Sliding windows analysis can undo the effects of preprocessing when applied to fMRI data

Resting-state fMRI (rs-fMRI) data is used to study the intrinsic functional connectivity (FC) in the human brain. Recently, interest has focused on studying the temporal dynamics of FC on short timescales, ranging from seconds to minutes. These studies of time-varying FC (TVFC) have enabled the classification of whole-brain dynamic FC profiles into distinct “brain states”, defined as recurring whole-brain connectivity profiles reliably observed across subjects and sessions. The analysis of rs-fMRI data is complicated by the fact that the measured BOLD signal consists of changes induced by neuronal activation, as well as non-neuronal nuisance fluctuations that should be removed prior to further analysis. Thus, the data undergoes significant preprocesing prior to analysis. In previous work [19], we illustrated the potential pitfalls involved with using modular preprocessing pipelines, showing how later preprocessing steps can reintroduce signal variation previously removed from the data. Here we show that the problem runs deeper, and that certain statistical analysis techniques can similarly interact with preprocessing and reintroduce previously removed signal variation. One such technique is the popular sliding window analysis, used to compute TVFC. In this paper, we discuss the problem both theoretically and empirically in application to test-retest rs-fMRI data. Importantly, we show that we are able to obtain essentially the same brain states and state transitions when analyzing motion induced signal as we do when analyzing the preprocessed but windowed data. Our results cast doubt on whether the estimated brain states obtained using sliding window analysis are neuronal in nature, or simply reflect non-neuronal nuisance signal variation (e.g., motion). We highlight the need to critically revisit previous work on rs-fMRI data work that may not have adequately controlled for these types of effects.