Sleep Deprivation and Phasic Activity of REM Sleep: Independence of ...

SLEEP DEPRIVATION AND PHASIC ACTIVITY OF REM SLEEP

Sleep Deprivation and Phasic Activity of REM Sleep: Independence of Middle-Ear Muscle Activity From Rapid Eye Movements Luigi De Gennaro PhD, and Michele Ferrara PhD Dipartimento di Psicologia - Università degli Studi di Roma "La Sapienza" Abstract: In the recovery nights after total and partial sleep deprivation there is a reduction of rapid eye movements during REM sleep as compared to baseline nights; recent evidence provided by a selective SWS deprivation study also shows that the highest percentage of variance of this reduction is explained by SWS rebound. The present study assesses whether the reduction of rapid eye movements (REMs) during the recovery night after total sleep deprivation is paralleled by a decrease of middle-ear muscle activity (MEMA), another phasic muscle activity of REM sleep. Standard polysomnography, MEMA and REMs of nine subjects were recorded for three nights (one adaptation, one baseline, one recovery); baseline and recovery night were separated by a period of 40 hours of continuous wake. Results show that, in the recovery night, sleep deprivation was effective in determining an increase of SWS amount and of the sleep efficiency index, and a decrease of stage 1, stage 2, intra-sleep wake, and NREM latencies, without affecting REM duration and latency. However, MEMA frequency during REM sleep did not diminish during these nights as compared to baseline ones, while there was a clear effect of REM frequency reduction. Results indicate an independence of phasic events of REM sleep, suggesting that the inverse relation between recovery sleep after sleep deprivation and REM frequency is not paralleled by a concomitant variation in MEMA frequency. Key words: Middle-ear muscle activity (MEMA); rapid eye movements (REMs); sleep deprivation; phasic activity; REM sleep INTRODUCTION

decrease was linearly related to the amount of sleep curtailment and to the SWS rebound.15 Finally, a decrease of REM frequency has been found in the recovery nights after selective SWS deprivation obtained without affecting total sleep time, that is an effect showing the independence of the REM frequency decrease from prior sleep duration.16 Furthermore, REM frequency correlated more with SWS amount than with arousal thresholds, suggesting that REMs are inversely correlated with SWS amount more than with sleep depth.16 It may be interesting to assess whether other phasic components also inversely covary with SWS and/or sleep depth, with the double aim of generalizing this inverse relationship and of providing a unifying framework for the phasic muscle activity of REM sleep. In fact, REMs, together with muscle twitches and middle-ear muscle activity (MEMA), reflect intense phasic neural discharge occurring in the brain during a state of activation phasically overcoming motoneural inhibition of REM sleep, and their occurrence does not determine sleep interruption, not needing to be inhibited during REM sleep.17 It has also been suggested that they could be a startle response which may

IT HAS FREQUENTLY BEEN REPORTED THAT RAPID EYE MOVEMENTS (REMS) DURING REM SLEEP INCREASE ACROSS SLEEP CYCLES1-2 and that, in ad libitum sleep, REMs are directly related to prior sleep duration, suggesting that they might be an index of sleep need or "satiety3-4 ". A decrease of REM density[1] was also found after total sleep deprivation5-6 and after the intake of sedative hypnotics.7-12 A negative relationship between REMs and sleep depth and/or sleep need has been more directly confirmed by experiments showing a significant reduction of REM density after a night in which sleep was restricted to 100 minutes,13 and after a night with the curtailment of the last four hours of sleep.14 Recently, variations in REM frequency were assessed in the recovery night following a gradual sleep restriction, obtained by postponing the sleep onset time while maintaining the final awakening time constant. Sleep curtailment decreased REM frequency in the ensuing recovery nights, and this Accepted for publication April 1999 Corresponding Author: Luigi De Gennaro, Ph.D., Sezione di Neuroscienze, Dipartimento di Psicologia - Università degli Studi di Roma "La Sapienza", Via dei Marsi, 78; 00185 ROMA, ITALY, Tel: +39-6-49917647, Fax: +39-6-4451667, E-mail: [email protected] SLEEP, Vol. 23, No. 1, 2000

1. Rapid eye movements during REM sleep have been conventionally defined as REM density, because they were not scored as individual events, but as the number of intervals (1-20 s long) with REM activity. The term REM frequency will be used in this paper only when the number of movements per minutes of REM sleep has been considered as a measure of rapid eye movements.

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Sleep Deprivation and Phasic Activity of REM Sleep—De Gennaro et al

operate through a central motor command center.18 Correlations have been empirically assessed between different types of phasic motor responses of REM sleep: a) motor activity in extremities and the head, and MEMA;19 b) periorbital integrated potentials and MEMA;20 c) REMs and MEMA;19,21 d) muscle tone changes and REMs;22 muscle twitches and REMs in narcoleptic and apneic subjects;23 muscle twitches and REMs in narcoleptic and normal subjects.24 However, the overall relationships are low and the correlational approach seems quite weak, leaving the problem of a unique center of phasic motor activity during REM sleep unsolved. The robust phenomenon of an inverse relation between REMs and SWS (and, secondarily, sleep depth) at present represents one of the clearest psychophysiological correlates of REMs, and it can also be a useful tool to re-examine the problem of a unique center of phasic motor activity during REM sleep. In other words, if there is a unique center of phasic motor activity during REM sleep, the decrease in REM frequency on nights in which there is an increase of SWS should be paralleled by a concomitant decrease of other phasic responses, such as MEMA. This muscle activity was first described in humans by Pessah and Roffwarg.21 Most of this activity takes place within REM sleep,21 more frequently just before or at the beginning of each REM period,25 with a weak correlation with REMs and periorbital integrated potentials20 and a stronger one with motor activity in the extremities and the head.19 The aim of the present study is to evaluate whether the inverse relationship between SWS amount and rapid eye movements is also true for middle-ear muscle activity. The hypothesis posits that in the recovery night after a total sleep deprivation there is a parallel decrease of MEMA and REM frequency, as a consequence of the increase of SWS.

started at about 11:30 PM and ended after 7.5 hours of accumulated sleep. In the adaptation and baseline nights, each subject arrived at the sleep laboratory at 9:00 PM and, after electrode montage, his undisturbed sleep was recorded. A 40-hour schedule of sleep deprivation began on morning awakening following the baseline night. Subjects remained in the sleep laboratory together with at least one experimenter throughout the whole course of the 40 hours of continuous wake. Every two hours, variations of alertness were assessed respectively by means of subjective ratings of sleepiness (i.e., the Stanford Sleepiness Scale (SSS) and the Sleepiness Visual Analog Scale (SVAS)) and of behavioral measures (i.e., a Letter Cancellation Test).26 Furthermore, oculomotor activity (smooth pursuit and saccadic eye movements, and optokinetic responses) was also recorded, as a possible indicator of sleepiness. Data related to these measures will be reported elsewhere. The duration of each session was 30 minutes. Subject's meals were scheduled at 8:30, 14:30, and 19:30; nonscheduled snacks were permitted. The 40-hour schedule of sleep deprivation ended at 10:00 PM; the recording of the recovery night began at about 11:30 PM. Subjects were awakened after 7.5 hours of accumulated sleep. Sleep Recording

Participants slept for three nights in a sound-proof, temperature controlled room: 1) Adaptation; 2) Baseline (BSL); 3) Recovery (REC). Every night, sleep recording

A VEGA 24 (Esaote Biomedica, Firenze) polygraph set at a paper speed of 10 mm/s was used for polygraphic recordings. EEG (C3-A2 and C4-A1) was recorded with an AC time constant of 0.3 seconds. Since MEMA can be inferred from variations in the compliance of the tympanic membrane or in pressure waves set up by movements of this membrane, this activity was recorded by a pressure-sensitive transducer (SenSym LX06001D) encased in silicone ear moulds fitted to the individual external auditory canals.27 The output of the transducer was routed directly to the polygraph's preamplifier. Sensitivity was adjusted so that the amplitude of the baseline pulse was 10 mm[3] and AC time constant was equal to 1 second. Bipolar horizontal and vertical eye movements were recorded with a time constant of 1 second. Bipolar horizontal EOG was recorded from electrodes placed about 1 cm from the medial and lateral canthi of the dominant eye, and bipolar vertical EOG from electrodes located about 3 cm above and below the right eye pupil. Equivalency between voltage and degree of visual angle was established through a calibration procedure utilizing REMs of known direction and amplitude. The calibrations were conducted in normal room illumination at the beginning of each recording session. Submental EMG was recorded with a time constant of

2. Subjects were selected if they usually went to sleep between 11:00 and 12:00pm and if they usually slept 7-8 hours per night [mean estimated duration of their own sleep was 472.5 min (s.d.=27.3)].

3. The rhythmically pulsating baseline signal recorded on the MEMA channels is presumably of vascular origin and it shows a coincidence with heart rate.21

MATERIALS AND METHODS Subjects Nine normal male subjects were selected as paid volunteers for the study (mean age=23.2 years). The requirements for inclusion were: normal sleep duration and schedule, no daytime nap habits, no excessive daytime sleepiness, no other sleep, medical, or psychiatric disorder, as assessed by a one-week sleep log[2] and by a clinical interview. Each subject was unaware of the purpose of the experiment and gave an informed consent; their rights were protected through the entire course of the experiment. Procedure

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REM frequency was expressed by the ratio: number of REMs /duration of REM sleep (in minutes). MEMA was also scored visually by two experimenters, who were blind with respect to the experimental condition; cases in which they were in disagreement were solved by a third blind scorer. MEMA was scored following the criteria proposed by Slegel et al.19 Any AC deflection was scored as MEMA when: a) its duration was greater than 0.3 sec; b) it exceeded twice the average baseline pulse amplitude; and c) there was no artifact. Any AC deflection within 0.3 sec after an EMG activation and/or with a concomitant activation of the strain gauge or of the microphone was considered an artefact. MEMA frequency was computed separately for NREM and REM sleep, as expressed by the ratio: number of valid MEMA deflections/duration of NREM or REM sleep (in minutes).

0.03 seconds as a standard measure of electromyographic activity during sleep. Because of the need to discriminate between endogenous MEMA and middle-ear variations evoked by motor activity (i.e., subvocalizations, mouth opening, swallowing, teeth clenching and head turning), supplemental bipolar EMGs with a time constant of 0.03 seconds were recorded by the right and left masseter muscles and by laryngeal surface electrodes. Electrode impedance was kept below 5 KOhms. Furthermore, we recorded each subject's head movements by means of a strain gauge connected to the ear mould, and monitored the sound level in the sleep room by means of a microphone placed at the level of the subject's head. Data Analysis Left central EEG (C3-A2), EMG, and horizontal and vertical EOG were used to visually score sleep stages, according to the standard criteria.28 With regard to SWS scoring, the amplitude criterion (>75 µV) was strictly followed. Horizontal and vertical EOG were scored visually by two experimenters, who were blind with respect to the experimental condition and with respect to the correspondence between upward/downward deflections of the pens and leftward/rightward or upward/downward direction of REMs. Cases of contemporary deflections of horizontal and vertical EOG (i.e., oblique rapid eye movements) were scored considering only the deflection with the greater amplitude, and cases in which the two scorers were in disagreement were solved by a third blind scorer. Any deflection of the pen equivalent to a REM greater than 3 deg and separated from another deflection by more than 200 msec was considered to be an independent REM. Minimal velocity to consider an eye movement as "rapid" was 50 deg/s.29 Blinks and other artefacts were excluded by visual analysis.

RESULTS Polysomnography Table 1 reports means, standard deviations, and ANOVA results of the sleep variables during baseline (BSL) and recovery (REC) nights. ANOVA results indicate that, as a result of sleep deprivation, recovery nights were characterized by a decrease of stage 1, stage 2, and intra-sleep waking; these decreases were paralleled by increases of SWS and of the sleep efficiency index, without affecting percentages of REM sleep. The whole latencies of NREM stages were significantly shortened. MEMA Frequency A two-way repeated measure ANOVA, 2x2, STAGE (NREM vs. REM sleep) x NIGHT (baseline vs. recovery) was carried out on the MEMA frequency. This analysis showed a significant main effect for the STAGE factor (F1,8=90.54; p=.00001), indicating a higher rate of MEMA during REM sleep (M=.72) as compared to NREM (M=.16) -Figure 1-. The main effect for NIGHT factor was not significant (BSL=.42, REC=.47; F1,8=.54, p=.48), and STAGE x NIGHT interaction was also not significant (F1,8=.01; p=.96). A further ANOVA was carried out on the MEMA frequency of REM sleep to assess whether the MEMA frequency after sleep deprivation could be modulated by the REM Cycle. The two-way repeated measure ANOVA, 4x2, REM Cycle (1st REMP vs. 2nd REMP vs. 3rd REMP vs. 4th REMP) x NIGHT (baseline vs. recovery) did not show any significant effect for REM Cycle factor (F3,24=1.24; p=.31)[4], for NIGHT factor (F1,8=.14; p=.72), and for REM Cycle x NIGHT interaction (F3,24=1.16; p=.34).

Table 1—Means and standard deviations (within brackets), and ANOVA results of the sleep variables (expressed as a function of Total Sleep Time) during baseline (BSL) and recovery (REC) nights. SWS= Slow-Wave Sleep (stages 3+4); ISW= Intra-Sleep Wake; SE= Sleep Efficiency index (Total Bed Time/Total Sleep Time x 100). Variables

BSL

REC

F(1,8)

p

% Stage 1 % Stage 2 % SWS % REM ISW (min) SE TST (min) S1 latency (min) S2 latency (min) SWS latency (min) REM latency (min)

8.21 (3.49) 56.99 (5.82) 10.94 (5.72) 23.86 (5.17) 17.33 (7.55) 92.54 (4.62) 449.61 (21.21) 16.67 (15.70) 19.11 (16.70) 45.94 (43.52) 85.83 (28.41)

4.47 (2.05) 51.07 (8.23) 22.12 (9.35) 22.34 (4.15) 9.33 (4.26) 97.24 (.87) 447.56 (17.76) 2.50 (1.84) 4.06 (2.43) 12.89 (6.27) 94.94 (52.63)

15.17 7.03 25.43 .60 5.88 9.33 .21 8.41 9.44 6.13 .35

.005 .03 .001 .46 .04 .02 .65 .02 .01 .04 .57

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4. Means as a function of REM CYCLE were: 1st REMP=.75, 2nd REMP=2.38, 3rdREMP=2.86, 4th REMP=.72.

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REM Frequency

Results of the polysomnographic data show that, in the recovery night, sleep deprivation was effective in determining an increase of SWS amount and of the sleep efficiency index, and a decrease of stage 1, stage 2, intra-sleep wake and NREM latencies, without affecting duration and latency of REM sleep. The main finding of the present study is that, in the recovery night after total sleep deprivation, there is an approximate 30% drop in REM frequency as compared to baseline nights (a 33% and 24% reduction respectively in

horizontal and vertical REM frequency). This result confirms previous observations,5-6 also suggesting a negative linear relationship between slow-wave sleep amount and REM frequency. The correlations between these two variables approached significance in the present sleep deprivation study, and they were significant after selective SWS deprivation.16 However, this evidence needs to be strengthened by a more direct evaluation by means of a correlation between slow-wave activity and REM frequency. With regard to the issue of the higher rate of vertical REMs as compared to horizontal ones, it is in line with the common knowledge of individual differences in REM frequency30 and in the preferred direction of REMs.31, 32 At variance with eye movements, middle ear muscle activity during REM sleep does not show any significant variation (MEMA frequency was equal to .70 and .75, respectively in the baseline and recovery nights), suggesting that sleep deprivation does not affect middle ear muscle activity. This negative finding also weakens the hypothesis of a unique centre of phasic motor activity during REM sleep: the decrease of REM frequency after a one-night total sleep deprivation is not paralleled by a concomitant decrease of phasic activity of middle-ear muscles. On the other hand, the present lack of a psychophysiological parallelism between muscle phasic events of REM sleep is not unique. In fact, there are at least two other phenomena for which REM frequency shows systematic variations not paralleled by concomitant variations in MEMA frequency: a) MEMA frequency during REM sleep, at variance with rapid eye movements, does not show the increase across sleep cycles;33 b) depressed patients show no difference in MEMA frequency as compared to normal subjects,33 while they show an increased REM frequency.34 A further evaluation of the hypothesis of a decrease—

Figure 1—Mean MEMA frequency and standard deviations recorded in REM and NREM sleep of baseline and recovery nights

Figure 2—Mean REM frequency and standard deviations recorded by horizontal and vertical EOG during baseline and recovery nights

A two-way repeated measure ANOVA, 2x2, CHANNEL (horizontal vs. vertical) x NIGHT (baseline vs. recovery) was carried out on the REM frequency. This analysis showed a significant main effect for the CHANNEL factor (F1,8=14.15; p=.005), indicating a higher rate of vertical REMs (M=16.52) as compared to horizontal ones (M=14.02), and a significant main effect for NIGHT factor (F1,8=23.79; p=.001) with the hypothesized decrease of REM frequency during the REC night (M=12.89) as compared to the BSL (M=17.65). The interaction CHANNEL x NIGHT was not significant (F1,8=.47; p=.51). Figure 2 reports means and standard deviations of this analysis. Since REM frequency has been found to be inversely correlated with SWS amount during the recovery nights after SWS deprivation,16 the relationship between REM frequency and SWS duration was also evaluated in the present study. During the recovery night after total sleep deprivation, correlations only approached statistical significance for either horizontal (r=-.65; p=.06) or vertical REMs (r=.60; p=.08). DISCUSSION

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