Aspects of the quantitative analysis of neurons in the cerebral cortex

We address three problems concerning the quantitative analysis of nerve cell distribution in the cerebral cortex: (i) preparatory tissue deformation (shrinkage); (ii) difficulties in differentiating between small neurons and astroglia; and (iii) the bias introduced by the counting method. We found that staining with Richardson's solution led to no shrinkage in Vibratome-cut sections of aldehyde-fixed rat brains, but did result in staining of the neurons and left the glial cells unstained. This was in striking contrast to Nissl staining which introduced a linear shrinkage of 20–30% and stained all kinds of cortical cells indiscriminately. A computer-based unbiased counting method was implemented by taking advantage of the stereological procedure referred to as the ‘optical disector’ (Gundersen, H.J.G) (1986) Stereology of arbitrary particles, J. Microsc., 143: 3–45).