Mechanism and Molecular Determinant for Regulation of Rabbit Transient Receptor Potential Type 5 (TRPV5) Channel by Extracellular pH

Abstract

The transient receptor potential type 5 (TRPV5) channel is present in kidney and intestine and important for transepithelial (re)absorption of calcium in these tissues. We report that in whole-cell patch clamp recording extracellular acidification inhibited rabbit TRPV5 with apparent pKa ∼6.55. The two extracellular loops between the fifth and sixth transmembrane segments of TRPV5 presumably form part of the outer opening of the pore and likely are important in binding and regulation by external protons. We found that mutation of glutamate 522 to glutamine (E522Q) decreased the sensitivity of the channel to extracellular acidification. Mutations of other titratable amino acids within the two extracellular loops to non-titratable amino acids had no effect on pH sensitivity. Substitutions of aspartate or other titratable amino acids for glutamate 522 conferred an increase in pH sensitivity. The pH sensitivity mediated by glutamate 522 was independent of extracellular or intracellular Mg2+. Single channel analysis revealed that extracellular acidification reduced single channel conductance as well as open probability of the wild type channel. In contrast to wild type channel, extracellular acidification did not reduce open probability for E522Q mutant. Methanethiosulfonate reagents inhibited the activity of glutamine 522 to cysteine mutant channel with a reaction rate constant approaching that with free thiols in solution, suggesting that glutamate 522 is located on the surface of the channel. These data suggest that glutamate 522 of the rabbit TRPV5 is a “pH sensor,” and extracellular protons inhibit TRPV5 likely by altering conformation of the channel protein. The transient receptor potential type 5 (TRPV5) channel is present in kidney and intestine and important for transepithelial (re)absorption of calcium in these tissues. We report that in whole-cell patch clamp recording extracellular acidification inhibited rabbit TRPV5 with apparent pKa ∼6.55. The two extracellular loops between the fifth and sixth transmembrane segments of TRPV5 presumably form part of the outer opening of the pore and likely are important in binding and regulation by external protons. We found that mutation of glutamate 522 to glutamine (E522Q) decreased the sensitivity of the channel to extracellular acidification. Mutations of other titratable amino acids within the two extracellular loops to non-titratable amino acids had no effect on pH sensitivity. Substitutions of aspartate or other titratable amino acids for glutamate 522 conferred an increase in pH sensitivity. The pH sensitivity mediated by glutamate 522 was independent of extracellular or intracellular Mg2+. Single channel analysis revealed that extracellular acidification reduced single channel conductance as well as open probability of the wild type channel. In contrast to wild type channel, extracellular acidification did not reduce open probability for E522Q mutant. Methanethiosulfonate reagents inhibited the activity of glutamine 522 to cysteine mutant channel with a reaction rate constant approaching that with free thiols in solution, suggesting that glutamate 522 is located on the surface of the channel. These data suggest that glutamate 522 of the rabbit TRPV5 is a “pH sensor,” and extracellular protons inhibit TRPV5 likely by altering conformation of the channel protein. Calcium (Ca2+) is the most abundant cation in the human body and critical for many processes such as bone mineralization, formation of blood clots, regulation of cell-cell adhesion, and intracellular signaling (1van Os C.H. Biochim. Biophys. Acta. 1987; 906: 195-222Crossref PubMed Scopus (114) Google Scholar, 2Brown E.M. Physiol. Rev. 1991; 71: 371-411Crossref PubMed Scopus (637) Google Scholar). Although the majority of Ca2+ is present in the bone and is in continuous turnover, there is little or no net gain or loss of Ca2+ from bone in normal young and healthy adults. To maintain calcium balance, the kidney excretes the same amount of calcium absorbed by the intestine. The amount of Ca2+ excreted by the kidney is about 2% of the total filtered load (3Friedman P.A. Gesek F. Physiol. Rev. 1995; 75: 429-471Crossref PubMed Scopus (201) Google Scholar). About 98% of the filtered Ca2+ is reabsorbed by the tubule. The absorption of Ca2+ in intestine and the reabsorption in kidney occur via both paracellular and transcellular pathways. In the kidney, the transcellular reabsorption of Ca2+ occurs mainly in the distal convoluted tubule (DCT) 1The abbreviations used are: DCTdistal convoluted tubuleTRPtransient receptor potentialTRPVV type subfamily of TRP channelsCHOChinese hamster ovaryECaCepithelial Ca2+ channelCaTCa2+ transporter proteinMTSmethanethiosulfonatepHeextracellular pHBAPTA1,2-bis(2-aminophenoxy)ethane-N,N,N′,N′-tetraacetic acidMTSEAmethanethiosulfonate ethylammoniumMTSETmethanethiosulfonate ethyltrimethylammoniumMTSESmethanethiosulfonate ethylsulfonateI-Vcurrent-voltage. and accounts for ∼15-20% of total reabsorption along the tubule. The relative contribution of transcellular versus paracellular absorption of Ca2+ along the intestinal tract is less clear. distal convoluted tubule transient receptor potential V type subfamily of TRP channels Chinese hamster ovary epithelial Ca2+ channel Ca2+ transporter protein methanethiosulfonate extracellular pH 1,2-bis(2-aminophenoxy)ethane-N,N,N′,N′-tetraacetic acid methanethiosulfonate ethylammonium methanethiosulfonate ethyltrimethylammonium methanethiosulfonate ethylsulfonate current-voltage. The transcellular (re)absorption of Ca2+ is a multistep process (3Friedman P.A. Gesek F. Physiol. Rev. 1995; 75: 429-471Crossref PubMed Scopus (201) Google Scholar). It begins with passive entry of Ca2+ through the Ca2+ channels in the apical membranes followed by diffusion of Ca2+ through cytosol facilitated by binding to Ca2+-binding protein calbindin-D28K and eventually extrusion of Ca2+ across the opposing basolateral membranes. The extrusion of Ca2+ across the basolateral membranes requires energy and is mediated by Na+/Ca2+ exchangers and Ca2+-ATPases operating against the electrochemical gradient for Ca2+. It is believed that the initial step of passive entry through Ca2+ channels in the apical membranes is likely the rate-limiting step of the transepithelial Ca2+ reabsorption in the distal nephron (3Friedman P.A. Gesek F. Physiol. Rev. 1995; 75: 429-471Crossref PubMed Scopus (201) Google Scholar). Several cDNAs for apical Ca2+ channels have been recently isolated from epithelial tissues. Hoenderop et al. (4Hoenderop J.G. van der Kemp A.W. Hartog A. van de Graaf S.F. van Os C.H. Willems P.H. Bindels R.J. J. Biol. Chem. 1999; 274: 8375-8378Abstract Full Text Full Text PDF PubMed Scopus (524) Google Scholar) isolated a cDNA from rabbit kidney and named it ECaC1 (for epithelial Ca2+channel). Northern blot analysis revealed that ECaC1 message is expressed in kidney, small intestine, and placenta. In the kidney, ECaC1 is localized to the apical membranes of DCT by immunofluorescent straining (4Hoenderop J.G. van der Kemp A.W. Hartog A. van de Graaf S.F. van Os C.H. Willems P.H. Bindels R.J. J. Biol. Chem. 1999; 274: 8375-8378Abstract Full Text Full Text PDF PubMed Scopus (524) Google Scholar, 5Hoenderop J.G. Hartog A. Stuiver M. Doucet A. Willems P.H. Bindels R.J. J. Am. Soc. Nephrol. 2000; 11: 1171-1178Crossref PubMed Google Scholar). Peng et al. (6Peng J.B. Chen X.Z. Berger U.V. Vassilev P.M. Tsukaguchi H. Brown E.M. Hediger M.A. J. Biol. Chem. 1999; 274: 22739-22746Abstract Full Text Full Text PDF PubMed Scopus (539) Google Scholar, 7Peng J.B. Chen X.Z. Berger U.V. Vassilev P.M. Brown E.M. Hediger M.A. J. Biol. Chem. 2000; 275: 28186-28194Abstract Full Text Full Text PDF PubMed Scopus (141) Google Scholar) isolated CaT1 (for Ca2+transporter protein) and CaT2 from rat intestine and kidney, respectively. CaT1 is also known as ECaC2. CaT2 is the rat ortholog of ECaC1. The ECaC/CaT channels belong to the superfamily of cation-permeable ion channels known as transient receptor potential (TRP) (8Montell C. Birnbaumer L. Flockerzi V. Bindels R.J. Bruford E.A. Caterina M.J. Clapham D.E. Harteneck C. Heller S. Julius D. Kojima I. Mori Y. Penner R. Prawitt D. Scharenberg A.M. Schultz G. Shimizu N. Zhu M.X. Mol. Cell. 2002; 92: 229-231Abstract Full Text Full Text PDF Scopus (562) Google Scholar). The TRP superfamily of ion channels can be divided into several families. The TRPV family is named after its first member, capsaicin (vanilloid) receptor (9Caterina M.J. Schumacher M.A. Tominaga M. Rosen T.A. Levine J.D. Julius D. Nature. 1997; 389: 816-824Crossref PubMed Scopus (7090) Google Scholar). ECaC1/CaT2 and ECaC2/CaT1 are now known as TRPV5 and TRPV6, respectively (8Montell C. Birnbaumer L. Flockerzi V. Bindels R.J. Bruford E.A. Caterina M.J. Clapham D.E. Harteneck C. Heller S. Julius D. Kojima I. Mori Y. Penner R. Prawitt D. Scharenberg A.M. Schultz G. Shimizu N. Zhu M.X. Mol. Cell. 2002; 92: 229-231Abstract Full Text Full Text PDF Scopus (562) Google Scholar). Overall cDNAs for TRP channels encode polypeptides of ∼700-1,000 amino acids with amino acid homology. Hydrophobicity analysis of the TRPV5 and TRPV6 polypeptides predicts a transmembrane topology of an amino-terminal cytoplasmic region containing many ankyrin repeats, six membrane-spanning domains with a putative pore-forming region similar to other Ca2+-permeable channels, and a carboxyl-terminal cytoplasmic terminus containing potential regulatory sites for protein kinases (10Hoenderop J.G. Nilius B. Bindels R.J. Annu. Rev. Physiol. 2002; 64: 529-549Crossref PubMed Scopus (208) Google Scholar, 11Peng J.B. Hediger M.A. Curr. Opin. Nephrol. Hypertens. 2002; 11: 555-561Crossref PubMed Scopus (38) Google Scholar). The epithelial Ca2+ channel in the apical of kidney and intestine is likely the for regulation of calcium by and (10Hoenderop J.G. Nilius B. Bindels R.J. Annu. Rev. Physiol. 2002; 64: 529-549Crossref PubMed Scopus (208) Google Scholar, 11Peng J.B. Hediger M.A. Curr. Opin. Nephrol. Hypertens. 2002; 11: 555-561Crossref PubMed Scopus (38) Google Scholar). The activity of TRPV5 and TRPV6 channels is inhibited by extracellular pH (4Hoenderop J.G. van der Kemp A.W. Hartog A. van de Graaf S.F. van Os C.H. Willems P.H. Bindels R.J. J. Biol. Chem. 1999; 274: 8375-8378Abstract Full Text Full Text PDF PubMed Scopus (524) Google Scholar, J.B. Chen X.Z. Berger U.V. Vassilev P.M. Tsukaguchi H. Brown E.M. Hediger M.A. J. Biol. Chem. 1999; 274: 22739-22746Abstract Full Text Full Text PDF PubMed Scopus (539) Google Scholar, 7Peng J.B. Chen X.Z. Berger U.V. Vassilev P.M. Brown E.M. Hediger M.A. J. Biol. Chem. 2000; 275: 28186-28194Abstract Full Text Full Text PDF PubMed Scopus (141) Google Scholar, R. J. Hoenderop J.G. Bindels R.J. G. Nilius B. PubMed Scopus Google Scholar). of TRPV5 and TRPV6 by the increase in Ca2+ by kidney in in pH Full Text PDF PubMed Scopus Google Scholar). In the present the and of regulation of TRPV5 by of cDNAs for rabbit (4Hoenderop J.G. van der Kemp A.W. Hartog A. van de Graaf S.F. van Os C.H. Willems P.H. Bindels R.J. J. Biol. Chem. 1999; 274: 8375-8378Abstract Full Text Full Text PDF PubMed Scopus (524) Google Scholar) and human TRPV5 J.B. Brown E.M. Hediger M.A. PubMed Scopus Google Scholar) by reaction rabbit and human kidney cDNA as by and into for transient in Chinese hamster ovary of TRPV5 was a from and by S. Nature. PubMed Scopus Google Scholar, S. A. 1999; PubMed Scopus Google Scholar). and in was in containing with cDNA for cDNAs for wild type or mutant TRPV5 and the by the About after by and in a for with an patch clamp as V. B. R. P.A. Am. J. 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Chem. 2000; 275: Full Text Full Text PDF PubMed Scopus Google Scholar). with for wild type or mutant TRPV5 in containing for and for was by a in an containing pH as and for the to and with containing 5 in and in for for was and the apparent rate constant for of channels by the constant was by the of of channels by reagents with a single The rate constant was by the of the constant by the of Zhu J. S. A. 1999; PubMed Scopus (38) Google Scholar). To the sensitivity of the channel to by extracellular relative with a the J. Biol. Chem. 2000; 275: Full Text Full Text PDF PubMed Scopus Google Scholar). are as of a of was of TRPV5 by activity of rabbit TRPV5 expressed in was whole-cell recording to other Ca2+ channels, TRPV5 Ca2+ are from R. J. Hoenderop J.G. Bindels R.J. G. Nilius B. PubMed Scopus Google Scholar). Ca2+ entry through TRPV5 intracellular Ca2+ in a the cytoplasmic of the pore and B. J. R. Hoenderop J.G. Bindels R.J. G. PubMed Scopus Google Scholar, B. J. Hoenderop J.G. R. S. G. Bindels R.J. J. Biol. Chem. 2002; Full Text Full Text PDF PubMed Scopus Google Scholar). We used as for to the effect of on TRPV5 activity from other potential through of a to from through TRPV5 and for of of for in did not the or potential The of on TRPV5 by in to pH in and from to a in The was as was The apparent pKa for acid of the was of apparent pKa is within the of pH in the of DCT in and J. Scopus Google Scholar). of of TRPV5 in of the by transmembrane topology of TRPV5 cation channels and six domains and a putative pore region between the fifth and sixth domains J.B. Hediger M.A. Curr. Opin. Nephrol. Hypertens. 2002; 11: 555-561Crossref PubMed Scopus (38) Google Scholar, R. J. Hoenderop J.G. Bindels R.J. G. Nilius B. PubMed Scopus Google Scholar). The two extracellular loops between the putative pore region and the two transmembrane domains presumably form part of the outer of the pore amino acids in these loops are likely for and the regulation of channel by protons. To the amino acids for of the titratable amino acids in the two extracellular loops to a non-titratable and the sensitivity of of the to by extracellular acidification We found that mutation of glutamate 522 to glutamine (E522Q) decreased the sensitivity of the channel to by extracellular acidification The apparent pKa for acid was pH by pH for versus for These suggest that of glutamate 522 by protons is in regulation of The pKa of free acid is C. Chem. Scholar). The between the pKa of TRPV5 conferred by glutamate 522 and that of a free glutamate in is likely to the that titratable of amino acids within are by the L. Nature. PubMed Scopus Google Scholar, Tominaga M. Julius D. S. A. 2000; PubMed Scopus Google Scholar). Mutations of other titratable amino acids did not sensitivity of the channels and of to mutant in and the on mutant not of by of TRPV5 is to Ca2+ J.B. Hediger M.A. Curr. Opin. Nephrol. Hypertens. 2002; 11: 555-561Crossref PubMed Scopus (38) Google Scholar, R. J. Hoenderop J.G. Bindels R.J. G. Nilius B. PubMed Scopus Google Scholar). We the of on Ca2+ through We Ca2+ through the channel in was used for of cDNAs for ECaC/CaT (4Hoenderop J.G. van der Kemp A.W. Hartog A. van de Graaf S.F. van Os C.H. Willems P.H. Bindels R.J. J. Biol. Chem. 1999; 274: 8375-8378Abstract Full Text Full Text PDF PubMed Scopus (524) Google Scholar, J.B. Chen X.Z. Berger U.V. Vassilev P.M. Tsukaguchi H. Brown E.M. Hediger M.A. J. Biol. Chem. 1999; 274: 22739-22746Abstract Full Text Full Text PDF PubMed Scopus (539) Google Scholar). We the for of its and TRPV5 in and for and with for of as TRPV5 did not was inhibited by to the of whole-cell recording extracellular acidification from to decreased of glutamate 522 the sensitivity of pH for versus for of of for 522 on the of of glutamate 522 for of TRPV5 by glutamate 522 by several amino acids with a pKa and in the titratable The pKa and of titratable of these amino acids are in and aspartate have the same titratable glutamate 522 to aspartate mutant had the same sensitivity as the wild type TRPV5 and acids and are also titratable by protons. We found that the apparent pKa for regulation of and and respectively and These pKa that for E522Q mutant versus pKa for the non-titratable with did not from to did not in the of for is not to a pKa for the mutant. are to the for of for mutant. The amino acid glutamate 522 of rabbit TRPV5 is not in human TRPV5 J.B. Brown E.M. Hediger M.A. PubMed Scopus Google Scholar, J.G. R. D. J. G. Bindels R.J. B. J. Physiol. PubMed Scopus Google Scholar). The human TRPV5 the non-titratable glutamine amino acid We found that the pKa for of human TRPV5 was not from that of E522Q mutant of rabbit TRPV5 versus not Overall these the that of amino acid the of glutamate 522 to of TRPV5 channel by extracellular protons. between apparent pKa for of TRPV5 and that of free amino acids in are likely by L. Nature. PubMed Scopus Google Scholar, Tominaga M. Julius D. S. A. 2000; PubMed Scopus Google Scholar). of on Single of channel conductance of TRPV5 was by single channel extracellular pH decreased as potential was reduced from to The was to a conductance of The of on single channel conductance by recording pH pH single channel conductance was pH pH and reduced to and respectively. with the in whole-cell the in single channel conductance by extracellular acidification was in whole-cell versus in single channel conductance by acidification from to no in the of channels, of whole-cell is to in single channel conductance open The suggest that extracellular acidification both single channel conductance and open probability of be in and in open probability is mediated by glutamate We the of extracellular acidification on single channel conductance on E522Q mutant. We found that single channel conductance was in E522Q mutant relative to wild type acidification also decreased the single channel conductance on E522Q by an similar to that on wild type In contrast to that for wild the of in single channel conductance and in whole-cell by extracellular acidification was the same for E522Q the of single channel conductance and whole-cell relative to for wild type TRPV5 and for E522Q in the effect of extracellular acidification for the two These suggest that of glutamate 522 of TRPV5 by protons is for in open probability not in single channel conductance of the channel. The and for in single channel conductance of TRPV5 of on and E522Q in the of and and extracellular to TRPV5 B. R. J. Hoenderop J.G. Bindels R.J. G. J. Physiol. 2000; PubMed Scopus Google Scholar, J. A. R. H. Hoenderop J.G. Bindels R.J. G. Penner R. Nilius B. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). The for intracellular is not The extracellular of TRPV5 by binding to aspartate in the putative pore region B. R. J. Hoenderop J.G. G. Bindels R.J. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). To extracellular acidification conformation of intracellular extracellular binding to increase of the channel, from both of the by in the and in the in whole-cell recording by after formation of whole-cell recording the of TRPV5 of less apparent as in the by in the of on by to for and from to the was to acidification from to reduced to of the acidification of The of and of was from to 5 for The pKa for of by extracellular acidification was not from that in the of We the effect of extracellular acidification on E522Q mutant in the of and intracellular Mg2+. The potential for E522Q was not from that for wild type with wild extracellular acidification a in on E522Q also on The and for E522Q 5 The pKa for of of E522Q by extracellular acidification was not from that in the of Overall these suggest that of TRPV5 from of glutamate 522 is independent of Mg2+. of on methanethiosulfonate reagents with the of cysteine Zhu J. S. A. 1999; PubMed Scopus (38) Google Scholar, Y. F. J. Biol. Chem. 2002; Full Text Full Text PDF PubMed Scopus Google Scholar). of by to of methanethiosulfonate ethylammonium to the a of The rate constant for was from did not by of to with the of of cysteine by type TRPV5 not inhibited by in the that by occurs the cysteine We the of reagents on Methanethiosulfonate ethyltrimethylammonium is versus Y. F. J. Biol. Chem. 2002; Full Text Full Text PDF PubMed Scopus Google and A. PubMed Scopus Google Scholar). Methanethiosulfonate ethylsulfonate a and is in between and Y. F. J. Biol. Chem. 2002; Full Text Full Text PDF PubMed Scopus Google Scholar, A. PubMed Scopus Google Scholar). The reaction rate for reagents with free thiols in or in for both and and for respectively A. PubMed Scopus Google Scholar). We found that inhibited with a rate similar to versus not The similar rate of by and a in the that the amino acid 522 is not located in the part of the ion with and the rate of by was of of by reagents is in with the rate for reaction of these reagents with free thiols in The rate constant for of the reaction rate of the reagents with free thiols in solution, suggesting that amino acid 522 on the surface of The rate constant for reagents to cysteine in the cytoplasmic and the pore region of ion channels is of D. Full Text Full Text PDF PubMed Scopus Google Scholar). To for with the reaction rate of of the cysteine a cysteine in two of these is glutamate is believed to in the pore region B. R. J. Hoenderop J.G. G. Bindels R.J. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). The other is is believed to be within the fifth transmembrane J.G. R. D. J. G. Bindels R.J. B. J. Physiol. PubMed Scopus Google Scholar) or in the between the fifth transmembrane and the to the outer J.G. van der Kemp A.W. Hartog A. van de Graaf S.F. van Os C.H. Willems P.H. Bindels R.J. J. Biol. Chem. 1999; 274: 8375-8378Abstract Full Text Full Text PDF PubMed Scopus (524) Google and We found that had no effect on suggesting that the of cysteine amino acid is not to the extracellular of had no on of inhibited The rate constant for of was that for of These are with the that glutamate 522 on the surface and glutamate in the pore region of the channel. of ion channels by pH occur via of the channel or be mediated by other In the present report that extracellular protons inhibit TRPV5 by glutamate 522 in the extracellular between the putative fifth transmembrane and the pore The of of glutamate 522 in and of TRPV5 is from a in sensitivity by mutation to The of other titratable amino acids to for glutamate 522 in sensitivity of amino acids by protons can in protein Several of suggest that of glutamate 522 TRPV5 activity by altering protein it open probability of the channel. amino acids with and a of der and for of and J. Mol. Biol. PubMed Scopus Google Scholar) can for glutamate der These suggest that glutamate 522 is not likely located in the region of the of the is by the cysteine These suggest that the in activity of the channel by of glutamate 522 is to of channel of of glutamate is of its in the extracellular be important in a of A. L. A. G. J. Physiol. 2002; PubMed Scopus Google Scholar). of channels by external protons is to be to of and in the extracellular A. L. A. G. J. Physiol. 2002; PubMed Scopus Google Scholar). The single channel conductance for E522Q mutant wild is with the that the of glutamate 522 be important for ion other titratable amino acids and can for glutamate 522 in an increase in pH sensitivity sensitivity is in pH pKa and and are pKa and in These suggest that the in channel activity from of glutamate 522 is to protein conformation of The for of TRPV5 is not for channels is to by intracellular and J. Physiol. 1995; PubMed Scopus Google Scholar). have that intracellular by not of TRPV6 J. A. R. H. Hoenderop J.G. Bindels R.J. G. Penner R. Nilius B. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google suggesting that other as by also be It is that in the for TRPV5 to with in and an extracellular acidification of the channel. It been that intracellular pH for via of a in the cytoplasmic carboxyl-terminal B. J. 1999; PubMed Scopus Google Scholar). The by extracellular pH of TRPV5 be an for The amino acid glutamate 522 of rabbit TRPV5 is not in human TRPV5 J.B. Brown E.M. Hediger M.A. PubMed Scopus Google Scholar, J.G. R. D. J. G. Bindels R.J. B. J. Physiol. PubMed Scopus Google Scholar). In human the amino acid is a non-titratable the pH mediated by the amino acid 522 is likely also important for regulation of epithelial Ca2+ in TRPV6, an of TRPV5 also present in kidney and intestine, a titratable the to the amino acid 522 of rabbit TRPV5 J.G. R. D. J. G. Bindels R.J. B. J. Physiol. PubMed Scopus Google Scholar). from the TRPV6 is likely as to as the rabbit found that the pKa for of TRPV6 was not from that of rabbit TRPV5 Hoenderop et al. J.G. S. F. J. Nilius B. Bindels R.J. J. PubMed Scopus Google Scholar) recently that TRPV5 and TRPV6 form and that have and of TRPV5 and It is likely that Ca2+ channels in human kidney and intestine are of TRPV5 and The from TRPV6 to pH for the Ca2+ Full Text PDF PubMed Scopus Google Scholar). protein pH acid and Ca2+ L. J. PubMed Scopus Google Scholar, L. Am. J. 2002; Full Text Full Text PDF PubMed Scopus Google Scholar). increase in Ca2+ to formation of kidney M. R. PubMed Scopus Google Scholar). of in normal human Ca2+ M. Full Text PDF PubMed Scopus Google Scholar). extracellular intracellular acid Ca2+ in and in protein is not The pH of in DCT is in the of B. J. Hoenderop J.G. R. S. G. Bindels R.J. J. Biol. Chem. 2002; Full Text Full Text PDF PubMed Scopus Google Scholar). The amount of Ca2+ excreted by the kidney is of the total filtered the 98% of the total filtered Ca2+ reabsorbed by the the DCT is for We found that extracellular acidification from pH to in activity for channels containing a titratable glutamate in rabbit or in human and for channels containing the non-titratable glutamine in human that Ca2+ channels are of and have an sensitivity of in channel activity pH acidification. acidification from to increase Ca2+ from to of the total filtered These the that of Ca2+ reabsorption in DCT by extracellular protons to increase in Ca2+ in the of with a pH and protein We J. M. Bindels for the of the and and Y. C. for and critical of the