fMRI Experimental design and data acquisition
The fMRI scanning procedure was conducted at Korea Advanced Institute of Science and Technology (KAIST), South Korea. 24 healthy subjects (15 males, 9 females, mean age 21, SD 0.8), with normal color vision, were enrolled and scanned in the study. Each subject was exposed to a procedure comprising alternating rest and task conditions while being examined by an fMRI scanner. The data obtained was preprocessed to identify and filter out datasets which contained head movement induced motion blur, background noise or low quality measurements. 4 subjects which produced the highest quality datasets were selected for onward methodological study and analysis.
Every experimental session lasted for 160 s and consisted of 8 blocks with each rest block leading a stimulus block. The duration of each rest and stimulus block was for 24 and 16 s, respectively. Within a single scanning session, 80 volume scans (32 with stimuli and 48 at rest) were obtained at intervals of 2 s. We applied cluster analysis on the resulting 80 data points corresponding to average activations in BA17, 18 and 19 voxels for the classification amongst HCC, HCF, RTC and RTX.
Four visual stimuli of different shapes and sizes were designed towards evaluating their potential hallucinogenic impact on the visual cortex. HCC comprised of concentric circles with varying diameters and colors (shades of grey). These circles were then set into synchronous rotation about the center point. RTC contained three static concentric circles with checkered boundaries and their center point indicated by a black spot. HCF pattern was a four-winged fan rotating about its center while the RTX was a stationary cross drawn using checkered lines (Fig. 1). Of the four sessions, each session was confined to a single visual stimulus. The experimental sessions were designed such that RTX was presented first followed by HCF, RTC and HCC respectively. A scan was acquired every 2 s while the stimulus was being shown. This pattern was repeated for the remaining three sessions as well. During each scan, the subjects were directed to continually focus on the presented stimulus and encouraged to keep their minds relaxed during the rest phase. To ensure high quality data from scanning procedure, their heads were placed in a brace and adjusted before a scan was performed.
Image acquisition
The images were acquired using a 3 Tesla (FORTE, Oxford magnet, Varian Console, built up by ISOL) instrument, with a quadrature head coil to get an anatomic scan and a surface coil to obtain the functional scan. High-resolution anatomic images (structural resolution 1.25 mm isotropic voxels) were acquired using an MPRAGE sequence (echo time TE = 3.7 ms, TR = 8.1 ms, flip angle = 8°, FOV = 256 × 256 mm) and functional data were acquired using echo planar imaging (EPI, TE = 37 ms, phase encoding = top to bottom, flip angle = 70°, TR = 2000 ms, matrix = 128 × 128 mm, slices = 15, voxels = 3 mm × 3 mm, no gap) as shown Fig. 1.
Image analysis
Data was analyzed using Statistical Parametric Mapping software (SPM8b; Wellcome Department of Cognitive Neurology, University College London, London, UK). Images realignment was performed to correct for the artifacts due to minor head vibrations and normalized to a standard Montreal Neurological Institute (MNI) template. Smoothing was done by 4-mm full width at half maximum smoothing to average the data with the neighboring data points. Images were analyzed using contrast vector C = [1 −1] corresponding to p < 0.05 [Family Wise Error (FWE) correction] (Fig. 3).
fMRI images configuration
Grey-scale fMRI images were used for onward investigations, with the darker regions having a higher pixel value while the lighter regions approaching to a zero on the pixel scale. A total of 80 scans were taken for each session so as to ascertain accuracy in the ensuing statistical analysis of these results. All volume scans were cut down into 15 slices. The dimension of a single volume was 128 × 128 × 15. The total number of voxels in a volume counted to 245,760. Each voxel in the study had a uniform size of 3 × 3 × 3 mm.
Statistical methods and techniques for fMRI data analysis
To test the variation and significance of data obtained after presentation of visual stimuli, analysis of variance (ANOVA) test [23] was employed. Upon ascertaining significance of impact on visual cortex, LSD [24] was applied to determine the individual contribution of each stimulus on the visual cortex. For classifying the mixed cortical activations into clusters, K-means clustering [25] was applied to the activation data. To compute the probabilistic relationship between each visual stimuli and BAs 17, 18 and 19, we used multinomial logistic regression (MLR) [26].