Differential Effect of Global Signal Regression Between Awake and Anesthetized Conditions in Mice

Introduction: In resting-state functional magnetic resonance imaging (rs-fMRI) studies, global signal regression (GSR) is a controversial preprocessing strategy. It effectively eliminates global noise driven by motion and respiration but also can introduce artifacts and remove functionally relevant metabolic information. Most preclinical rs-fMRI studies are performed in anesthetized animals, and anesthesia will alter both metabolic and neuronal activity.Methods: In this study, we explored the effect of GSR on rs-fMRI data collected under anesthetized and awake state in mice (n = 12). We measured global signal amplitude, and also functional connectivity (FC), functional connectivity density (FCD) maps, and brain modularity, all commonly used data-driven analysis methods to quantify connectivity patterns.Results: We found that global signal amplitude was similar between the awake and anesthetized states. However, GSR had a different impact on connectivity networks and brain modularity changes between states. We demonstrated that GSR had a more prominent impact on the anesthetized state, with a greater decrease in functional connectivity and increased brain modularity. We classified mice using the change in amplitude of brain modularity coefficient (Delta Q) before and after GSR processing. The results revealed that, when compared with the largest Delta Q group, the smallest Delta Q group had increased FCD in the cortex region in both the awake and anesthetized states. This suggests differences in individual mice may affect how GSR differentially affects awake versus anesthetized functional connectivity.Discussion: This study suggests that, for rs-fMRI studies which compare different physiological states, researchers should use GSR processing with caution. Impact statement Global signal regression (GSR) is a commonly used denoising technique, however, it can also introduce specific artifacts. In functional connectivity (FC) studies of functional magnetic resonance imaging, it is crucial to examine the impact of GSR on animals under various anesthesia conditions. Our results showed that GSR had distinct effects on FC and modularity of awake versus anesthetized mice. Therefore, caution should be taken when using GSR to remove noise, especially when comparing different neurometabolic states.