Inhibitors of ATP Release Inhibit Vesicular Nucleotide Transporter

Kato, Yuri; Omote, Hiroshi; Miyaji, Takaaki

doi:10.1248/bpb.b13-00544

Cited by 17 publications

(11 citation statements)

References 22 publications

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“…Thus, it is highly probable that there are VNUT-specific inhibitors among the intermediates of glycolysis, the TCA cycle, and β-oxidation or their derivatives. Consistent with this speculation, arachidonic acid inhibits VNUT with an ID50 of 6.5 μM [134]. Proteoliposomes containing purified VNUT and other vesicular neurotransmitter transporters provide a suitable screening system for exploring vesicular ATP release blockers.…”

Section: Development Of a Vesicular Atp Release Blockermentioning

confidence: 58%

“…The facts that enhanced purinergic chemical transmission causes a wide variety of pathophysiological phenomena [1,2] and that VNUT −/− mice appear to be healthy and exhibit therapeutic effects towards the metabolic syndrome anticipate the development of vesicular ATP release blockers for therapeutic purposes. However, the VNUT inhibitors known so far are toxic and not specific to VNUT [4,34,134]. For example, glibenclamide, an inhibitor of the ABC transporter, also inhibits VNUT, with an ID50 being 1.6 μM [134].…”

Section: Development Of a Vesicular Atp Release Blockermentioning

confidence: 99%

“…However, the VNUT inhibitors known so far are toxic and not specific to VNUT [4,34,134]. For example, glibenclamide, an inhibitor of the ABC transporter, also inhibits VNUT, with an ID50 being 1.6 μM [134]. Arachidonic acid, a max-anion channel blocker, also inhibits VNUT, with ID50 of 6.5 μM.…”

Section: Development Of a Vesicular Atp Release Blockermentioning

confidence: 99%

“…Evans blue and DIDS strongly inhibit VNUT, with ID50s of 40 nM and 1.5 μM, respectively. They also inhibit other SLC17 members, anion channels, and ion-transporting ATPases, with inhibition by some of these compounds being irreversible under natural circumstances [134,135]. Therefore, when these compounds are used to characterize the types of ATP release, researchers must remember that these compounds are not specific to the target protein.…”

Section: Development Of a Vesicular Atp Release Blockermentioning

confidence: 99%

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Vesicular nucleotide transporter (VNUT): appearance of an actress on the stage of purinergic signaling

Moriyama

Hiasa

Sakamoto³

et al. 2017

Purinergic Signalling

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Vesicular storage of ATP is one of the processes initiating purinergic chemical transmission. Although an active transport mechanism was postulated to be involved in the processes, a transporter(s) responsible for the vesicular storage of ATP remained unidentified for some time. In 2008, SLC17A9, the last identified member of the solute carrier 17 type I inorganic phosphate transporter family, was found to encode the vesicular nucleotide transporter (VNUT) that is responsible for the vesicular storage of ATP. VNUT transports various nucleotides in a membrane potential-dependent fashion and is expressed in the various ATP-secreting cells. Mice with knockout of the VNUT gene lose vesicular storage and release of ATP from neurons and neuroendocrine cells, resulting in blockage of the initiation of purinergic chemical transmission. Thus, VNUT plays an essential role in the vesicular storage and release of ATP. The VNUT knockout mice exhibit resistance for neuropathic pain and a therapeutic effect against diabetes by way of increased insulin sensitivity. Thus, VNUT inhibitors and suppression of VNUT gene expression may be used for therapeutic purposes through suppression of purinergic chemical transmission. This review summarizes the studies to date on VNUT and discusses what we have learned about the relevance of vesicular ATP release as a potential drug target.

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Section: Development Of a Vesicular Atp Release Blockermentioning

confidence: 58%

Section: Development Of a Vesicular Atp Release Blockermentioning

confidence: 99%

Section: Development Of a Vesicular Atp Release Blockermentioning

confidence: 99%

Section: Development Of a Vesicular Atp Release Blockermentioning

confidence: 99%

See 2 more Smart Citations

Vesicular nucleotide transporter (VNUT): appearance of an actress on the stage of purinergic signaling

Moriyama

Hiasa

Sakamoto³

et al. 2017

Purinergic Signalling

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“…More recently it was proposed that VNUT-dependent ATP release from spinal cord dorsal root neurons might originate neuropathic pain in mice ( Masuda et al, 2016 ). Furthermore, clodronate was identified as a novel inhibitor of VNUT for the treatment of neuropathic and inflammatory pain, opening up new therapeutic possibilities for other pathologies ( Kato et al, 2013 , 2017 ).…”

Section: Discussionmentioning

confidence: 99%

Specific Temporal Distribution and Subcellular Localization of a Functional Vesicular Nucleotide Transporter (VNUT) in Cerebellar Granule Neurons

et al. 2017

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Adenosine triphosphate (ATP) is an important extracellular neurotransmitter that participates in several critical processes like cell differentiation, neuroprotection or axon guidance. Prior to its exocytosis, ATP must be stored in secretory vesicles, a process that is mediated by the Vesicular Nucleotide Transporter (VNUT). This transporter has been identified as the product of the SLC17A9 gene and it is prominently expressed in discrete brain areas, including the cerebellum. The main population of cerebellar neurons, the glutamatergic granule neurons, depends on purinergic signaling to trigger neuroprotective responses. However, while nucleotide receptors like P2X7 and P2Y13 are known to be involved in neuroprotection, the mechanisms that regulate ATP release in relation to such events are less clearly understood. In this work, we demonstrate that cerebellar granule cells express a functional VNUT that is involved in the regulation of ATP exocytosis. Numerous vesicles loaded with this nucleotide can be detected in these granule cells and are staining by the fluorescent ATP-marker, quinacrine. High potassium stimulation reduces quinacrine fluorescence in granule cells, indicating they release ATP via calcium dependent exocytosis. Specific subcellular markers were used to assess the localization of VNUT in granule cells, and the transporter was detected in both the axonal and somatodendritic compartments, most predominantly in the latter. However, co-localization with the specific lysosomal marker LAMP-1 indicated that VNUT can also be found in non-synaptic vesicles, such as lysosomes. Interestingly, the weak co-localization between VNUT and VGLUT1 suggests that the ATP and glutamate vesicle pools are segregated, as also observed in the cerebellar cortex. During post-natal cerebellar development, VNUT is found in granule cell precursors, co-localizing with markers of immature cells like doublecortin, suggesting that this transporter may be implicated in the initial stages of granule cell development.

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Reconstitution and Transport Analysis of Eukaryotic Transporters in the Post-Genomic Era

Omote

Moriyama

2017

Methods in Molecular Biology

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Measuring transport activity through reconstituted proteoliposomes is a key technique to resolve numerous problems found in the traditional methods. The system includes overexpression, purification, and reconstitution of transporters. Mixing of purified transporter with lipid and dilution below the critical micelle concentration result in rapid generation of proteoliposomes. Incubation of proteoliposomes in the presence of a driving force initiates substrate uptake. After starting the reaction, samples are passed through a gel filtration column to separate proteoliposomes from the reaction mixture. Here, we describe step-by-step procedures for such reconstitution assays.

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Inhibitors of ATP Release Inhibit Vesicular Nucleotide Transporter

Cited by 17 publications

References 22 publications

Vesicular nucleotide transporter (VNUT): appearance of an actress on the stage of purinergic signaling

Vesicular nucleotide transporter (VNUT): appearance of an actress on the stage of purinergic signaling

Specific Temporal Distribution and Subcellular Localization of a Functional Vesicular Nucleotide Transporter (VNUT) in Cerebellar Granule Neurons

Reconstitution and Transport Analysis of Eukaryotic Transporters in the Post-Genomic Era

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