P2X4 receptor regulation of transient receptor potential melastatin type 6 (TRPM6) Mg2+ channels
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The transient receptor potential melastatin type 6 (TRPM6) ion channel regulates the body Mg2+ homeostasis by mediating transcellular Mg2+ absorption in kidney and intestine. Here, the P2X4 receptor was established as a novel regulator of TRPM6 activity. Using RT-qPCR on a mouse tissue panel, P2x4 and P2x6 were shown to be expressed in the epithelium of the colon and of the kidney, two major sites of Mg2+ reabsorption. While P2x4 was highly expressed in the colon, both P2x4 and P2x6 mRNA were prominently expressed in the distal convoluted tubule segment of the kidney, a segment with high Trpm6 expression. Using whole-cell patch clamp, an inhibitory role of P2X4 on TRPM6 activity was determined. Expression of P2X6, which does not form functional channels in mammalian cells, did not affect the function of TRPM6. The inhibition was dependent on the activity of P2X4, since a P2X4 mutant with altered ATP sensitivity was not able to inhibit TRPM6. Additionally, P2X4 was unable to inhibit TRPM7, a close homologue of TRPM6, suggesting that the inhibition is specific for TRPM6. To identify the intracellular signaling molecules that mediate the P2X4-dependent inhibition of TRPM6, the cells were treated with inhibitors of protein kinase c, protein kinase a, and phosphoinositide 3-kinase. However, none of these inhibitors prevented the inhibition of TRPM6 by P2X4. In conclusion, we propose that P2X4 receptor mediated purinergic signaling is a new regulatory mechanism of TRPM6 Mg2+ channels.
KeywordsMagnesium Colon Purinergic receptor P2X4 TRPM6
This work was supported by grants from the Netherlands Organization for Scientific Research (ZonMw 9120.8026, NWO ALW 818.02.001), a NWO Vici Grant for J.G.J. Hoenderop (016.130.668) and EURenOmics funding from the European Union seventh Framework Program (FP7/2007–2013, agreement no 305608). The authors want to state their gratitude to AnneMiete van der Kemp, Lonneke Duijkers, Fariza Bouallala, and Pauline de Bruijn for their excellent technical support. In particular, we would like to thank Dr. Sjoerd Verkaart for valuable discussions.
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