Active transport of gamma-aminobutyric acid and glycine into synaptic vesicles.

Abstract

Although y-aminobutyric acid (GABA) and glycine are recognized as major amino acid inhibitory neurotransmitters in the central nervous system, their storage is poorly understood. In this study we have characterized vesicular GABA and glycine uptakes in the cerebrum and spinal cord, respectively. We present evidence that GABA and glycine are each taken up into isolated synaptic vesicles in an ATPdependent manner and that the uptake is driven by an electrochemical proton gradient. Uptake for both amino acids exhibited kinetics with low affinity (Km in the millimolar range) similar to vesicular glutamate uptake. The ATP-dependent GABA uptake was not inhibited by the putative amino acid neurotransmitters glycine, taurine, glutamate, or aspartate or by GABA analogs, agonists, and antagonists. Similarly, ATPdependent glycine uptake was hardly affected by GABA, taurine, glutamate, or aspartate or by glycine analogs or antagonists. The GABA uptake was not affected by chloride, which is in contrast to the uptake of the excitatory neurotransmitter glutamate, whereas the glycine uptake was slightly stimulated by low concentrations of chloride. Tissue distribution studies indicate that the vesicular uptake systems for GABA, glycine, and glutamate are distributed in different proportions in the cerebrum and spinal cord. These results suggest that the vesicular uptake systems for GABA, glycine, and glutamate are distinct from each other. y-Aminobutyric acid (GABA) and glycine are the major inhibitory neurotransmitters in the vertebrate central nervous system (1, 2). Recently, the primary structures of GABAA and glycine receptors have been deduced; the subunits of these receptors have been shown to have substantial sequence homology, particularly in the region thought to be involved in conducting chloride (3). GABA and glycine are released upon membrane depolarization, both in a calcium-dependent manner (4-6) and in a calciumindependent manner (4, 7). Recent evidence indicates that the calcium-dependent release of GABA originates from a noncytoplasmic compartment (8). There are also observations indicating that GABA and glycine are concentrated in distinct nerve terminals (9, 10). However, localization of endogenous amino acids in synaptic vesicles has not been clearly demonstrated, either with intact tissues or isolated vesicle preparations. In addition, there has been little study on the vesicular GABA and glycine uptake processes. We have previously provided evidence that glutamate is taken up into synaptic vesicles by a proton-motive force generated by a proton-pump ATPase in the vesicle (11-13). In this communication, we have studied vesicular GABA and glycine uptake, using a synaptic vesicle preparation different from that previously used for vesicular glutamate uptake. We provide evidence that suggests that GABA and glycine are also accumulated into synaptic vesicles in an ATP-dependent manner and that their uptake is driven by an electrochemical proton gradient. We have characterized these uptake systems with r...

Unique pH dynamics in GABAergic synaptic vesicles illuminates the mechanism and kinetics of GABA loading

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Vesicular Inhibitory Amino Acid Transporter Is a Cl−/γ-Aminobutyrate Co-transporter

“…GABAergic inhibitory signaling occurs in the brain, whereas both GABAergic and glycinergic signaling occur primarily in spinal cord and brain stem. Like other neurotransmitters such as L-glutamate and acetylcholine, GABA and glycine are actively accumulated in synaptic vesicles through vesicular neurotransmitter transporters (3)(4)(5)(6)(7). Currently, only one type of transporter, SLC32A1, is known to be responsible for this process and is referred to as either vesicular GABA transporter or vesicular inhibitory amino acid transporter (VIAAT) (2,8,9).…”

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“…Recent evidence has demonstrated that glutamate, GABA and glycine are actively taken up and stored in isolated synaptic vesicles [53,54]. The vesicular loading occurs exclusively for glutamate or GABA using purified cerebrum vesicles and for glycine using spinal cord synaptic vesicles.…”