Slow cortico-cortical connectivity (2-5Hz) is a new robust signature of conscious states

While long-range cortico-cortical functional connectivity has been reported by several studies as a necessary condition of conscious state, precise empirical evidence is still scarce. In the present work we provide such a direct and conclusive evidence in a set of three experiments. In the two first experiments intracranial-EEG was recorded during four distinct states in the same individuals: conscious wakefulness (CW), rapid-eye-movement sleep (REM), stable periods of slow-wave sleep (SWS) and deep propofol anaesthesia (PA). We discovered that long-range FC, computed by the weighted Symbolic-Mutual-Information (wSMI) in the 2-5Hz frequency band was a specific marker of conscious states that could discriminate CW and REM from SWS and PA. In the third experiment, we generalized this original finding on a large cohort of brain-injured patients by revealing that wSMI in the 2-5 Hz range was also able to accurately discriminate patients in the vegetative state (or unresponsive wakefulness syndrome) from patients in the minimally conscious state. Taken together the present results suggest that 2-5Hz FC is a new and robust signature of conscious states.

In order to address these important and unsolved questions, we explored cortico-cortical 58 connectivity on direct intracranial EEG (iEEG) recordings performed in the same subjects 59 across various conscious and unconscious states. More precisely, we recorded iEEG in 12 60 drug-resistant epileptic patients undergoing a stereoelectroencephalography (SEEG) for pre-61 surgical evaluation with large and brain-scale implantations. Each patient was recorded in two 62 3 states that are associated with conscious experience (conscious wakefulness (CW) and rapid-63 eye movement sleep (REM) that is typically associated with conscious dreaming), as well as 64 in two unconscious states: very stable periods of slow-wave sleep (SWS) that are usually free 65 of conscious dreaming (Siclari et al., 2017), and deep propofol anaesthesia (PA). 66 We estimated cortico-cortical connectivity by computing the weighted symbolic mutual 67 information (wSMI) in various frequency bands. We previously conceived wSMI and used 68 this measure to distinguish conscious and minimally conscious (MCS) patients from 69 vegetative state/unresponsive wakefulness syndrome (VS/UWS) patients (King et al., 2013).  (Corazzol et al., 2017). In these two studies, wSMI 77 computed in the 4-10Hz (=32ms) distinguished MCS from VS/UWS states, while wSMI 78 computed in higher frequencies failed to do so. 79 We conducted two consecutive SEEG studies: in study I, five patients were recorded during 4 80 nights in three stages (CW, REM, SWS), whereas in study II, 7 additional patients were 81 recorded during short periods of 10 minutes in four stages (CW, REM, SWS and PA). Finally, 82 we conducted a third study using high-density scalp EEG on 145 patients suffering from 83 disorder of consciousness (MCS and VS/UWS) (Naccache, 2018), in order to determine if our 84 findings would discriminate between these two states. wSMI 4-10Hz was higher during REM than during SWS in only 5 out of 12 patients, while 99 the 7 remaining patients showed an opposite significant pattern. Finally, only 2 out of 7 100 patients showed larger wSMI 4-10Hz in REM than in PA, while the five remaining ones also 101 showed the reverse pattern (see Figure 1a and Figure S2b). 102 We then discovered that the wSMI calculated on slower frequencies (wSMI 2-5Hz; =64ms) 103 actually succeeded much better to discriminate robustly the two conscious states from the two 104 unconscious ones: 10 out of 12 patients showed significant larger mean values during CW 105 than during SWS, while the remaining two patients (who both had short recordings) did not 106 show significant differences across these two states. The very same result was found when 107 comparing REM to SWS. A mirror observation was made in the 8-20 Hz (=16ms) and in the 108 32-80Hz (=4ms) where mean wSMI was significantly larger (p<0.001) in the SWS as 109 compared to CW and REM in all patients (Figure 1 b). Concerning PA, all seven patients had 110 a larger wSMI 2-5Hz during CW than PA, and during REM than PA (see Figure 1b and S2c).

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As a control test, we checked that wSMI computed in a slower frequency band (1-2.5Hz; 112 =128ms) did not perform well to discriminate conscious from unconscious states (Fig 1a and 113 S1c). 114 We also noticed that PA, unlike SWS, was associated with a massive, diffuse and systematic 115 increase of functional connectivity in high frequencies (32-80Hz (=2ms), see Figure 1b

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In the present iEEG study, we first showed that long-range cortico-cortical connectivity 138 measured with the wSMI computed in the theta-alpha band (4-10Hz), -that was previously 139 shown to be higher for patients in the MCS from those in the VS/UWS -, was larger during 140 CW than during SWS. However, and unexpectedly, this measure failed to discriminate both 141 REM from SWS, and REM and CW from PA. In other terms, FC in the 4-10Hz does not 142 seem to be a general signature of conscious states. We then discovered that FC computed in a 143 slower delta-theta band (2-5Hz) actually succeeded much better to discriminate correctly 144 conscious states (CW and REM) from the two unconscious states investigated here (SWS and 145 PA). This measure was found larger in CW and in REM as compared to SWS and to PA in 146 the vast majority of patients. Notably, none of the 12 recorded patients showed a reverse 147 pattern of connectivity between conscious and unconscious states in this delta-theta band.

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In the light of these results and of previous reports, we propose a two-frequency hypothesis of 170 cortical connectivity that introduces a distinction between signatures of conscious state and 171 signatures of conscious access.

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In two previous SEEG studies, we identified brain-scale increases of connectivity in the  conscious GW. This last measure seems more specific than the proposed signature of 189 conscious access, but the reasons of this lack of specificity require additional studies. 190 Finally, we will also discuss our findings related to increase of FC in the gamma-band during 191 unconscious states. Recently, loss of consciousness has also been related to hyper-correlated

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First and second experiment 209 The two first experiments of this study used a very similar approach. We therefore report their 210 respective experimental procedures in a single section.       exceeded a 100 μv peak-to-peak amplitude in more than 50% of the epochs were rejected.

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Channels that exceeded a z-score of 4 across all the channels mean variance were rejected.

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This step was repeated two times. Epochs that exceeded a 100 μv peak-to-peak amplitude in 366 more than 10% of the channels were rejected. Channels that exceeded a z-score of 4 across all 367 the channels mean variance (filtered with a high pass of 25 Hz) were rejected. This step was 368 repeated two times. The remaining epochs were digitally transformed to an average reference.

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Rejected channels were interpolated. Finally, EEG were deemed to pass this preprocessing 370 step if at least 70% of the channels and at least 30% of the epochs were kept. Where z is the z-statistic of the Mann-Whitney U test and N the size of the population.

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This effect size measure was computed both at the level of the wSMI mean , but also at the level 404 of the cluster (effect size computed on the mean over electrodes belonging to the cluster).