SIMULTANEOUS ISOLATION OF GLIAL AND NEURONAL FRACTIONS FROM RAT-BRAINHOMOGENATES - COMPARISON OF HIGH-AFFINITY L-GLUTAMATE TRANSPORT-PROPERTIES

An improved three-step Percoll density gradient centrifugation techniq ue is described for simultaneous isolation of glial plasmalemmal vesic les (GPV) and synaptosomal vesicles (SYN) from a rat brain homogenate. While electron microscopy revealed that fractions contained intact ve sicles with markedly distinct morphological features, measures of high -affinity [H-3]choline uptake, glutamine synthetase and carbonic anhyd rase activities, as well as Western blot analyses for glial fibrillary acidic protein and neuron specific enolase, served to confirm the low level of neuronal contamination in GPV fractions as well as the low l evel of glial contamination in SYN fractions. In addition, GPV and SYN fractions were used to characterize the kinetic and pharmacological p roperties of sodium-dependent [H-3]L-glutamate transport. In conclusio n, these results demonstrate the usefulness of this method for obtaini ng highly-enriched, functionally viable populations of glial and neuro nal elements which are suitable for studies of their respective cell f unctions in vitro.