Uptake and release of [3H]gamma-aminobutyric acid by embryonic spinal cord neurons in dissociated cell culture.

Open Access
Publisher Website
Google Scholar
Abstract
The uptake and release of [3H].gamma.-aminobutyric acid (GABA) by embryonic chick spinal cord cells maintained in culture was investigated. Cells dissociated from 4 or 7 day old embryos were studied 1-3 wk after plating. At 3.degree. C, [3H]GABA was accumulated by a high affinity (Km .simeq. 4 .mu.M) and a low affinity (Km .simeq. 100 .mu.M) mechanism. The high affinity transport was markedly inhibited in low Na+ media, by ouabain, at 0.degree. C and by 2,4-diaminobutyric acid. Autoradiography, after incubation in 0.1 .mu.M [3H]GABA, showed that .apprx. 50% (range = 30-70%) of the multipolar cells were labeled. These cells were neurons rather than glia; action potentials and/or synaptic potentials were recorded in cells subsequently found to be labeled. Non-neuronal, fibroblast-like cells and co-cultured myotubes were not labeled under the same conditions. Studies of intact adult tissue indicate that high affinity transport of [3H]GABA may be unique to neurons that use GABA as a neurotransmitter. Of 15 physiologically identified cholinergic neurons, i.e., cells that innervated nearby myotubes, none were heavily labeled after incubation in 0.1 .mu.M [3H]GABA is significant in this regard. The newly taken up [3H]GABA was not metabolized in the short run. It was stored in a form that could be released when the neurons were depolarized in a high K+ (100 mM) medium. As expected for a neurotransmitter, the K+-evoked release was reversibly inhibited by reducing the extracellular Ca2+/Mg2+ ratio.