163 Axonal Loss in Narcolepsy Type 1 The Independent Review Board of the Vrije University Medical Center (registered with the US office of Human Research Protections as number 00002991 under Federal wide Assurance number 00003703) approved the procedures of the Netherlands Brain Bank concerning “Donation of brain material for scientific research” under reference number 2009/148. Informed consent for the autopsy, and use of brain material and clinical data for research purposes was obtained from the donors or next of kin. Tissue dissection and regions of interest Locations included the brainstem which is the most heavily affected brain region in previous DTI studies,[84, 104-107, 109, 238], the anterior corpus callosum as it is a dense white matter tract where DTI studies reported profoundly lower fractional anisotropy [105-107, 238], white matter regions adjacent to the anterior cingulate and occipital cortices to also include multiple white matter regions in proximity to the cortex where DTI differences have been reported [84, 105-107, 238], and the vermis as a control region in which almost no DTI differences have been reported [84, 106, 109, 110, 238]. The same anatomical regions were selected in different subjects using the following anatomical landmarks during the dissection process: transversal sections from the midbrain were included at the level of the superior colliculus and included the substantia nigra, red nucleus, reticular formation and pyramidal tract. The proximal and distal corpus callosum and the ventral anterior cingulate cortex were identified from the same tissue block using slices perpendicular to and just caudal from the genu of the corpus callosum. Deep white matter below the cingulate sulcus was identified using a three mm straight line from the origin of the cingulate sulcus. The transversal occipital sections had to include the primary visual (V1) and secondary visual (V2) cortices, where V1 was recognized by the band of Gennari. The dentate nucleus was used as a landmark in the sagittal cerebellum sections. Nine regions of interest (ROIs) were drawn per subject (Table 1, and Figure 1 for examples) using QuPath (version 0.4.3). Circular ROIs with a perimeter of 5000 µm were used for all ROIs except for the occipital lobe. The white matter within gyri in the occipital lobe was generally too narrow for a ROI with this perimeter and three smaller circular ROIs were drawn separately for V1 and V2. Irregularities such as tissue tears, folds and unfocused areas were removed from the ROIs. Anatomical locations of ROIs were visually inspected for correspondence within subjects across stainings, and across different subjects. ROIs were drawn blinded by one researcher (JKG). One anterior 6
RkJQdWJsaXNoZXIy MjY0ODMw