Compartmentation and release of exogenous GABA in sheep brain synaptosomes

Exogenous tritiated gamma-aminobutiric acid ([3H]GABA) is retained in two compartments in sheep cortex synaptosomes, corresponding to cytoplasmic and vesicular spaces, assuming that freeze-thawing the synaptosomes loaded with [3H]GABA releases the cytoplasmic [3H]GABA (81 +/- 3.9%), and that subsequent solubilization of the synaptosomes with 1% sodium cholate releases the vesicular [3H]GABA (19 +/- 3.9%). Depolarization of synaptosomes with 40 mM K+ in a Na+-medium, in the absence of Ca2+, releases 20.3 +/- 2.7% of the [3H]GABA retained in the synaptosomes. The [3H]GABA released under these conditions comes predominantly from the cytoplasm. The presence of 1 mM Ca2+ during depolarization releases an additional 13% (a total of about 33.5 +/- 9.9%) of the releasable [3H]GABA, and the [3H]GABA release which is Ca2+-dependent also comes mostly from the cytoplasmic compartment. When choline replaces external Na+, the [3H]GABA release is absolutely Ca2+-dependent, and the [3H]GABA released also comes mostly from the cytoplasmic pool. Therefore, it appears that [3H]GABA taken up by synaptosomes is accumulated mostly in the cytoplasmic compartment from which it is released upon depolarization. The technique described permits distinguishing the effect of different factors on the two pools of accumulated [3H]GABA.