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After each neurotransmitter release, MK801, an open-channel blocker that can only block a channel that has been activated, decreases initial current (which has been normalized for wild type and mutant cells). In the colocalization model, VGLUT1 and VGLUT2 occupy the same vesicles filling them partially. The commingling model proposes that transporters occupy distinct vesicles in the same synapse. The segregation model proposes that the transporters are found in different synapses.
A brain without glutamate, the major excitatory neurotransmitter in mammals, is quite literally unthinkable. Yet, knockout mice lacking the crucial vesicular glutamate transporter, VGLUT1, can live for up to a few months. This unexpected finding has enabled two research groups to investigate the function of VGLUT1.
With potential links to learning and memory, as well as neurological diseases such as epilepsy, approaching glutamate transmission through the protein believed to load it...
TOGETHER OR APART
Today, both labs are still keeping pace, having independently created mouse lines that lack VGLUT1. The mice show little synaptic transmission as evidenced by their virtual lack of excitatory postsynaptic potentials.23 Some activity does remain, however, and both groups uncovered that this is due to VGLUT2 expression, which is coexpressed in the same cells as VGLUT1 in the first weeks after birth, before being shut off. "That's why the mice didn't die right after birth," says Rosenmund.
How VGLUT1 and VGLUT2 might mingle in these cells, and potentially on the same vesicles, is still a matter of debate. Edwards and colleagues argue that VGLUT1 and VGLUT2 reside in different synapses.2 This position is based partly on their finding that VGLUT1 knockout mice have a reduced frequency, but unchanged amplitude, of miniature excitatory postsynaptic current (mEPSC), an indirect indicator of the amount of neurotransmitter in a vesicle. "This could only be explained by complete segregation of the two isoforms," says Edwards.
In contrast, Rosenmund's group found shifted amplitudes for the mEPSC of VGLUT1 knockout cells.3 This, they argue in part, reveals that knockout cells release half-full vesicles, thus placing VGLUT1 and VGLUT2 together on the same vesicles. Thomas Otis, a researcher from UC, Los Angeles Medical School, attributes the differences to the different neuronal preparations used in the two papers; Rosenmund's group used primary cultured cells, and Edwards's group used brain slices to measure mEPSCs. "But I'm sure a lot of people will be doing these experiments in a variety of different contexts, trying to see which side things fall on," says Otis.
Regardless of the answer, the research is beginning to tease apart the mechanism of vesicle loading says Erik Jorgensen, associate professor at the University of Utah. In the past, it was thought that only vesicles that were completely filled could be released, "Rosenmund and colleagues have excluded the hypothesis, producing a very strong result," says Jorgensen.
Vladimir Parpura, assistant professor at UC-Riverside, says the two papers point out the underappreciated complexity of glutamate neurotransmission. He and colleagues recently found that astrocytes also release glutamate by using VGLUT1 and VGLUT2.4 This is possibly a mechanism for storing or recycling glutamate says Jorgensen, suggesting that the glutamate transporters have multiple effects.
A number of drugs, such as amphetamines and norepinephrine transporters, manipulate vesicular transporters, but nothing yet manipulates glutamate transport. This research has opened a window on designing molecules that could be used to change synaptic strength, says Otis. "This has ramifications for altering signaling in the brain during learning, memory, and disease."
David Secko
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