Blocks of the hippocampal subiculum, hippocampus, intracerebellar nuclei, cerebellar cortex and visual cortex were isolated from adult mice in the evening, at midnight or in the next morning. They were embedded in paraffin, cut into sections, stained with cationic iron colloid or aldehyde fuchsin and counter-stained with nuclear fast red or carbol-thionin. Specimens prepared at midnight contained a markedly increased number of dark neurons. In the cerebellar cortex, 50% or more of Punkinje cells were dark at midnight. The dark neurons, except for Purkinje cells, were usually provided with extracellular sulfated proteoglycans reactive to cationic iron colloid or aldehyde fuchsin. The specimens, including those of intracerebellar nuclei and cerebellar cortex, prepared in the next morning (or when the animals were sleeping) contained few dark neurons. These findings suggest that the dark neurons are exhausted or tired cells, which are restored to normal or light cells with sleep (resting of neurons), and that they are neither poorly fixed nor degenerative cells.

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