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Abstract
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Serotonin Transporter Genetic Variation and the Response of the Human Amygdala
Ahmad R. Hariri, Venkata S. Mattay, Alessandro Tessitore, Bhaskar Kolachana, Francesco Fera, David Goldman, Michael F. Egan, and Daniel R. Weinberger

Supporting Online Material

Materials and Methods

DNA isolation and analysis. DNA isolation and analysis was conducted on blood samples obtained from all subjects who gave informed consent according to NIMH IRB guidelines. Oligonucleotide primers flanking the 5-HTTLPR and corresponding to the nucleotide positions -1416 to -1397 (stpr5, 5' -GGCGTTGCCGCTCTGAATGC) and -910 to -888 (stpr3, 5' -GAGGGACTGAGCTGGACAACCAC) of the 5-HTT gene 5' -flanking regulatory region were used to generate 484- and 528-bp fragments. PCR amplification was accomplished using the methods of Heils et al. (S1). Based on this assay, individuals possessing either one (n = 8; 4 in each cohort) or two copies (n = 6; 3 in each cohort) of the s allele were included in the s group, and those homozygous for the l allele (n = 14; 7 in each cohort) were included in the l group.

Analysis of imaging data. Each subject was scanned using a GE Signa 3T scanner with a real-time functional imaging upgrade (Milwaukee, WI). An automated shim procedure was applied to minimize possible magnetic field inhomogeneities. Blood oxygenation-level dependent (BOLD) functional images were acquired with a gradient echo EPI sequence, and covered 24 axial slices (4 mm thick, 1 mm gap) that began at the cerebral vertex and encompassed the entire cerebrum and the majority of the cerebellum (TR/TE = 2000/28 msec, FOV = 24 cm, matrix = 64 x 64). Whole-brain image analysis was completed using SPM99 (http://www.fil.ion.ucl.ac.uk/spm). Images for each subject were realigned to the first volume in the time series to correct for head motion, spatially normalized into a standard stereotactic space (Montreal Neurological Institute template) using a 12 parameter affine model and smoothed to minimize noise and residual differences in gyral anatomy with a Gaussian filter, set at 8 mm full-width at half-maximum, producing an effective spatial resolution of 11.4 x 11.5 x 10.9 mm. Voxel-wise signal intensities were ratio normalized to the whole-brain global mean. Predetermined condition effects at each voxel were calculated using a t-statistic, producing a statistical image for the contrast of the emotion task versus the sensorimotor control for each subject. These individual contrast images were then used in second-level random effects models, that account for both scan-to-scan and subject-to-subject variability, to determine task-specific regional responses at the group-level with one-sample (main effects of task) and paired t-tests (direct comparisons). Because of our strong a priori hypothesis regarding the differential response of the amygdala and our use of a rigorous statistical model, a statistical threshold of P < 0.05, with a small volume correction for multiple comparisons, was used to identify significant responses for all comparisons.

Reference

S1. A. Heils et al., J. Neurochem.66, 2621 (1996).