Stereological studies on rat spinal neurons during postnatal development: estimates of mean perikaryal and nuclear volumes free from assumptions about shape.

A stereological method for estimating the mean volumes of particles independent of assumptions about their shapes is illustrated using neurons in the ventral horn of rat cervical spinal cord. Male rats of 20 and 120 days post partum age were killed by intracardiac perfusion with formaldehyde/glutaraldehyde solutions. Cervical enlargements were removed, trimmed and embedded in resin. Randomised sections of ventral horn were photographed in a systematic pattern and used to estimate the volume-weighted mean volumes of neuronal perikarya and their nuclei. Volumes were estimated from point-sampled intercepts using rulers to classify intercept lengths. Classifying motoneuron perikarya was extremely reproducible, group means (coefficients of variation) at 120 days post partum being 25,190 microns3 (23%) and 24,250 microns3 (25%) in two separate trials. Classifying all neurons at the same age gave values of 20,520 microns3 (22%) and 22,490 microns3 (28%) in two trials. The mean perikaryal volumes of motoneurons at 20 and 120 days of age were not significantly different but nuclear volumes increased from 1,580 microns3 (16%) at 20 days to 2,660 microns3 (28%) at 120 days. These results illustrate the value of the method for obtaining unbiased and efficient estimates of the sizes of irregular perikarya and their nuclei. The benefit is that sizes can be estimated without biases due to simplifying assumptions about perikaryal/nuclear shape or nucleolar location. The influence of section thickness (even of thick paraffin sections) on the estimates is also negligible.