning revealed through event-related functional magnetic resonance imaging. J. Neurosci. 19, 10869–10876 (1999).

36. Fischer, H., Furmark, T., Wik, G. & Fredrikson, M. Brain representation of habituation to repeated complex visual stimulation studied with PET. Neuroreport 11, 123–126 (2000).

37. Bradley, M. M. & Lang, P. J. Measuring emotion: the self-assessment manikin and the semantic differential. J. Behav. Ther. Exp. Psychiatry 25, 49–59 (1994).

38. Iber, C., Ancoli-Israel, S., Chesson, A. & Quan, S. The AASM Manual for the Scoring of Sleep and Associated Events (American Academy for Sleep Medicine, 2007).