Local differences on a nodal level in 6-OHDA rats compared to shams.

<p>Glass brains depicting nodes that showed significantly altered (A) node degree, (B) clustering coefficient and (C) node efficiency in 6-OHDA rats compared to shams. Nodes are represented as spheres and located in the glass brain according to their corresponding anatomical region’s centre mass. Blue spheres depict significantly lower, red spheres significantly higher values in 6-OHDA rats compared to shams. Significance was assessed using two-sample t-test by comparing the AUC of these network measures for each node across all network sparsities (0.05</p

Connection-wise comparison reveals significantly reduced connections in 6-OHDA rats compared shams.

<p>Glass brain depicting edges and their associated nodes that showed significantly reduced functional connectivity in 6-OHDA rats in comparison to sham rats. The size of a node represents the number of abnormal connections associated with that node. Significance was assessed using non-parametric permutation tests (N = 10000). A p-value below 0.05 was considered significant (FWER corrected). The results shown were generated at a primary NBS threshold of t = 3.1 with p = 0.0425, FWER corrected. 6-OHDA rats N = 13; sham rats N = 11. Bold L marks the lesion side. R, contralateral side.</p

Rat brain parcellation scheme showing 150 ROIs (75 on each hemisphere).

<p>Each bilateral ROI is labelled differently and overlaid onto coronal slices of the rat brain template.</p

Global network measures of segregation and integration across different network sparsities in 6-OHDA rats and shams.

<p>(A) Mean clustering coefficient C normalized to a random reference network. (B) Mean characteristic path length L normalized to a random reference network. (C) Small-worldness S. (D) Number of modules. (E) Modularity Q. (F) Global efficiency. Data are expressed as mean ± SEM. 6-OHDA rats N = 13; sham rats N = 11.</p

Tensor-based morphometry (TBM) of T₂-weighted images to contrast anatomical changes in MPTP-treated marmosets.

<p>Color scales are for volume difference (green), the raw t-statistical value at each voxel and the direction of the change (warm colors = expansion, cold colors = atrophy). Volume changes which survive multiple comparisons corrections across all voxels in the brain (False Discovery Rate with q<0.05) are also shown. <b>A.</b> 6 degrees of freedom (dof) TBM reveals local changes, but does not control for brain size differences. The t maps reveal sub-regional changes in the MPTP-treated animals compared to controls. The directionality map indicates that regional increases are mostly reflected in the ventricular system or the space between the cerebellum and spinal cord. Atrophy is seens mostly in the motor, temporal and parietal cortex, but little change in observed in sub-cortical structures on these comparisons. None of these changes survive a FDR correction for multiple statistical comparisons potentially due to variability and insufficient power. One source of variation that can dramatically affect sub-regional detection of changes is the difference in brain size due to animals of different ages being included here. <b>B.</b> Accounting for 9 dof, TBM can reveal sub-region-specific changes that are corrected for global changes (i.e. differences in brain volume). Indeed, this correction produces more consistent effects that survive FDR. Still, cortical areas show a clearer pattern of atrophy compared to sub-cortical regions, with only very subtle effects evident in the caudate or putamen. (Ctx = Cortex, Ento = Entorhinal, Temp = temporal; SMC = somatosensory cortex, SN = substantia nigra, VTA = ventral tegmental area, Hypothal = hypothalamus).</p

Animal characteristics.

<p>Animal characteristics.</p

Definition and delineation of regions of interests on T₂-weighted MRI scans.

<p>To control for overall age and growth differences between animals, total skull volume was measured, as this is considered an independent measurement that is not affected by changes in brain tissue and allows a contrast with total brain volume. Using the marmoset atlas [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0180733#pone.0180733.ref049" target="_blank">49</a>], the caudate, the putamen, the substantia nigra (SN), and hippocampus were delineated. Cortical thickness was evaluated in the motor, parietal and temporal cortex (Ctx) using five measurements each. For relative signal measurements, a ROI in the visual cortex (purple square) served as internal control.</p

Delineation and distinction of anatomical structures on T₂-weighted MR images.

<p>The high spatial resolution used here (0.0875 mm³/voxel) afforded the clear delineation of anatomical regions in the midbrain, such as the deep mesencephalic nucleus, red nucleus, ventral tegmental area, rostral subnucleus. subthalamic nucleus, as well as the substantia nigra. However, the lack of well-defined signal differences between these did not allow separate volumetric quantification of these. The putamen in contrast could be reliable delineated against surround structures, such as the external globus pallidus, the internal capsule, the caudate, as well as the insularis cortex. The caudate was also easily distinguished from surrounding structures, such as the corona radiate, the putamen and overlaying cortical areas, such as the motor and parietal cortex. Although no clear anatomical distinction between motor and parietal cortex was possible based on signal intensity, the distinctive shapes of the corona radiate allowed us to perform separate and consistent measurements within the motor and parietal cortical areas. The hippocampus was clearly resolved on T₂-weighted images and afforded a reliable measurement with a high inter- and intra-rater reliability (>96%), but sub-structures, such as the CA1-4, granular layer of the dentate gyrus (GrDG), alveus, or the parasubiculum (PaS), prosubiculum (ProS), presubiculum (PrS), subiculum (S) could not be sufficiently and consistently resolved for separate volumetric measurements.</p

Motor performance and loss of dopaminergic innervation of the nigral-striatal pathway.

<p><b>A.</b> Motor activity and motor disability in MPTP treated common marmoset treated with a single dose of 12.5mg/kg levodopa plus 12.5 mg/kg carbidopa. Administration of levodopa led a marked increased in motor activity (left panel) and marked reduction in motor disability (right panel). Both motor activity and motor disability peak at around 90 min of levodopa oral administration. Each data point is a mean ± s.e.m, n = 7. <b>B.</b> A denervation of tyrosine hydroxylase (TH) positive dopaminergic fibers is seen in the caudate and putamen after MPTP administration. <b>C.</b> A higher magnification of the caudate and putamen further highlights the dramatic loss of dopaminergic fibers in both structures with an even loss throughout the structure. <b>D.</b> A quantification of TH+ neurons in the substantia nigra further demonstrated a significant (p<0.001) loss of ~80% of these neurons due to MPTP treatment.</p

Calculation of statistical power.

<p><b>A.</b> Graphical representation of total sample size required for a given effect size to achieve different levels of statistical power for two-tailed (i.e. if no hypothesis regarding direction of effect is available) t-test comparisons. The effect size for different comparisons is indicated in this graph with the corresponding statistical power. With N = 11 in the current experiment, a sufficient statistical power 1-β>0.8 is achieved to avoid Type II errors (i.e. false negatives) with a Type I error (false positives) rate set at 5% (i.e. p<0.05). <b>B.</b> Illustration of a power analysis for Pearson’s correlations indicates that small r values (<0.4) require substantial sample sizes to achieve sufficient power to afford a valid comparison at the 5% Type I error rate. In primate studies small sample size are typical due to availability of subjects. A total sample size of 11 is insufficient to achieve an 80% power even with r = 0.9. Significant correlations with medium to high associations of measures would require sample sizes >50 subjects. In rats, we have previously demonstrated that significant correlations between neuropathological measurements and MRI can be performed with N = 15 with high associations (r>0.7) [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0180733#pone.0180733.ref025" target="_blank">25</a>, <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0180733#pone.0180733.ref027" target="_blank">27</a>].</p