Subjects:
Thirty-nine subjects with migraine (29 females; mean ± SEM age, 29.97 ± 1.55 years) and 39 pain-free controls (23 females; mean ± SEM age 30.70 ± 2.01 years) were recruited from the general population using an advertisement. There were no significant differences in age (t-test, p>0.05) or gender composition (χ2 test, p>0.05) between the control and migraine group. Migraine subjects were diagnosed according to the International Classification of Headache Disorders (ICHD), 3rd edition, sections 1.1 and 1.2 (15). Ten migraineurs reported aura associated with their migraines and the remaining 29 reported no aura. All migraine subjects were scanned during the interictal period (the pain- and symptom- free period between migraine attacks), at least 72 hours after and 24 hours prior (subsequently verified with a headache diary) to a migraine event.
The exclusion criteria for migraineurs included the presence of any other pain condition or neurological disorder. Controls were exempt from the study if they had a family history of migraines, currently used analgesic medications or if they suffered from any other pain condition or neurological disorder. All participants were subject to standard MRI exclusion criteria. All migraine subjects indicated the pain intensity during their most recent migraine (6-point visual analogue scale; 0 = no pain, 5 = most intense imaginable pain) and specified the distribution of their pain commonly experienced during a migraine attack on a face map. Furthermore, each subject described the qualities of their migraine pain as well as any current treatments and medications used to prevent or abort a migraine once initiated. This study was approved by the Institutional Human Research Ethics Committee at the University of Sydney and informed written consent was obtained for all participants in accordance with the Declaration of Helsinki. Data from 30 of the 39 migraineurs were used in previous investigations (16-19).
MRI acquisition:
Subjects lay supine on the bed of a 3-T MRI scanner (Phillips, Achieva) with their head immobilized in a 32-channel head coil. In each subject a high-resolution 3D T1-weighted anatomical image set covering the entire brain was collected (turbo field echo; echo time = 2.5ms, repetition time = 5600ms, flip angle 8⁰, voxel size 0.8 x 0.8 x 0.8 mm). In addition, a high-resolution diffusion tensor image (DTI) image set covering the entire brain was collected using a single-shot, multi section, spin-echo echo-planar pulse sequence (repetition time = 8788 ms; flip angle = 90⁰, matrix size 112 x 112, field of view 224 x 224 mm, slice thickness = 2.5 mm, 55 axial slices). For each slice, diffusion gradients were applied along 32 independent orientations with b = 1000 s/mm2 after the acquisition of b = 0 s/mm2 (b0) images. The b0 value reflected the strength and timing of the gradients used to generate DTI; the high b value of 1000 s/mm2 generated images with stronger gradients and faster slew rates.
MRI analysis:
Trigeminal nerve volume and maximum cross-sectional area analysis:
Using Statistical Parametric Mapping (SPM) version 12 software (20), the T1-weighted anatomical image from each subject was resampled at a higher resolution of 0.3x0.3x0.3 mm in order to improve visualization of the trigeminal nerve and to reduce the inclusion of surrounding tissue and cerebrospinal fluid. Using the resampled images, the left and right trigeminal nerves within the root entry zone were isolated in each subject. The root entry zone is the section of the trigeminal nerve that lies within the pontine cistern, i.e. from where it emerges from the pons, to the point at which it exits the pontine cistern anteriorly (Figure 1).
All three orthogonal planes were used in defining the nerve on both sides, with the axial plane being the first plane used, followed by coronal and sagittal views. A volume of interest encompassing the entire trigeminal root entry zone was created on each subject’s T1-weighted image. The total volumes (mm3) within the isolated nerves were calculated by extracting and averaging the volume from each voxel inside the isolated region of the trigeminal root entry zone in the left and right nerve for each individual control and migraine subject. Furthermore, the cross-sectional volume of the nerve in each coronal slice was calculated and the maximum coronal cross-sectional area value (mm2) was determined. For both volume and cross-sectional area, the mean of the left and right trigeminal nerves was calculated, resulting in a single value for each measurement in each control and migraine subject.
For the control and migraine groups, mean (±SEM) volume and maximum cross-sectional areas were calculated. Significant differences in volume and cross-sectional areas between control and migraine groups were determined using unpaired t-tests (one-tailed, p<0.05). In addition, linear relationships between volume and cross-sectional areas in migraineurs with migraine duration and migraine intensity were determined (Pearson’s correlation, p<0.05).
Trigeminal nerve diffusion analysis:
All diffusion tensor image sets from each subject were motion corrected, based on b0 images within each series. Using diffusion-weighted images collected from 32 directions and b0 images, the diffusion tensor was calculated from all the images using a linear model (21). Once the elements of diffusion tensor were calculated, whole-brain maps of fractional anisotropy (FA), mean diffusivity (MD), axial diffusion (AX) and radial diffusion (RD) were calculated. All images remained in native space for these calculations. The DTI images were shifted to the subject’s anatomical space by coregistering the b0 DTI image to the T1-weighted image.
The diffusion images were then resampled to a voxel size of 0.3x0.3x0.3 mm. Using these resampled images, the left and right trigeminal nerves within the root entry zone were isolated as aforementioned. Using the FA image, all three orthogonal planes were used in defining the nerve and a volume of interest encompassing the entire trigeminal root entry zone was created for each subject. The mean (±SEM) FA, MD, AX and RD values were calculated for the left and right trigeminal nerves in all subjects. For all four diffusion measures, the mean of the left and right trigeminal nerves was calculated, resulting in a single value for each measurement in each control and migraine subject.
For the control and migraine groups, mean (±SEM) FA, MD, AX and RD values were calculated. Significant differences in all four diffusivity values between control and migraine groups were determined using unpaired t-tests (one-tailed, p<0.05). In addition, linear relationships between diffusivity values in migraineurs with migraine duration and migraine intensity were determined (Pearson’s correlation, p<0.05).
To study nerve fiber demography, the trigeminal root entry zone was divided into caudal, middle and rostral thirds (Figure 1). FA, MD, AX and RD values were then calculated for each of these three regions for the left and right nerves in each control and migraine subject. For all four diffusion measures, the mean of the left and right trigeminal nerves was calculated, resulting in a single value for each measurement in each control and migraine subject at each of the caudal, middle and rostral parts. For the control and migraine groups, mean (±SEM) FA, MD, AX and RD values were calculated at each level. Significant differences in diffusion values between groups were determined using unpaired t-tests (one-tailed, p<0.05).