FIGURE 5.

Measurement of astrocyte density using the optical fractionator. Four panels illustrating the implementation of the optical fractionator at a single site within the white matter. The counting brick is composed of a counting frame with a red exclusion line, a green inclusion line, and a fraction of the volume of the measured thickness of the tissue. (A) At the top of the section, visible astrocytes are not yet counted as they are outside the focal plane. (B) As the probe descends through the z dimension, several astrocytes come into fine focus and are counted. Arrows show that 2 cells can be counted: 1 is in the counting frame and 1 is touching the green inclusion line. The cell marked by the arrowhead is not counted because it contacts the red exclusion line. (C, D) The labeled cells are not in focus and are not counted. Repeating these procedures at each site within the white matter produces an unbiased estimate of the number of astrocytes within the counting bricks. Scale bar for A =  10 µm.