Review articleVGLUTs: ‘Exciting’ times for glutamatergic research?
Introduction
Glutamate is an ubiquitous anionic amino acid that is present in all cell types in our body. In the brain, however, it acts as a signaling molecule, being stored and released from a subpopulation of neurons (so-called glutamatergic neurons). In a resting neuron, glutamate is stored in a tiny membrane-bound organelle found within the synaptic terminal – the synaptic vesicle – and it is released from the neuron by the fusion of this vesicle with the plasma membrane. This process, which occurs in response to neuronal activity, is known as exocytosis. This released glutamate is actually capable of eliciting a response in most of the neurons so far examined in the mammalian central nervous system (CNS).
As is the case for other small classical neurotransmitters, such as GABA and biogenic monoamines, glutamate is synthesized in the presynaptic cytoplasm with the aid of a synthesizing enzyme (a phosphate-activated glutaminase (PAG)) and translocated into the vesicle lumen via a glutamate transport protein (VGLUT) located on the vesicle. Glutamate uptake into synaptic vesicles was initially characterized using a reasonably pure vesicle fraction isolated from mammalian brain (Disbrow et al., 1982, Naito and Ueda, 1985). As is the case for other classical neurotransmitters, glutamate uptake is driven by a proton-dependent electrochemical gradient that exists across the vesicle membrane and which is created by a vacuolar-type ATPase. It exhibits stringent substrate specificity (highly specific for l-glutamate) and a relatively low affinity (Km = 1–3 mM). It depends predominately on the existence of a vesicular membrane potential gradient, rather than a pH gradient, unlike other vesicular neurotransmitter transporters that depend on the existence of a pH gradient as the driving force. A low concentration of chloride (2–5 mM) is necessary for its maximum activity. Although the first successful cloning of a vesicular monoamine transporter stimulated the search to identify and characterize other vesicular neurotransmitter transporters (Liu et al., 1992, Gasnier, 2000), molecular cloning of the VGLUT gene(s) had not been accomplished. In 2000, work from the laboratories of Jahn (Takamori et al., 2000) and Edwards (Bellocchio et al., 2000) shed first light on these enigmatic proteins, allowing for their subsequent molecular characterization and manipulation. In this review, I would like to overview the recent progress on VGLUT proteins and their physiological functions. In addition to the functional studies on VGLUTs in the nervous system, on which I will focus in this article, there is accumulating evidence that VGLUTs are also expressed in peripheral tissues, indicating that intercellular glutamate signaling is not just restricted to the CNS, but actually more broadly adopted in mammalian tissues. Readers who are interested in this aspect should refer to other recent reviews (Li et al., 2005, Moriyama and Yamamoto, 2004).
Section snippets
Vesicular glutamate transporters uncovered
Although it had long been conceptually envisaged that the process of loading glutamate into synaptic vesicles might be a key step in glutamatergic neurotransmission, the protein responsible for this process eluded detection for many years. When the protein was eventually identified it came as a surprise because the protein had already been cloned in 1994, but had been disguised as a different transporter protein (Ni et al., 1994). This protein, originally termed BNP1 (brain-specific Na+
Three VGLUTs in mammalian CNS
In addition to the previously known C. elegans orthologue of VGLUT, eat-4, multiple VGLUT isoforms have been identified in many other organisms such as Drosophila (Daniels et al., 2004), zebrafish (Higashijima et al., 2004) and frog (Gleason et al., 2003), in which glutamate is mainly used as a neurotransmitter at the neuromuscular junction. It seems that VGLUT genes are evolutionary conserved, concomitant with the organisms employing some form of glutamatergic signaling. Such a finding raises
What do mice without VGLUT1 tell us?
One of the main interests in the field is to clarify the functional differences between the three VGLUT isoforms. As mentioned above, intensive biochemical investigations on the three identified VGLUTs have yet to yield satisfactory answers. Moreover, because the VGLUTs (VGLUT1 and VGLUT2) were originally isolated as plasma membrane carriers for inorganic phosphate (Aihara et al., 2000, Ni et al., 1994), and VGLUT1 was shown to exhibit an additional chloride conductance associated with the
Do VGLUTs contribute to presynaptic plasticity?
The finding that the residual excitatory neurotransmission in VGLUT1 knockout mice appeared to be mediated by VGLUT2 led two groups to explore more precisely both how VGLUT1 and VGLUT2 are co-expressed in a single neuron and how VGLUT expression could contribute to synaptic plasticity in glutamatergic neurotransmission. Since it is now evident that the release of a quantum of glutamate (from a single vesicle) does not saturate postsynaptic glutamate receptors (Ishikawa et al., 2002, McAllister
VGLUT expression and pathology
Despite the current uncertainty regarding whether changes in VGLUT expression result in the modulation of synaptic efficacy, reports have started to establish a link between alterations in VGLUT isoform expression and pathological processes and also in the response to pharmacological treatments. For instance, VGLUT1 expression in the cerebral cortex and the hippocampus is increased by chronic administration of antidepressant drugs (Moutsimilli et al., 2005, Tordera et al., 2005). In addition,
Concluding remarks
Recent successes in the molecular identification of VGLUTs have promoted rapid progress in the anatomical identification of glutamatergic synapses in the CNS. Amongst the three VGLUT isoforms, VGLUT1 and VGLUT2 are largely segregated in the brain and are expressed in the presynaptic terminals of asymmetric synapses, revealing most, if not all, authentic glutamatergic synapses in the CNS. By contrast, VGLUT3 expression is less abundant compared to VGLUT1 and VGLUT2 and is expressed in
Acknowledgements
This review was written to acknowledge the grateful receipt of the Japan Neuroscience Society Young Investigator Award in 2005. This work was supported, in part, by a grant from the Japanese Ministry of Education, Science, Sports, Culture and Technology (no. 17680032) and by the 21st Century COE Program on “Brain Integration and its Disorder” at Tokyo Medical and Dental University. I am grateful to Reinhard Jahn (Göttingen, Germany) for his continuous support throughout my work on VGLUTs. I am
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