pCASL Analysis

Calculation of rCBF was done using the FSL software package (https://fsl.fmrib.ox.ac.uk/fsldownloads_registration). First, the “Brain extraction Tool” was used to remove non-brain tissue from a T1-weighted image. Second, the pCASL data obtained before, during, and after ketamine infusion were co-registered with the skull-stripped T1-weighted image, and, lastly, the T1-weigthed image was nonlinearly co-registered to Montreal Neurological Institute (MNI) space and the combined transformation was applied to the rCBF maps.

The effects of S-ketamine on rCBF were investigated with both voxel-based and region of interest (ROI) analyses. Voxel-based analysis identified areas with the most significant increase of rCBF using an analysis of variance (ANOVA) model and permutation-based statistical inference. We further used threshold-free cluster enhancement to account for spatial dependencies. The statistical maps were thresholded at p < 0.05 and corrected for multiple comparisons (FWE correction). ROI analyses were performed in two defined cortical areas with the first corresponding to the position of the MRS voxel in ACC and the second to the region with most significant voxel-based changes of rCBF, and in the subcortical regions provided by the MNI atlas from FSL (left and right Thal, caudate, accumbens, and putamen). The rCBF was calculated as both absolute values in mL/100 g/min and as normalised values by dividing each voxel with the global mean for each subject to reduce inter-subject variation caused by a difference in global rCBF. Normalised rCBF was used as the primary outcome to enhance the sensitivity to regional changes, but absolute rCBF values are reported as well.