The effect of loading and extraction temperatures, composition of medium, and molecular size on retention of extracellular space marker by mouse cerebrum slices

Abstract

Mouse cerebrum slices were incubated for 1 hour at either 37° or 0° in media containing the ‘extracellular space markers’, inulin, sucrose, D-mannitol, D-sorbitol, sulfate, or thiocyanate. They were then transferred to marker-free medium, and the marker was extracted for 1 hour. With both loading temperatures, some marker was retained at 37°, and an equal or greater amount was retained at 0°. The greater the molecular weight of the carbohydrate marker, the higher the NaCl concentration needed for greater retention upon extraction at 0° than at 37°. Slices loaded at 0° required a higher salt concentration than those loaded at 37° for greater retention to occur upon extraction at 0°. Experiments in media made hypertonic with D-mannitol, D-sorbitol, or choline chloride instead of additional NaCl show that the effect of additional salt is not simply due to increased tonicity. Unlike the other markers, thiocyanate did not show a retention difference. Although the results correlate poorly with ‘second marker spaces’ computed from uptake data, they agree with uptake studies in suggesting compartmentation of the ‘extracellular’ space.