FIGURE 3.

Light micrographs showing rat striatal ChAT immunostained neurons (arrowheads) in the same location within two consecutive sections (A) and (B), with all disector neurons identified in (C) and (D). The blood vessel labeled BV in the middle of both sections is a key reference point (see also (C) and (D)), as are other vessels denoted BV. To identify disector neurons, an acetate sheet (i.e., a clear overhead projector sheet) was overlaid on (A). The location of each neuron and the location of the central blood vessel were drawn on the sheet. The sheet was then moved to (B), and the location of the central blood vessel and the √√ neurons (i.e., the neurons in both sections) was aligned. This alignment is indicated by each solid red outline that has been drawn over each specific √√ neuron in (C) and (D). On the basis of this alignment, it can be seen that this is the same region in the two consecutive sections. The numbers 97 and 98 refer to the specific section numbers. (C) and (D) also indicate how disector neurons were counted using the (physical) disector method for the two serial sections/photos. √x indicates that the nucleus is in the first section and is counted for that section. This is because there is no nucleus in the second section. √√ indicates the nucleus was present in both sections, and hence this neuron was not counted as a disector neuron for either of these sections. √‐ indicates that the nucleus is present in the first section and counted, but the cell is not in the second section. ‐√ indicates the opposite. Thus, the nucleus is in the second section and counted for that section, but the cell is not in the first section. xx indicates that the cell had no nucleus in either section. If there was a nucleus in the second section but not in the first section, then the neuron would be counted as an x√ disector neuron belonging to the second section. Note that in this example there are no x√ disector neurons. Note also that determination of whether the nucleus was present or not was completed for each neuron at a magnification of ×3450. We acknowledge that a disector analysis can now be completed using computer software (e.g., Photoshop) instead of using acetate sheets as described here for each sampled region within the structure of interest. Scale bar in (A), 200 µm. Same scale applies in (A)–(D).