Comparison of extracellular space in the mature and aging rat brain using a new technique

A new technique for measuring extracellular space in the rat brain has been developed. It involves opening the blood-brain barrier with a bolus of hyperosmotic sucrose followed by a high-pressure perfusion of the cerebral vasculature with an isotonic solution containing an impermeant radioactive tracer, [3H]sucrose. After allowing the concentration of tracer in the brain to reach a plateau, the amount of radioactivity/mg of brain tissue is expressed as a percentage of the amount of radioactivity/mg of perfusate to obtain a value for extracellular space. The addition of glutaraldehyde to the perfusate results in the brain being fixed simultaneously for electron microscopy. Reproducible estimates of extracellular space were obtained similar to those obtained by other methods (e.g. Levin et al. 1970). As it is impossible to be sure of the validity of the absolute value of extracellular space obtained by any method using perfused solutions we have used our method for comparative purposes. Extracellular space was measured in mature (control) and ageing rats to test the claim that the volume of space in the cerebral cortex is substantially reduced with ageing (Bondareff and Narotsky 1972). We found a consistent tendency for the extracellular space to increase with age in the 6 regions of brain examined. This was not statistically significant except in the group of ageing rats on a food-restricted diet. Therefore, these results do not support a generalisation that the extracellular space decreases in the ageing brain. In both control and ageing rats, extracellular space was shown to be unevenly distributed in the brain, the largest space being present in the cerebellum, olfactory bulb, inferior and superior colliculi and the least space in the white matter. These are the first measurements of extracellular space in which assessment is possible by both electron microscopy and by the measurement of a chemical tracer.