Time of day is associated with paradoxical reductions in global signal fluctuation and functional connectivity

The brain exhibits substantial diurnal variation in physiology and function, but neuroscience studies rarely report or consider the effects of time of day. Here, we examined variation in resting-state functional MRI (fMRI) in around 900 individuals scanned between 8 AM and 10 PM on two different days. Multiple studies across animals and humans have demonstrated that the brain’s global signal (GS) amplitude (henceforth referred to as “fluctuation”) increases with decreased arousal. Thus, in accord with known circadian variation in arousal, we hypothesised that GS fluctuation would be lowest in the morning, increase in the midafternoon, and dip in the early evening. Instead, we observed a cumulative decrease in GS fluctuation as the day progressed. Although respiratory variation also decreased with time of day, control analyses suggested that this did not account for the reduction in GS fluctuation. Finally, time of day was associated with marked decreases in resting-state functional connectivity across the whole brain. The magnitude of decrease was significantly stronger than associations between functional connectivity and behaviour (e.g., fluid intelligence). These findings reveal time of day effects on global brain activity that are not easily explained by expected arousal state or physiological artefacts. We conclude by discussing potential mechanisms for the observed diurnal variation in resting brain activity and the importance of accounting for time of day in future studies.


Reviewer #1
R1Q1 Matthew F. Glasser: The authors have addressed my prior concerns. I would request a minor tweak of the future section to read "There are many avenues to extend the current study. For example, it will be interesting to explore whether the same effects can be seen during task-fMRI. In addition, Glasser and colleagues proposed the use of temporal ICA (Glasser et al., 2018) to decompose the fMRI data into multiple components, some of which appear to reflect "global" artefacts, which can then be more selectively removed, and some of which may relate to neural signals for arousal or eyes open versus closed. It would be interesting to investigate how these distinct global components might relate to time of day. Furthermore, some of these "global" components are present only during resting-fMRI, but not task-fMRI (Glasser et al., 2018). Thus, some of the effects we observe in this study might not appear in task-fMRI." We have updated the following passage as requested.

Section 4.11 of Discussion
There are many avenues to extend the current study. For example, it will be interesting to explore whether the same effects can be seen during task-fMRI. In addition, Glasser and colleagues proposed the use of temporal ICA (Glasser et al., 2018) to decompose the fMRI data into multiple components, some of which appear to reflect "global" artefacts, which can then be more selectively removed and some of which may relate to neural signals for arousal or eyes open versus closed. It would be interesting to investigate how these distinct "global" components might relate to time of day. Furthermore, some "global" components are present only during resting-fMRI, but not task-fMRI (Glasser et al., 2018). Thus, some of the effects we observe in this study might not appear in task-fMRI.
We have also added a URL where the list of subjects who passed our QC of the physiological data can be freely accessed.

Section 5.4.2 of Materials and Methods
The list of subjects who passed visual quality screening of their pulse and respiratory data is publicly available at the GitHub repository maintained by the Computational Brain Imaging Group (https://github.com/ThomasYeoLab/CBIG/tree/master/stable_projects/preprocessing/ Orban2020_tod/data_release).

Reviewer #3
The authors have made substantial revisions that largely address the prior concerns. I just have two remaining minor comments.
R3Q1: In the abstract, the authors now state: "These findings reveal unexpected effects of time of day on global brain activity that are not easily explained by arousal or physiological artefacts." However, I think this is still misleading as the authors did not actually make any measurements of arousal state. I think what they mean to say is the "expected arousal" state. As noted in the prior comments and as acknowledged by the authors in the revised work, the actual arousal state may have differed from the expected arousal state due to a number of factors.
We agree with the reviewer's point and have further refined this sentence in the abstract as suggested.
Abstract: These findings reveal time of day effects on global brain activity that are not easily explained by expected arousal state or physiological artefacts. Figure S1, please add a colorbar to describe the color scheme used.

R3Q2: For
We have now added colourbars to Figure S1. High density of data points around 12:30 pm is consistent with the planned timing of resting state scans based on the HCP study protocol (HCP Reference Manual -1200 Subjects Release; Page 33). These same results are presented and described in more detail in Fig 1, 3, S3, S4, S8 and S9 without colour-coding of data point density.