3D Reconstruction and Standardization of the Rat Vibrissal Cortex for Precise Registration of Single Neuron Morphology
Figure 8
Registration of 3D neuron morphologies to the standardized barrel cortex.
(A) Example of a L5 thick-tufted neuron reconstructed from 100 µm thick sections. Outlines of pia, WM and barrels are added to the reconstruction in the coordinate system given by the slicing direction. (B) Side view of (A). The slicing direction does not match the orientation of the column containing the neuron soma. (C) Reconstruction of landmarks in 3D and registration of the barrels to the standardized barrel field. It may be necessary to correct the orientation of the neuron to match the direction of the local column axis (gray – before rotation, red – after rotation). The histograms show the rotation angle used to align the barrel field outlines with the standardized barrel field (global orientation) and the angle of the subsequent rotation aligning the neuron orientation with the local column orientation. (D) The barrel outlines in the reconstruction are of lower resolution along the slicing direction and thus show a systematic offset compared to the standardized barrel landmarks. This is corrected for by translation along the local column axis. (E) The variability between different reconstructions is minimized by scaling the supragranular, granular and infragranular structures such that the landmarks of the reconstructed neuron coincide with the standardized landmarks. The average scaling factors for the individual layers are very close to 1. (F) Registration of the neuron to the standardized barrel cortex allows objective determination of anatomical parameters such as the soma location in 3D. Comparison of the registered depth of 56 neurons with the penetration depth of the pipette recorded during the experiment shows that this recording depth is on average 46 µm lower than the registered depth, but varies in a range of up to 200 µm around the registered depth.