Neonate and Juvenile
NSA’s FDA Neurotoxicity Presentation 2015
Developmental Neurotoxicity Assessment: Neonatal Programmed Cell Death
The assessment of permanent damage in neonates and juveniles is difficult due to the normal occurrence of programmed cell death in the developing brain, and timelines for disintegration (and the corresponding windows to view these) are shortened from days to hours. Patterns of degeneration must be compared to a baseline level to determine changes in cell death activity. Architecting a study design to account for the earlier and shorter observable opportunity of cell death becomes a critical consideration.
The disruption of the chronology of programmed cell death (apoptosis) in the developing brain is an increasing issue of concern. As an example, the Disintegrative Degeneration stains i.e. CuAg have been used to show how alcohol causes extensive cell death in neonate rats (Ikonomidou et al. Science 257: 1056, 2000).
The distinct pattern of degenerated cells in the ventral border of the anterior ventral thalamus is shown below.
Adjacent sections from neonate rat brain stained with thionine and the Cupric Silver method by NSA.
Thionine Nissl Stain
CuAg Disintegrative Degeneration Stain
The top two images are from two adjacent sections of the anterior thalamus from a 3 day old rat in which abundant programmed cell death (apoptosis) is taking place. The upper left image is stained for nissl substance to show neuron cell bodies and the upper right with the cupric silver stain to show disintegrative degeneration-seen as black dots. The ease of detection of degeneration in the cupric silver stained section vs. the cell body only nissl stain is quite clear. A more highly magnified view of both of these stains is shown in the lower two images. Numerous dark blue dots can be detected among normal cells in the nissl stained image which are the pyknotic nuclei cells undergoing apoptosis. In the lower right image the black irregular profiles of degenerating cells are quite conspicuous and easily detected.
The temporal histogram below illustrates peak times of cell death in the ventral septum of rat brain. The tabulation of all neuronal populations will identify critical periods during which brain development might be perturbed.
Degeneration in the Lateral Septum; Postnatal Day 3
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