Klaus M. Giehl

Rubrospinal neurons (RSNs) undergo a marked atrophy in the second week after cervical axotomy. This delayed atrophy is accompanied by a decline in the expression of regeneration-associated genes such as GAP-43 and Talpha1-tubulin, which are initially elevated after injury. These responses may reflect a deficiency in the trophic support of axotomized RSNs. To test this hypothesis, we first analyzed the expression of mRNAs encoding the trk family of neurotrophin receptors. In situ hybridization revealed expression of full-length trkB receptors in virtually all RSNs, which declined 7 d after axotomy. Full-length trkC mRNA was expressed at low levels. Using RT-PCR, we found that mRNAs encoding trkC isoforms with kinase domain inserts were present at levels comparable to that for the unmodified receptor. TrkA mRNA expression was not detected in RSNs, and the expression of p75 was restricted to a small subpopulation of axotomized cells. In agreement with the pattern of trk receptor expression, infusion of recombinant human BDNF or NT-4/5 into the vicinity of the axotomized RSNs, between days 7 and 14 after axotomy, fully prevented their atrophy. This effect was still evident 2 weeks after the termination of BDNF treatment. Moreover, BDNF or NT-4/5 treatment stimulated the expression of GAP-43 and Talpha1-tubulin mRNA and maintained the level of trkB expression. Vehicle, NGF, or NT-3 treatment had no significant effect on cell size or GAP-43 and Talpha1-tubulin expression. In a separate experiment, infusion of BDNF also was found to increase the number of axotomized RSNs that regenerated into a peripheral nerve graft. Thus, in BDNF-treated animals, the prevention of neuronal atrophy and the stimulation GAP-43 and Talpha1-tubulin expression is correlated with an increased regenerative capacity of axotomized RSNs.

The unprocessed precursor of the neurotrophin nerve growth factor (NGF), proNGF, has been suggested to be a death-inducing ligand for the neurotrophin receptor p75. Whether proNGF is a true pathophysiological ligand that is secreted, binds p75, and activates cell death in vivo, however, has remained unknown. Here, we report that after brain injury, proNGF was induced and secreted in an active form capable of triggering apoptosis in culture. We further demonstrate that proNGF binds p75 in vivo and that disruption of this binding results in complete rescue of injured adult corticospinal neurons. These data together suggest that proNGF binding to p75 is responsible for the death of adult corticospinal neurons after lesion, and they help to establish proNGF as the pathophysiological ligand that activates the cell-death program by means of p75 after brain injury. Interference in the binding of proNGF to p75 may provide a therapeutic approach for the treatment of disorders involving neuronal loss.

Brain-derived neurotrophic factor (BDNF) and neurotrophin-3 (NT-3) have been identified as survival factors for adult axotomized rat corticospinal neurons (CSN) in vivo. Axotomy of corticospinal neurons at the level of the internal capsule induced death of 46% of the CSN within the first week after axotomy. The surviving population of CSN displayed severe atrophy with mean cross-sectional area 49% of their unlesioned contralateral counterparts 7 days after axotomy. Using in situ hybridization to assess the expression of the receptors for the family of neurotrophins, we found trkB and trkC but not trkA mRNA expression in CSN. Intraparenchymal application of BDNF or NT-3 at doses of 12 microg/day for 7 days via an osmotic minipump fully prevented the axotomy-induced death of CSN. Interestingly, no neuronal atrophy was seen after BDNF application while NT-3 had only a partial effect on the size of the axotomized CSN. Nerve growth factor did not prevent death or cell atrophy, consistent with lack of trkA mRNA expression in these neurons. These findings show that BDNF and NT-3 are survival factors for adult rat CSN in vivo, and may contribute to the development of therapeutic strategies aiming at the prevention of CSN degeneration in human motor neuron diseases.

TRPM3 is a poorly understood member of the large family of transient receptor potential (TRP) ion channels. Here we describe five novel splice variants of TRPM3, TRPM3α1–5. These variants are characterized by a previously unknown amino terminus of 61 residues. The differences between the five variants arise through splice events at three different sites. One of these splice sites might be located in the pore region of the channel as indicated by sequence alignment with other, better-characterized TRP channels. We selected two splice variants, TRPM3α1 and TRPM3α2, that differ only in this presumed pore region and analyzed their biophysical characteristics after heterologous expression in human embryonic kidney 293 cells. TRPM3α1 as well as TRPM3α2 induced a novel, outwardly rectifying cationic conductance that was tightly regulated by intracellular Mg2+. However, these two variants are highly different in their ionic selectivity. Whereas TRPM3α1-encoded channels are poorly permeable for divalent cations, TRPM3α2-encoded channels are well permeated by Ca2+ and Mg2+. Additionally, we found that currents through TRPM3α2 are blocked by extracellular monovalent cations, whereas currents through TRPM3α1 are not. These differences unambiguously show that TRPM3 proteins constitute a pore-forming channel subunit and localize the position of the ion-conducting pore within the TRPM3 protein. Although the ionic selectivity of ion channels has traditionally been regarded as rather constant for a given channel-encoding gene, our results show that alternative splicing can be a mechanism to produce channels with very different selectivity profiles. TRPM3 is a poorly understood member of the large family of transient receptor potential (TRP) ion channels. Here we describe five novel splice variants of TRPM3, TRPM3α1–5. These variants are characterized by a previously unknown amino terminus of 61 residues. The differences between the five variants arise through splice events at three different sites. One of these splice sites might be located in the pore region of the channel as indicated by sequence alignment with other, better-characterized TRP channels. We selected two splice variants, TRPM3α1 and TRPM3α2, that differ only in this presumed pore region and analyzed their biophysical characteristics after heterologous expression in human embryonic kidney 293 cells. TRPM3α1 as well as TRPM3α2 induced a novel, outwardly rectifying cationic conductance that was tightly regulated by intracellular Mg2+. However, these two variants are highly different in their ionic selectivity. Whereas TRPM3α1-encoded channels are poorly permeable for divalent cations, TRPM3α2-encoded channels are well permeated by Ca2+ and Mg2+. Additionally, we found that currents through TRPM3α2 are blocked by extracellular monovalent cations, whereas currents through TRPM3α1 are not. These differences unambiguously show that TRPM3 proteins constitute a pore-forming channel subunit and localize the position of the ion-conducting pore within the TRPM3 protein. Although the ionic selectivity of ion channels has traditionally been regarded as rather constant for a given channel-encoding gene, our results show that alternative splicing can be a mechanism to produce channels with very different selectivity profiles. The transient receptor potential (TRP) 1The abbreviations used are: TRP, transient receptor potential; TRPM, TRP melastatin subfamily; TRPV, TRP vanilloid receptor subfamily; GFP, green fluorescent protein; EYFP, enhanced yellow fluorescent protein; HEK-293, human embryonic kidney 293; MagNuM, magnesium nucleotide-regulated metal ion current; MIC, magnesium-inhibited cation current. gene family comprises at least 28 mammalian genes divided into seven subfamilies (1Clapham D.E. Nature. 2003; 426: 517-524Crossref PubMed Scopus (2164) Google Scholar, 2Montell C. Science's STKE 2005. 2005; : re3Google Scholar). Most of the encoded proteins exhibit common structural features such as six predicted transmembrane (TM) domains with a putative pore loop between TM5 and TM6 and the so-called TRP box after TM6 (1Clapham D.E. Nature. 2003; 426: 517-524Crossref PubMed Scopus (2164) Google Scholar, 2Montell C. Science's STKE 2005. 2005; : re3Google Scholar). Although all members of this group have been reported to form cationic channels, their mechanisms of activation, their regulation, and their biological functions are remarkably diverse. They also display a large variety of different cation selectivities (1Clapham D.E. Nature. 2003; 426: 517-524Crossref PubMed Scopus (2164) Google Scholar, 2Montell C. Science's STKE 2005. 2005; : re3Google Scholar). For example, TRPM4 and TRPM5 have been described as impermeable for divalent cations (3Launay P. Fleig A. Perraud A.L. Scharenberg A.M. Penner R. Kinet J.P. Cell. 2002; 109: 397-407Abstract Full Text Full Text PDF PubMed Scopus (550) Google Scholar, 4Nilius B. Prenen J. Droogmans G. Voets T. Vennekens R. Freichel M. Wissenbach U. Flockerzi V. J. Biol. Chem. 2003; 278: 30813-30820Abstract Full Text Full Text PDF PubMed Scopus (271) Google Scholar, 5Hofmann T. Chubanov V. Gudermann T. Montell C. Curr. Biol. 2003; 13: 1153-1158Abstract Full Text Full Text PDF PubMed Scopus (326) Google Scholar), whereas TRPV5 and TRPV6 appear to be exclusively permeable for Ca2+ (6Vennekens R. Hoenderop J.G. Prenen J. Stuiver M. Willems P.H. Droogmans G. Nilius B. Bindels R.J. J. Biol. Chem. 2000; 275: 3963-3969Abstract Full Text Full Text PDF PubMed Scopus (274) Google Scholar, 7Wissenbach U. Niemeyer B. Fixemer T. Schneidewind A. Trost C. Cavalié A. Reuss K. Meese E. Bonkhoff H. Flockerzi V. J. Biol. Chem. 2001; 276: 19461-19468Abstract Full Text Full Text PDF PubMed Scopus (240) Google Scholar). The diversity of TRP channels is further increased by the fact that most members of the TRP gene family can give rise to several different transcripts due to alternative splicing (8Lis A. Wissenbach U. Philipp S.E. Naunyn-Schmiedeberg′s Arch. Pharmacol. 2005; (in press)PubMed Google Scholar). In a few cases, the functional consequences of these alternative splice events are now beginning to emerge. For example, miss-plicing of TRPM6 transcripts is associated with a hereditary disorder called hypomagnesemia with secondary hypocalcemia (9Schlingmann K.P. Weber S. Peters M. Niemann N.L. Vitzthum H. Klingel K. Kratz M. Haddad E. Ristoff E. Dinour D. Syrrou M. Nielsen S. Sassen M. Waldegger S. Seyberth H.W. Konrad M. Nat. Genet. 2002; 31: 166-170Crossref PubMed Scopus (655) Google Scholar, 10Walder R.Y. Landau D. Meyer P. Shalev H. Tsolia M. Borochowitz Z. Boettger M.B. Beck G.E. Englehardt R.K. Carmi R. Sheffield V.C. Nat. Genet. 2002; 31: 171-174Crossref PubMed Scopus (476) Google Scholar), and an amino-terminal-truncated variant of TRPM4 appears to modulate Ca2+ oscillations after receptor stimulation in T lymphocytes (11Launay P. Cheng H. Srivatsan S. Penner R. Fleig A. Kinet J.P. Science. 2004; 306: 1374-1377Crossref PubMed Scopus (258) Google Scholar). However, up to now, the largest number of different splice variants for any TRP family member has been described for TRPM3 (12Lee N. Chen J. Wu S. Sun L. Huang M. Levesque P.C. Rich A. Feder J.N. Gray K.R. Lin J.H. Janovitz E.B. Blanar M.A. J. Biol. Chem. 2003; 278: 20890-20897Abstract Full Text Full Text PDF PubMed Scopus (164) Google Scholar, 13Grimm C.M. Kraft R. Sauerbruch S. Schultz G. Harteneck C. J. Biol. Chem. 2003; 278: 21493-21501Abstract Full Text Full Text PDF PubMed Scopus (277) Google Scholar). Lee et al. (12Lee N. Chen J. Wu S. Sun L. Huang M. Levesque P.C. Rich A. Feder J.N. Gray K.R. Lin J.H. Janovitz E.B. Blanar M.A. J. Biol. Chem. 2003; 278: 20890-20897Abstract Full Text Full Text PDF PubMed Scopus (164) Google Scholar) reported six splice variants of human TRPM3, which they named TRPM3a–f. Their lengths range from 1544 to 1579 amino acid residues. Functional data were only reported for the TRPM3a splice variant. Heterologously expressed TRPM3a exhibits a constitutive, Ca2+ concentration-dependent Ca2+ entry that can be augmented by Ca2+ store depletion or by stimulation of muscarinic receptors (12Lee N. Chen J. Wu S. Sun L. Huang M. Levesque P.C. Rich A. Feder J.N. Gray K.R. Lin J.H. Janovitz E.B. Blanar M.A. J. Biol. Chem. 2003; 278: 20890-20897Abstract Full Text Full Text PDF PubMed Scopus (164) Google Scholar). In addition, Grimm et al. (13Grimm C.M. Kraft R. Sauerbruch S. Schultz G. Harteneck C. J. Biol. Chem. 2003; 278: 21493-21501Abstract Full Text Full Text PDF PubMed Scopus (277) Google Scholar) reported one further human variant that has a total of only 1325 residues because of its considerably shorter carboxyl terminus (13Grimm C.M. Kraft R. Sauerbruch S. Schultz G. Harteneck C. J. Biol. Chem. 2003; 278: 21493-21501Abstract Full Text Full Text PDF PubMed Scopus (277) Google Scholar). On its amino terminal end, however, this variant, which we will refer to as TRPM31325 throughout this report, possesses an additional 153 amino acids completely missing in the other reported TRPM3 variants. As is the case with TRPM3a, TRPM31325 generates constitutively active, Ca2+-permeable channels when heterologously expressed in human embryonic kidney 293 (HEK-293) cells. The activity of TRPM31325 channels can reportedly be enhanced by hypo-osmotically induced cell swelling (13Grimm C.M. Kraft R. Sauerbruch S. Schultz G. Harteneck C. J. Biol. Chem. 2003; 278: 21493-21501Abstract Full Text Full Text PDF PubMed Scopus (277) Google Scholar) and by d-erythro-sphingosine (14Grimm C. Kraft R. Schultz G. Harteneck C. Mol. Pharmacol. 2005; 67: 798-805Crossref PubMed Scopus (131) Google Scholar). In this report we describe five novel TRPM3 splice variants that we cloned as full-length cDNA constructs from mouse brain. These variants are 1699–1721 amino acid residues long and are characterized by a novel amino terminal sequence of 61 residues not described so far. Interestingly, two of the novel variants, designated TRPM3α1 and TRPMα2, differ only in a region between the fifth and transmembrane domains the pore-forming region of the channel is region is amino acids in TRPM3α1 with a is by an The of our is that this of the a large in the ionic selectivity of the ion channels. We alternative splicing as a mechanism to the selectivity of TRPM3 channels. of TRPM3 and were used to transcripts after of mouse total For we used the and the of the at by at and at of five three TRPM3α2, and three and one was found for splice variant We the and a to the of and which were cloned into J. Philipp S. S. Flockerzi V. Cavalié A. J. Scopus Google Scholar) for transient expression of TRPM3 with the green fluorescent the TRPM3α1 and TRPM3α2 were to the enhanced yellow fluorescent cDNA in the expression of The of and have been to and and of were in with in a were a they For transient to were with TRPM3α2, or were used or after to cell For of or were for in were by cell a cell and were for their and for expression of TRPM3 in and of TRPM3 and and the sequence of the mouse were by with and as for were to with at for they were and with a cDNA of the human as In were of K. P. PubMed Scopus Google Scholar) and from or with from were for for in three in and and in and and with terminal the of with a of in were to for and to for were as described S. Trost C. J. J. N. G. A. M. Flockerzi V. J. Biol. Chem. 2000; 275: Full Text Full Text PDF Scopus Google Scholar). from to were in of and analyzed and cell were with an by of in and were with were by their green an and used after were at and the was at with from to that the between was increased to the the at and and the potential were by (in or or was to the The were with and are indicated in the was to and and to with or potential of was were the at in extracellular were was to of cation and and to or were for by a was at from the of extracellular The (in of the that were used for extracellular in of in a of were as described previously S. J. L. G. Freichel M. M. Cavalié A. Flockerzi V. J. PubMed Scopus Google Scholar). In and cell proteins were to and an were with for at in (in with for and analyzed in a were and analyzed were to Ca2+ after of the a and S.E. were from from and with a with the data used in or Ca2+ were from are given as and the was used to and of TRPM3 from we two human TRPM3 cDNA and from a gene human The mouse gene a highly with 28 and mouse gene number and were from the mouse Their are located and to an with a predicted that is also in the human cDNA of this cDNA from mouse total were by and were from mouse not the sequence of mouse We five different splice variants, designated TRPM3α2, and Their amino acid up to amino which are encoded by and The predicted and gene number are not in the cDNA of these TRPM3 variants from amino acid residues encoded by the have been in the described human variant (12Lee N. Chen J. Wu S. Sun L. Huang M. Levesque P.C. Rich A. Feder J.N. Gray K.R. Lin J.H. Janovitz E.B. Blanar M.A. J. Biol. Chem. 2003; 278: 20890-20897Abstract Full Text Full Text PDF PubMed Scopus (164) Google Scholar). a acid sequence that to the amino terminal of the human TRPM31325 variant (13Grimm C.M. Kraft R. Sauerbruch S. Schultz G. Harteneck C. J. Biol. Chem. 2003; 278: 21493-21501Abstract Full Text Full Text PDF PubMed Scopus (277) Google Scholar) is in the variants of for a completely novel amino terminal sequence that is not in any of the described human TRPM3 variants. mouse cDNA we not a one that with that these are expressed in a be by tightly regulated or by expression of these variants from alternative from however, mouse TRPM3 proteins show amino acid sequence to the human variants. mouse TRPM3α1 to human to to to and to The between the mouse and human transcripts that not only the amino acid sequence also the splice events are highly between mouse and is highly of functional of TRPM3 proteins also of their and by alternative of analyzed the expression of the mouse gene by and by in We found expression in and with transcripts of and In with results (13Grimm C.M. Kraft R. Sauerbruch S. Schultz G. Harteneck C. J. Biol. Chem. 2003; 278: 21493-21501Abstract Full Text Full Text PDF PubMed Scopus (277) Google Scholar), we not transcripts in mouse kidney by not In transcripts in several of the mouse such as the the the and the and expression was found in the of transcripts be in of total TRPM3α1 and TRPM3α2 by splicing within of the mouse and the human TRPM3 gene to the of additional amino acid residues and the additional of an by a (12Lee N. Chen J. Wu S. Sun L. Huang M. Levesque P.C. Rich A. Feder J.N. Gray K.R. Lin J.H. Janovitz E.B. Blanar M.A. J. Biol. Chem. 2003; 278: 20890-20897Abstract Full Text Full Text PDF PubMed Scopus (164) Google Scholar). Interestingly, this which is in and (12Lee N. Chen J. Wu S. Sun L. Huang M. Levesque P.C. Rich A. Feder J.N. Gray K.R. Lin J.H. Janovitz E.B. Blanar M.A. J. Biol. Chem. 2003; 278: 20890-20897Abstract Full Text Full Text PDF PubMed Scopus (164) Google Scholar) in all other variants, between the presumed fifth and transmembrane domains In to the members of the T. Nilius B. 2003; PubMed Scopus Google Scholar), is that this of the to the pore-forming region of the channel The variants TRPM3α1 and TRPM3α2 differ only in this region and We to this in the sequence the biophysical characteristics of the ion channels. this end, we expressed TRPM3 variants in which not TRPM3 with we that not exhibit a constitutively active, outwardly rectifying current. was after the cell and the extracellular cations by the cation the potential from to that the was by cations we used currents increased within the of cell and when we increased the in the to and currents in the currents of were These data that TRPM3α1 is regulated by of intracellular which is found to be in the of A.M. A. 2000; PubMed Google Scholar). We to TRPM3α2 and this variant also a functional In extracellular TRPM3α2 currents were and increased only However, when we all extracellular cations with we large which as as were and currents were not in The of currents these to the conductance is permeable to We intracellular also regulated TRPM3α2 channels. to our results with we found large currents in the of intracellular currents with in the by intracellular appears to be a common of the two splice variants. by intracellular however, is not a of TRPM3 channels has also been described for the ion channels TRPM6 T. Nilius B. S. Droogmans G. Bindels R.J. Hoenderop J.G. J. Biol. Chem. 2004; Full Text Full Text PDF PubMed Scopus Google Scholar) and K. Perraud A.L. T. Kinet J.P. Penner R. Scharenberg A.M. Fleig A. Nature. 2001; PubMed Scopus Google Scholar). has been to be expressed in and is to be the of an that has been named K. Perraud A.L. T. Kinet J.P. Penner R. Scharenberg A.M. Fleig A. Nature. 2001; PubMed Scopus Google Scholar) or J. 2002; PubMed Scopus Google Scholar). In the we have several to that the of the not our We that in were in and and with divalent extracellular were within of the cell and and that the is very K. Perraud A.L. T. Kinet J.P. Penner R. Scharenberg A.M. Fleig A. Nature. 2001; PubMed Scopus Google and data not For the in we to the any of in with the in not these however, the that the outwardly rectifying currents not through TRPM3 proteins from of channels. We however, that the currents in a to to described for and B. Prenen J. Droogmans G. Voets T. Vennekens R. Freichel M. Wissenbach U. Flockerzi V. J. Biol. Chem. 2003; 278: 30813-30820Abstract Full Text Full Text PDF PubMed Scopus (271) Google Scholar, 5Hofmann T. Chubanov V. Gudermann T. Montell C. Curr. Biol. 2003; 13: 1153-1158Abstract Full Text Full Text PDF PubMed Scopus (326) Google Scholar, D. Nature. 2002; PubMed Scopus Google Scholar). encoded by or TRPM6 not show such a T. Nilius B. S. Droogmans G. Bindels R.J. Hoenderop J.G. J. Biol. Chem. 2004; Full Text Full Text PDF PubMed Scopus Google Scholar, L. D.E. Science. 2001; PubMed Scopus Google Scholar). when we to an extracellular only the large a of the be TRPM3α1 and TRPM3α2 channels show a of biophysical that are not found in any other member of the channel the of TRPM3 the ionic selectivity of the TRPM3α1 and TRPM3α2 splice variants, we currents and analyzed the potential that the potential is in TRPM3α2 with when Ca2+ and are the only cations in the extracellular results were with of of the the B. of Scholar) that TRPM3α2 channels are at least permeable for Ca2+ and at least permeable for TRPM3α1 channels large in ion that the ion-conducting pore of TRPM3 channels is by the sequence differences between the two splice variants and that this region of the is of the ion-conducting pore of this that the currents after TRPM3 proteins are by these proteins and are not the of through selectivity for divalent cations of the TRPM3α2 variant not that this variant is of divalent However, when we extracellular of Ca2+ or currents of up to currents were in in cells. These data in to TRPM3α2-encoded channels not only have a rather selectivity for divalent cations are also of divalent TRPM3 by of TRPM3α1 and TRPM3α2 currents in extracellular and a as the only cation that currents through TRPM3α2 are by cations in the and We were by the fact that this only currents through TRPM3α2 not through We the of this that only one permeable cation in to We found that Ca2+ currents through TRPM3α2 as well as through TRPM3α1 and results were for On the other the monovalent cations and TRPM3α2 whereas of currents through TRPM3α1 and TRPM3α2 is by all cations the extracellular is to however, that at their of the cations blocked currents through TRPM3α2 these results that TRPM3 channel activity is tightly regulated by the of extracellular TRPM3α1 channels are only to divalent cations, TRPM3α2 channels are to all extracellular of TRPM3 intracellular Ca2+ in to TRPM3α1 or TRPM3α2 at In increased Ca2+ of with and we the to an extracellular of Ca2+ intracellular Ca2+ to of the expression of TRPM3 channels of extracellular an rise of the intracellular Ca2+ an was in in These data that an increased for Ca2+ extracellular ionic that the intracellular Ca2+ On the other to have a for Ca2+ with as by the Ca2+ after extracellular Ca2+ and the intracellular Ca2+ These with our data that TRPM3α2 channels have a divalent with TRPM3α1 channels and They also well with that the human TRPM3a and TRPM31325 variants, which the pore region as TRPM3α2, Ca2+ entry (12Lee N. Chen J. Wu S. Sun L. Huang M. Levesque P.C. Rich A. Feder J.N. Gray K.R. Lin J.H. Janovitz E.B. Blanar M.A. J. Biol. Chem. 2003; 278: 20890-20897Abstract Full Text Full Text PDF PubMed Scopus (164) Google Scholar, 13Grimm C.M. Kraft R. Sauerbruch S. Schultz G. Harteneck C. J. Biol. Chem. 2003; 278: 21493-21501Abstract Full Text Full Text PDF PubMed Scopus (277) Google Scholar). In this report, we have described five novel splice variants expressed in mouse and that from the of within the are of the and sequence of the and have been described N. Chen J. Wu S. Sun L. Huang M. Levesque P.C. Rich A. Feder J.N. Gray K.R. Lin J.H. Janovitz E.B. Blanar M.A. J. Biol. Chem. 2003; 278: 20890-20897Abstract Full Text Full Text PDF PubMed Scopus (164) Google and 13Grimm C.M. Kraft R. Sauerbruch S. Schultz G. Harteneck C. J. Biol. Chem. 2003; 278: 21493-21501Abstract Full Text Full Text PDF PubMed Scopus (277) Google and this The gene for a of different of which have up to now been we have splice variants that differ only in a predicted to to the ion-conducting the of TRPM3 selectivity of ion channels is to be by the and of the selectivity as the of the channel pore B. of Scholar). all members of an ion channel such as channels, common ionic The TRP family of ion channels is in this as members with cationic selectivity in D.E. Nature. 2003; 426: 517-524Crossref PubMed Scopus (2164) Google Scholar). The gene to this because two channels can be expressed from this gene with different ionic One monovalent cation whereas TRPM3α2 divalent In such a in ionic selectivity be to have consequences for the of the channel and the of the cell that has been that of amino acid residues can the divalent selectivity of ion channels. channels that are to Ca2+ by a an to a are an H. K. S. Nature. PubMed Scopus Google Scholar). However, these in the amino acid sequence have been whereas the in selectivity we describe for TRPM3 have and in Although alternative splicing is as a mechanism to ion channel 2001; PubMed Scopus Google Scholar), has not been in the selectivity of a Interestingly, the of also splice variants that differ in the pore-forming region (8Lis A. Wissenbach U. Philipp S.E. Naunyn-Schmiedeberg′s Arch. Pharmacol. 2005; (in press)PubMed Google Scholar) and might a further for such a of is mechanism of in of a selected in transcripts of receptors to the of a by an within the channel has been to the Ca2+ selectivity of channels with consequences for their H. K. S. Nature. PubMed Scopus Google Scholar). the in of ionic selectivity in TRPM3 variants is by a of amino acid residues and further within the between the presumed fifth and transmembrane The differences in ion selectivity for the TRPM3 splice variants that this the pore of Although this be to be the ion-conducting due to for and TRPV6 channels T. Nilius B. 2003; PubMed Scopus Google Scholar), this has not been up to now for any member of the with the presumed pore of other members of the TRP the pore loop of TRPM3 is considerably by and additional amino acid residues The domains that the selectivity of the channels T. A. Droogmans G. Nilius B. J. Biol. Chem. 2004; Full Text Full Text PDF PubMed Scopus Google Scholar) are in TRPM3 The splicing within the TRPM3 channel pore amino acid residues into this might the Ca2+ of TRPM3α1 with TRPM3α2, because of increased In with this in and receptors the of a by a by the Ca2+ selectivity of channels as well P.H. J. Curr. 2003; 13: PubMed Scopus Google Scholar). residues in the pore of channels their divalent selectivity H. K. S. Nature. PubMed Scopus Google Scholar). of TRPM3 by and data show that TRPM3α1 and TRPM3α2 are regulated by of intracellular to members of the family such as TRPM6 and T. Nilius B. S. Droogmans G. Bindels R.J. Hoenderop J.G. J. Biol. Chem. 2004; Full Text Full Text PDF PubMed Scopus Google Scholar, K. Perraud A.L. T. Kinet J.P. Penner R. Scharenberg A.M. Fleig A. Nature. 2001; PubMed Scopus Google Scholar). the human variant TRPM31325 has been reported to increased entry when the of the extracellular was from to (13Grimm C.M. Kraft R. Sauerbruch S. Schultz G. Harteneck C. J. Biol. Chem. 2003; 278: 21493-21501Abstract Full Text Full Text PDF PubMed Scopus (277) Google Scholar), a that swelling of the cells. the that we in cell a of the to in the intracellular cation and the cell are and of TRPM3 activity by intracellular might TRPM3 increased or activity in and were in these we found that TRPM3 currents are also tightly regulated by extracellular However, only TRPM3α2 channel activity was by the extracellular of monovalent cations, whereas TRPM3 variants were by extracellular divalent by extracellular is a highly of ion channels, not E. A. Arch. J. PubMed Scopus Google Scholar) H. J.P. Nat. 2000; PubMed Scopus Google Scholar) channels have also been to be by extracellular Although the of TRPM3α2 by extracellular in the range to be in not appear to be The intracellular Ca2+ in was also extracellular Ca2+ can the cell through TRPM3α2 channels at a to be in in Ca2+ Functional of found that TRPM3 is expressed in the in the of the is for the and the of has been that the of such as Ca2+ in are regulated and are of in the of these T. 2004; PubMed Scopus Google Scholar). The expression in the might of TRPM3 proteins in the of or the of its ionic we not which of the TRPM3 variants are in the However, the selectivity for divalent cations of TRPM3α2 channels this variant a to a in the of divalent cation in We and for and and Cavalié for and of the