Manual

Introduction

The neurobiology of birdsong, as a model for human speech, is a fast growing area of research in the neurosciences and involves electrophysiological, histological and more recently magnetic resonance imaging (MRI) approaches. Many of these studies require the identification and localization of different brain areas (nuclei) involved in the sensory and motor control of song.
Until now, the only published atlases of songbird brains consisted in drawings based on histological slices of the canary and of the zebra finch brain.
Taking advantage of high-magnetic field (7 Tesla) MRI technique, we present the first high-resolution (80 × 160 × 160 µm) 3-D digital atlas in stereotaxic coordinates of a male zebra finch brain, the most widely used species in the study of birdsong neurobiology. Image quality allowed us to discern most of the song control, auditory and visual nuclei.
This zebra finch MRI atlas should become a very useful tool for neuroscientists working on birdsong, especially for co-registration of MRI data but also for determining accurately the optimal coordinates and angular approach for injections or electrophysiological recordings.
 

The atlas can be freely downloaded and can be interactively explored with MRIcro.

This digital three-dimensional MRI atlas of the zebra finch brain is available to the scientific community. Detailed description of this atlas can be found in the accompanying paper, when using this data-set in a publication of your own, please cite the following paper:
Poirier C., Vellema, M., Verhoye, M., Van Meir, V., Wild, J.M., Balthazart, J., Van Der Linden, A. (2008). A three-dimensional MRI atlas of the zebra finch brain in stereotaxic coordinates. NeuroImage, Volume 41, Issue 1.
The present web page mainly contains information about how to visualize the data in MRIcro environment. As we would like to keep track of who uses the atlas for our strictly internal documentation purposes, we kindly ask you not to re-distribute the datasets yourself but instead to refer interested scientists to this website.

Data format

The MRI atlas data (16-bit int) and delineation data (8-bit int) are provided in Raw data (file_name.raw), Amira format (file_name.am) and in Analyze format (file_name.img and file_name.hdr).
Important note: Analyze files (.img and .hdr) must be saved in the same directory.

Name of the files:

/en/rg/bio-imaging-lab/research/mri-atlases/zebra-finch-brain-atlas/manual/

  • MRI brain data: atlas
  • MRI brain data in skull format: atlas_for_skull
  • CT skull data: skull

Delineation data:

  • brain_delineations
  • nuclei_delineations

Delineation data in skull format:

  • brain_delineations_for_skull
  • nuclei_delineations_for_skull
  • skull_delineations
  • /en/rg/bio-imaging-lab/research/mri-atlases/zebra-finch-brain-atlas/manual/

MRI data description

  • Dimensions: 256*256*256 (pixels)
  • Resolution: 0.08*0.08*0.08 (mm)
  • Sequence: Spin Echo sequence (TE/TR: 30/3000 ms), 4 averages

CT data description

  • Dimensions: 300*300*512 (pixels)
  • Resolution: 0.08*0.08*0.08 (mm)

Definition of the stereotaxic frame

This frame was defined by the symmetrical axis of the brain and the base of the brain on the mid-sagittal slice tilted by 10 deg/en/rg/bio-imaging-lab/research/mri-atlases/zebra-finch-brain-atlas/manual/rees. The Origin was defined as the crossing point between the mid-sagittal plane and the Commissura Posterior.

I- Visualization of the atlas dataset with MRIcro

  1. Download MRIcro and install it
  2. Run MRIcro
  3. In File menu, Open image…[Analyse or VoxBo]
  4. Select the file named atlas.hdr. A horizontal view should be displayed on the right panel.
  5. Adjust the contrast by dragging over the image while depressing the RIGHT mouse button.
  6. Select in the Slice Viewer section (left panel, purple spot in fig. 1) the icon representing the three orthogonal views. The coronal view is now displayed on the top/ left corner, the sagittal view is on the top/right corner and the horizontal view in the bottom/left corner of the right panel.
  7. Press on the LR/RL icon (orange spot in fig. 1) to select SPM2 default orientation. After this manipulation, the left hemisphere will be represented on the left side of the coronal and horizontal views (neurological orientation). This means that now, you look at the coronal view from the back.

II- Reading of relative coordinates

  1. In the Slice Viewer section, select the voxel with the following coordinates: X = 132, Y = 120, Z = 128. These coordinates are expressed in voxels. Then press the red cross button (yellow spot in fig. 1). The red cross should target what we have chosen as the origin of the atlas: the intersection of the posterior commisura and the mid-sagittal plane.
  2. To read the coordinates related to the origin, adjust the origin in the Header Information section (grey spot in fig. 1) to X = 125; Y = 120; Z = 128. Then press on the icon positioned on the left side of the red cross icon (blue spot in fig. 1). Coordinates relative to the origin will be displayed on the bottom right corner. They are now of 0, 0, 0 since the red cross is positioned at the origin. What you see on your screen should correspond to figure 1.
  3. When you open the atlas dataset, voxel size is automatically expressed in the Header Information section in hundreds of microns (0.8 *0.8*0.8). Adjust the voxel size to 80*80*80 (the new unit will be now microns). Now if you click on one voxel of one view, the 3 orthogonal slices containing this voxel will be displayed. Relative coordinates will be automatically adjusted and will be expressed in microns (and not in mm as written on the screen). The first number corresponds to the X value, the second one to the Y value and the third one to the Z value. The X axis corresponds to the left/right axis (with positive values for the right side), the Y axis corresponds to the rostral/caudal axis (with positive values for the rostral side) and the Z axis corresponds to the dorsal/ventral axis (with positive values for dorsal side). Absolute coordinates, expressed in voxels in the Slice Viewer section will also be automatically adjusted.

Fig. 1: Illustration of MRIcro display. The red cross is positioned on the Commissura Posterior on the mid-sagittal slice, which was used as the origin of the stereotaxic frame.

Options (on the left panel):

  1. Image(s) can be enlarged by using the zoom button (red spot in fig. 1).
  2. It is possible to read the coordinates of one voxel without changing the three orthogonal slices displayed on the right panel. For this, position your mouse cross on this voxel (without clicking). Relatives coordinates will be displayed at the bottom of the left panel (green spot in Fig. 1).

III- Visualization of the delineations superimposed on the 3D dataset

To superimpose delineations, go in the menu Overlay, and in the sub-menu Load image overlay… and select a delineation data set (for instance nuclei_delineations.hdr). It will take a few seconds before delineations appear.
Important note: this step needs to be done with the original values of voxels size (0.8, 0.8, 0.8) and origin (0, 0, 0) (in the Header Information section). If you want to read relative coordinates in microns, adjust these values after the superimposition (see steps 9 and 10 in Reading of relative coordinates).
Option: Transparency degree and color of delineations can be changed in Overlay menu.

Legend:
At the bottom of the left panel, the last number (between []) corresponds to the number of the labeled structure where the mouse cross is positioned (pink spot in fig. 2). The numbers indicate the structures as following: 1.024: L, 2.047: HVC, 3.02: X, 4.043: RA, 5.016: E, 6.039: LMAN, 7.012: MLd, 8.035: DLM, 9.008: Ov, 10.03: Olfactory bulb, 11: nXIIts, 12.03: Rt, ≥13: TeO

When the three views are displayed, sometimes MRIcro encounters problems to display this number. Displaying only one view at a time (by selecting this view in the Slice Viewer section) solves this problem.

Fig. 2: Superimposition of nuclei delineations to the atlas dataset. The mouse was positioned in the Optic Tectum (green delineations) when a copy of the screen image was saved. The number > 13 is thus displayed at the bottom of the left panel (pink spot).

IV- Reading of coordinates in the brain in relation to the skull

The atlas dataset and brain and nuclei delineations are provided in an image matrix of 256*256*256. This matrix fits the brain size. Because the skull is bigger than the brain, we needed to increase this image matrix to the one of the skull (300*300*512) to allow superimposition in MRIcro. Atlas dataset, brain and nuclei delineations are thus also provided with this bigger matrix.

To read coordinates on the brain while visualizing the skull at the same time, the coordinates of the zero need to be adjusted to this new matrix.

  1. Open the file named atlas_for_skull.hdr.
  2. Adjust the contrast
  3. Display the three orthogonal views.
  4. Verify that the SPM2 default orientation is still selected (L/R button) to visualize the left part of the skull on the left part of the coronal and horizontal images.
  5. Overlay the file named skull.hdr (this operation can be quite long).
  6. Now adjust the voxel size to 80, 80, 80 (microns) and the origin to X = 147, Y = 142, Z = 255.
  7. In the Slice Viewer section select the voxel with the following coordinates: X = 154, Y = 142, Z = 255. Press on the red cross icon and on the icon just on its left to display the coordinates in the right bottom corner of the right panel. What you see on your screen should correspond to figure 3. The red cross is positioned at the origin (relative coordinates: 0, 0, 0).
    You can now navigate into the brain/skull, the relative coordinates will be automatically adjusted.

Figure 3: Superimposition of the skull to the atlas dataset.