Microglia release ATP by exocytosis

Abstract

Microglia survey the brain environment by sensing several types of diffusible molecules, among which extracellular nucleotides released/leaked from damaged cells have central roles. Microglia sense ATP or other nucleotides by multiple P2 receptors, after which they change into several different phenotypes. However, so far, it is largely unknown whether microglia themselves release ATP and, if so, by what mechanism. Here we show that exocytosis is the mechanism by which microglia release ATP. When we stimulated microglia with ionomycin, they released ATP and the release was dependent on Ca²⁺, vesicular H⁺-ATPase, or SNAREs but independent of connexin/pannexin hemichannels. VNUT was found to be expressed in microglia and exhibited no colocalization with lysosome. We also visualized the exocytosis of ATP by a quinacrine-based fluorescent time-lapse imaging. Moreover, we found that lipopolysaccharide increased the ionomycin-induced release of ATP, which was dependent on the increase in VNUT. Taken together, our data suggested that exocytosis is the mechanism of ATP release from microglia. When activated, they would release ATP by increasing VNUT-dependent exocytotic mechanisms.

Role of vesicular nucleotide transporter VNUT (SLC17A9) in release of ATP from AR42J cells and mouse pancreatic acinar cells

“…1 and 2), confirms our early studies and validity of using such fluorescent markers [4]. VNUT and [10][11][12][13][14][15]. In freshly isolated pancreatic acini, secretory/zymogen granule marker Rab3D co-localizes with VNUT ( Fig.…”

“…Soon after, we provided evidence that VNUT is expressed in pancreatic zymogen granules and is engaged in ATP uptake [9]. Since then, expression of VNUT has been demonstrated in several cell types indicating the general importance of VNUT for ATP release [10][11][12][13][14][15].…”

“…The two original templates were the VNUT plasmid [8,14] and the Clontech mCherry (from pmCherry-C1 vector) together with plasmid pPAP7160. Plasmid pPAP7160 was generated by homologous recombination in yeast strain PAP1500 [19] between a polymerase chain reaction (PCR) fragment encompassing nucleotides 1-2,580 from pEGFP-N1 (Clontech, USA) and pEMBLyex4 [20].…”

SLC17A9 Protein Functions as a Lysosomal ATP Transporter and Regulates Cell Viability

“…However, the mechanisms underlying ATP accumulation in those organelles remained unclear. Emerging evidence suggests that VNUT (also called SLC17A9) could be a lysosomal ATP transporter (15), but controversy remains (16). In particular, evidence supporting a lysosomal localization of endogenous SLC17A9 or lysosomal ATP transport activity via SLC17A9 was lacking.…”

Immunological Identification of Vesicular Nucleotide Transporter in Intestinal L Cells

“…[2][3][4] Recent evidence indicated that VNUT is responsible for the vesicular storage of ATP and plays an essential role in the secretion of ATP, since VNUT is expressed and localized with ATP-storing secretory vesicles such as adrenal chromaffin granules, insulin granules of islet β cells, and glutamate-containing synaptic vesicles of hippocampal neurons, and its suppression of expression by RNA interference causes decreased secretion of ATP. [2][3][4][5][6][7] Thus, it is regarded that VNUT is a potential and useful marker for sites of vesicular storage and secretion.…”