Biomedical subjects

G Droogmans

Publications and source records attributed to G Droogmans.

At least 19 recordsLinked to original sources

PubMed2004

Cell volume response to hyposmotic shock and elevated cAMP in bovine trabecular meshwork cells.

PURPOSE: Hyposmolar perfusion of intact trabecular meshwork (TM) induces a decrease in its hydraulic conductivity (Lp). However, exposure to agents that elevate intracellular cAMP in TM cells increases Lp. Since volume of TM cells could directly influence porosity of the TM and hence Lp, this study has investigated changes in volume in response to acute hyposmotic shock (i.e. regulatory volume decrease or RVD) and elevated cAMP in cultured TM cells. METHODS: Bovine trabecular meshwork cells (BTMC), grown on glass coverslips and loaded with the fluorescent dye MQAE, were used to measure rapid changes in cell volume using the principle of dynamic fluorescence quenching. Activation of volume-regulated anion channels (VRAC) was assessed by measuring volume-sensitive Cl(-) currents (I(Cl,swell)) in the whole cell configuration of the patch clamp technique and by determining the swelling-induced enhancement in I(-) permeability using the halide-sensitivity of MQAE. Expressions of ClC (chloride channels of the ClC gene family), P-glycoprotein (Pgp), and cystic fibrosis transmembrane regulator (CFTR) Cl(-) channels were examined by RT-PCR. Elevation of cAMP in response to forskolin was confirmed by determining the phosphorylation of cAMP response element-binding protein and activating transcription factor-1 (CREB, ATF-1), which form the downstream targets of protein kinase A. RESULTS: As a response to hyposmotic shock, there was an acute increase in cell volume but there was no robust RVD. Patch clamp experiments showed activation of a characteristic Cl(-) current in response to cell swelling. This Cl(-) current was inhibited by NPPB (100microM) and fluoxetine (50microM), both of which are known blockers of VRAC. Experiments, which used the halide-sensitivity of MQAE, also indicated a 9-fold increase in I(-) influx upon cell swelling (8.9+/-4.6; n=9), consistent with activation of a VRAC-like Cl(-) current. To examine whether RVD is limited by K(+) conductance, the swollen cells were exposed to gramicidin, which is known to induce cation channel activity. Such a maneuver led to secondary swelling with [Na(+)](o)=140mM but a rapid shrinkage [Na(+)](o)=8mM indicating that the RVD is limited by cationic conductance necessary for K(+) efflux. Exposure to forskolin, which resulted in CREB and ATF-1 phosphorylation, caused a reversible decrease in cell volume (14.5+/-5%; n=20) under isosmotic and hyposmotic conditions. RT-PCR analysis confirmed expression of ClC-2, ClC-5, and Pgp Cl(-) channels in bovine TM cells. However, ClC-3 and CFTR were not expressed. CONCLUSIONS: TM cells respond to acute hyposmotic shock in an osmometric manner, but their RVD is limited by K(+) conductance. The lack of CFTR expression and decrease in cell volume in response to forskolin concomitant with hyposmolarity suggest that elevated cAMP activates a K(+) conductance. Thus, the altered resistance to aqueous outflow in response to hyposmotic perfusion of the TM and elevated cAMP may be attributed to persistent cell swelling and cell shrinkage, respectively.

Animals↗

PubMed2003

Cell swelling, heat, and chemical agonists use distinct pathways for the activation of the cation channel TRPV4.

TRPV4 is a Ca(2+)- and Mg(2+)-permeable cation channel within the vanilloid receptor subgroup of the transient receptor potential (TRP) family, and it has been implicated in Ca(2+)-dependent signal transduction in several tissues, including brain and vascular endothelium. TRPV4-activating stimuli include osmotic cell swelling, heat, phorbol ester compounds, and 5',6'-epoxyeicosatrienoic acid, a cytochrome p450 epoxygenase metabolite of arachidonic acid (AA). It is presently unknown how these distinct activators converge on opening of the channel. Here, we demonstrate that blockers of phospholipase A(2) (PLA(2)) and cytochrome p450 epoxygenase inhibit activation of TRPV4 by osmotic cell swelling but not by heat and 4alpha-phorbol 12,13-didecanoate. Mutating a tyrosine residue (Tyr-555) in the N-terminal part of the third transmembrane domain to an alanine strongly impairs activation of TRPV4 by 4alpha-phorbol 12,13-didecanoate and heat but has no effect on activation by cell swelling or AA. We conclude that TRPV4-activating stimuli promote channel opening by means of distinct pathways. Cell swelling activates TRPV4 by means of the PLA(2)-dependent formation of AA, and its subsequent metabolization to 5',6'-epoxyeicosatrienoic acid by means of a cytochrome p450 epoxygenase-dependent pathway. Phorbol esters and heat operate by means of a distinct, PLA(2)- and cytochrome p450 epoxygenase-independent pathway, which critically depends on an aromatic residue at the N terminus of the third transmembrane domain.

Animals↗

PubMed2003

Amazing chloride channels: an overview.

AIM: This review describes molecular and functional properties of the following Cl- channels: the ClC family of voltage-dependent Cl- channels, the cAMP-activated transmembrane conductance regulator (CFTR), Ca2+ activated Cl- channels (CaCC) and volume-regulated anion channels (VRAC). If structural data are available, their relationship with the function of Cl- channels will be discussed. We also describe shortly some recently discovered channels, including high conductance Cl- channels and the family of bestrophins. We illustrate the growing physiological importance of these channels in the plasma membrane and in intracellular membranes, including their involvement in transepithelial transport, pH regulation of intracellular organelles, regulation of excitability and volume regulation. Finally, we discuss the role of Cl- channels in various diseases and describe the pathological phenotypes observed in knockout mice models.

Animals↗

PubMed2002

Block of volume-regulated anion channels by selective serotonin reuptake inhibitors.

We have used the whole-cell patch clamp technique to study the effects of the commonly used antidepressants sertraline, paroxetine, citalopram and fluvoxamine on the volume-regulated anion channel (VRAC) in endothelial cells. It was the purpose of the present experiments to investigate whether VRAC block is a general property of this group of selective serotonin reuptake inhibitors (SSRIs). At pH 7.4, all SSRIs induced a fast and reversible block of the volume-sensitive chloride current ( I(Cl,swell)), with an IC(50) value of 2.1+/-0.5 microM for sertraline, 2.7+/-0.2 microM for paroxetine, 12.3+/-1.4 microM for fluvoxamine and 27.7+/-2.8 microM for citalopram. The block was enhanced at more alkaline pH, indicating that it is mediated by the uncharged form. This study describes the effects of a variety of SSRIs on an anion channel. Our data reveal a potent block and suggest a hydrophobic interaction of high affinity between the uncharged SSRI and volume-regulated anion channels. We conclude that VRAC block is a general property of this pharmacological class of selective serotonin reuptake inhibitors.

Antidepressive Agents↗

PubMed2002

Current understanding of mammalian TRP homologues.

Calcium influx into the cell from the extracellular medium is crucial for important processes including muscle contraction, secretion and gene expression. This calcium influx is mainly mediated through calcium influx channels, which on the basis of their activation mechanism can be subdivided in voltage-gated calcium channels, which have already been thoroughly characterized and non-voltage-gated calcium permeable channels. This latter group includes ion channels activated by binding of extra and intracellular messengers, mechanical stress or depletion of intracellular calcium stores. Currently little molecular data is available concerning this class of calcium influx channels. However, recent studies have indicated that members of the transient receptor potential (TRP) family of ion channels can function as calcium influx channels both in excitable and non-excitable tissues. On the basis of structural information the TRP family is subdivided in three main subfamilies: the TRPC (canonical) group, the TRPV (vanilloid) group and the TRPM (melastatin) group. The cloning and characterization of members of this cation channel family has exploded during recent years, leading to a plethora of data concerning TRPs in a variety of tissues and species, including mammals, insects and yeast. This review summarizes the currently available information concerning members of the TRP family expressed in mammalian tissues.

Animals↗

PubMed2001

Function and expression of the epithelial Ca(2+) channel family: comparison of mammalian ECaC1 and 2.

1. The epithelial Ca(2+) channel (ECaC) family represents a unique group of Ca(2+)-selective channels that share limited homology to the ligand-gated capsaicin receptors, the osmolarity-sensitive channel OTRPC4, as well as the transient receptor potential family. Southern blot analysis demonstrated that this family is restricted to two members, ECaC1 and ECaC2 (also named CaT1). 2. RT-PCR analysis demonstrated that the two channels are co-expressed in calbindin-D-containing epithelia, including small intestine, pancreas and placenta, whereas kidney and brain only express ECaC1 and stomach solely ECaC2. 3. From an electrophysiological point of view, ECaC1 and ECaC2 are highly similar channels. Differences concern divalent cation permeability, the kinetics of Ca(2+)-dependent inactivation and recovery from inactivation. 4. Ruthenium red is a potent blocker of ECaC activity. Interestingly, ECaC2 has a 100-fold lower affinity for ruthenium red (IC(50) 9 +/- 1 microM) than ECaC1 (IC(50) 121 +/- 13 nM). 5. ECaCs are modulated by intracellular Mg(2+) and ATP. ECaC1 and ECaC2 activity rapidly decay in the absence of intracellular ATP. This effect is further accelerated at higher intracellular Mg(2+) concentrations. 6. In conclusion, ECaC1 and ECaC2 are homologous channels, with an almost identical pore region. They can be discriminated by their sensitivity for ruthenium red and show differences in Ca(2+)-dependent regulation.

Adenosine Triphosphate↗

PubMed2001

CaT1 and the calcium release-activated calcium channel manifest distinct pore properties.

The calcium release-activated calcium channel (CRAC) is a highly Ca(2+)-selective ion channel that is activated on depletion of inositol triphosphate (IP(3))-sensitive intracellular Ca(2+) stores. It was recently reported that CaT1, a member of the TRP family of cation channels, exhibits the unique biophysical properties of CRAC, which led to the conclusion that CaT1 comprises all or part of the CRAC pore (Yue, L., Peng, J. B., Hediger, M. A., and Clapham, D. E. (2001) Nature 410, 705-709). Here, we directly compare endogenous CRAC with heterologously expressed CaT1 and show that they manifest several clearly distinct properties. CaT1 can be distinguished from CRAC in the following features: sensitivity to store-depleting agents; inward rectification in the absence of divalent cations; relative permeability to Na(+) and Cs(+); effect of 2-aminoethoxydiphenyl borate (2-APB). Moreover, CaT1 displays a mode of voltage-dependent gating that is fully absent in CRAC and originates from the voltage-dependent binding/unbinding of Mg(2+) inside the channel pore. Our results imply that the pores of CaT1 and CRAC are not identical and indicate that CaT1 is a Mg(2+)-gated channel not directly related to CRAC.

Calcium Channels↗

PubMed2001

Aspartate residues of the Glu-Glu-Asp-Asp (EEDD) pore locus control selectivity and permeation of the T-type Ca(2+) channel alpha(1G).

The structural determinant of the permeation and selectivity properties of high voltage-activated (HVA) Ca(2+) channels is a locus formed by four glutamate residues (EEEE), one in each P-region of the domains I-IV of the alpha(1) subunit. We tested whether the divergent aspartate residues of the EEDD locus of low voltage-activated (LVA or T-type) Ca(2+) channels account for the distinctive permeation and selectivity features of these channels. Using the whole-cell patch-clamp technique in the HEK293 expression system, we studied the properties of the alpha(1G) T-type, the alpha(1C) L-type Ca(2+) channel subunits, and alpha(1G) pore mutants, containing aspartate-to-glutamate conversions in domain III, domain IV, or both. Three characteristic features of HVA Ca(2+) channel permeation, i.e. (a) Ba(2+) over Ca(2+) permeability, (b) Ca(2+)/Ba(2+) anomalous mole fraction effect (AMFE), and (c) high Cd(2+) sensitivity, were conferred on the domain III mutant (EEED) of alpha(1G). In contrast, the relative Ca(2+)/Ba(2+) permeability and the lack of AMFE of the alpha(1G) wild type channel were retained in the domain IV mutant (EEDE). The double mutant (EEEE) displayed AMFE and a Cd(2+) sensitivity similar to that of alpha(1C), but currents were larger in Ca(2+)- than in Ba(2+)-containing solutions. The mutation in domain III, but not that in domain IV, consistently displayed outward fluxes of monovalent cations. H(+) blocked Ca(2+) currents in all mutants more efficiently than in alpha(1G). In addition, activation curves of all mutants were displaced to more positive voltages and had a larger slope factor than in alpha(1G) wild type. We conclude that the aspartate residues of the EEDD locus of the alpha(1G) Ca(2+) channel subunit not only control its permeation properties, but also affect its activation curve. The mutation of both divergent aspartates only partially confers HVA channel permeation properties to the alpha(1G) Ca(2+) channel subunit.

Aspartic Acid↗

PubMed2001

Inhibition of volume-regulated anion channels by dominant-negative caveolin-1.

Caveolae are flask-shaped invaginations of the plasma membrane formed by the association of caveolin proteins with lipid rafts. In endothelial cells, caveolae function as signal transduction centers controlling NO synthesis and mechanotransduction. We now provide evidence that the endothelial volume-regulated anion channel (VRAC) is also under the control of the caveolar system. When calf pulmonary artery endothelial (CPAE) cells were transfected with caveolin-1 Delta1-81 (deletion of amino acids 1 to 81), activation of VRAC by hypotonic cell swelling was strongly impaired. Concomitantly, caveolin-1 Delta1-81 disturbed the formation of caveolin-1 containing lipid rafts as evidenced by sucrose density gradient centrifugation. In nontransfected cells, endogenous caveolin-1 typically associated with low-density, detergent-resistant lipid rafts. However, transient expression of caveolin-1 Delta1-81 caused a redistribution of endogenous caveolin-1 to high-density, detergent-soluble membrane fractions. We therefore conclude that the interaction between caveolin-1 and detergent-resistant lipid rafts is an important prerequisite for endothelial VRAC activity.

Animals↗

PubMed2001

Functional interaction between TRP4 and CFTR in mouse aorta endothelial cells.

BACKGROUND: This study describes the functional interaction between the putative Ca2+ channel TRP4 and the cystic fibrosis transmembrane conductance regulator, CFTR, in mouse aorta endothelium (MAEC). RESULTS: MAEC cells express CFTR transcripts as shown by RT-PCR analysis. Application of a phosphorylating cocktail activated a Cl- current with characteristics similar to those of CFTR mediated currents in other cells types (slow activation by cAMP, absence of rectification, block by glibenclamide). The current is present in trp4 +/+ MAEC, but not in trp4 -/- cells, although the expression of CFTR seems unchanged in the trp4 deficient cells as judged from RT-PCR analysis. CONCLUSIONS: It is concluded that TRP4 is necessary for CFTR activation in endothelium, possibly by providing a scaffold for the formation of functional CFTR channels.

Animals↗

PubMed2001

Pore properties and ionic block of the rabbit epithelial calcium channel expressed in HEK 293 cells.

We have used the whole-cell patch-clamp technique to analyse the permeation properties and ionic block of the epithelial Ca2+ channel ECaC heterologously expressed in human embryonic kidney (HEK) 293 cells. Cells dialysed with 10 mM BAPTA and exposed to Ca2+-containing, monovalent cation-free solutions displayed large inwardly rectifying currents. Their reversal potential depended on the extracellular Ca2+ concentration, [Ca2+]o. The slope of the relationship between reversal potential and [Ca2+]o on a logarithmic scale was 21 +/- 4 mV, compared with 29 mV as predicted by the Nernst equation (n = 3-5 cells). Currents in mixtures of Ca2+ and Na+ or Ca2+ and Ba2+ showed anomalous mole fraction behaviour. We have described the current-concentration plot for Ca2+ and Na+ by a kinetic permeation model, i.e. the "step" model. Extracellular Mg2+ blocked both divalent and monovalent currents with an IC50 of 62 +/- 9 microM(n = 4) in Ca2+-free conditions and 328 +/- 50 microM (n = 4-9) in 100 microM Ca2+ solutions. Mono- and divalent currents through ECaCs were blocked by gadolinium, lanthanum and cadmium, with a blocking order of Cd2+ >> Gd3+ > La3+. We conclude that the permeation of monovalent and divalent cations through ECaCs shows similarities with L-type voltage-gated Ca2+ channels, the main differences being a higher Ca2+ affinity and a significantly higher current density in micromolar Ca2+ concentrations in the case of ECaCs.

Animals↗

PubMed2001

The single pore residue Asp542 determines Ca2+ permeation and Mg2+ block of the epithelial Ca2+ channel.

The epithelial Ca(2+) channel (ECaC), which was recently cloned from rabbit kidney, exhibits distinctive properties that support a facilitating role in transcellular Ca(2+) (re)absorption. ECaC is structurally related to the family of six transmembrane-spanning ion channels with a pore-forming region between S5 and S6. Using point mutants of the conserved negatively charged amino acids present in the putative pore, we have identified a single aspartate residue that determines Ca(2+) permeation of ECaC and modulation by extracellular Mg(2+). Mutation of the aspartate residue, D542A, abolishes Ca(2+) permeation and Ca(2+)-dependent current decay as well as block by extracellular Mg(2+), whereas monovalent cations still permeate the mutant channel. Variation of the side chain length in mutations D542N, D542E, and D542M attenuated Ca(2+) permeability and Ca(2+)-dependent current decay. Block of monovalent currents through ECaC by Mg(2+) was decreased. Exchanging the aspartate residue for a positively charged amino acid, D542K, resulted in a nonfunctional channel. Mutations of two neighboring negatively charged residues, i.e. Glu(535) and Asp(550), had only minor effects on Ca(2+) permeation properties.

Amino Acid Sequence↗

PubMed2001

The amino side of the C-terminus determines fast inactivation of the T-type calcium channel alpha1G.

We analysed the kinetic properties of the fast inactivating T-type calcium channel alpha1G in HEK 293 cells transfected with different alpha1G chimeras, containing the N-terminus, III-IV linker or various C-terminal regions of the slowly inactivating L-type alpha1C. A highly negatively charged region of 23 amino acids at the amino side of the intracellular carboxy terminus of alpha1G was found to be critical for fast inactivation. The N-terminus of alpha1G does not seem to be necessary for inactivation of the T-type calcium channel because replacement of the a1G N-terminus with the alpha1C N-terminus did not influence channel kinetics at all. Replacing the III-IV linker of alpha1G with that of a1C decreased the rate of inactivation at -20 mV from 15.8 +/- 1.8 to 8.5 +/- 1.1 ms, and shifted the potential for half-maximal inactivation from -69.6 +/- 0.8 to -54.0 +/- 1.7 mV. However, these parameters were not significantly different at other potentials. We suggest a putative 'ball-and-chain'-like mechanism for inactivation in which the negative charges function as an acceptor domain for a ball, hypothetically located at a different intracellular part of the channel. In addition, transferring the IQ motif and EF hand of alpha1C to alpha1G does not confer Ca2+-dependent inactivation on alpha1G, suggesting that other sequences besides the C-terminus are needed for Ca2+-dependent inactivation of alpha1C.

Algorithms↗

PubMed2001

Modulation of the epithelial Ca2+ channel ECaC by extracellular pH.

We investigated the effect of extracellular pH on whole-cell currents through the epithelial Ca2+ channel, ECaC, expressed in HEK 293 cells. Both mono- and divalent current densities were significantly smaller at pH 6.0 than at pH 7.4. At pH 8.5 they were slightly larger. Lowering extracellular pH enhanced the slow component of monovalent current activation at negative potentials but had no significant effect on the kinetics of Ca2+ currents. The kinetics of block of monovalent cation current by extracellular Mg2+ was significantly changed at high and low pH. The time constant of the time- and voltage-dependent current component during a voltage step to -140 mV was significantly larger at pH 8.5 than at pH 7.4. At pH 6.0 it was almost absent. The [Mg2+] inhibiting 50% of monovalent current through ECaC at pH 6.0 (IC50) was 323 +/- 23 microM (n = 8), compared with 62 +/- 9 microM (n = 4) at pH 7.4 and 38 +/- 4 microM (n = 8) at pH 8.5. The affinity of ECaC for Ca2+ was also affected by extracellular pH, shifting from 4.8 +/- 0.7 microM (n = 6) at pH 6.0 to 161 +/- 30 nM (n = 5) at pH 7.4 and 425 +/- 117 nM (n = 8) at pH 8.5.

Calcium↗

PubMed2001

Ca(2+)-activated Cl- channels in Ehrlich ascites tumor cells are distinct from mCLCA1, 2 and 3.

Using the whole-cell patch-clamp technique, we have studied the electrophysiological and pharmacological properties of the Ca(2+)-activated Cl- current present in Ehrlich cells. The currents activated slowly upon depolarization, deactivated upon hyperpolarization, and showed strong outward rectification. An increase in [Ca2+]i activated the current with an EC50 of 165.2 nM. Extracellular application of niflumic acid (100 microM) rapidly blocked the current in a voltage-dependent manner whereas sulfhydryl-modifying agents such as dithiothreitol (DTT, 1-2 mM) and N-ethylmaleimide (NEM, 100 microM) had no effect on Ca(2+)-activated currents in Ehrlich cells. Members of the recently discovered CLCA gene family are the only molecular candidates for Ca(2+)-activated Cl- channels cloned so far. Using RT-PCR we demonstrated that the appearance of a Ca(2+)-activated Cl- current in Ehrlich cells is not associated with the expression of the murine members of the CLCA family (mCLCA1-mCLCA3). Correspondingly, the kinetic and pharmacological properties of the Ca(2+)-activated current in Ehrlich cells differ from those of CLCA-associated currents, which are time independent and DTT sensitive. Thus, phenotypic differences in combination with RT-PCR data point to the existence of different molecular species for Ca(2+)-activated Cl- channels.

Animals↗

PubMed2001

The C-terminal part of the R-domain, but not the PDZ binding motif, of CFTR is involved in interaction with Ca(2+)-activated Cl- channels.

Expression of the cystic fibrosis transmembrane conductance regulator (CFTR) inhibits Ca(2+)-activated Cl- channels (CaCC) by an unknown mechanism. This inhibition does not require CFTR activation (activity-independent inhibition), but is potentiated when CFTR is activated (activity-dependent inhibition). In this study, we evaluated, in endothelial cells, possible structural determinants for this interaction. Bovine pulmonary artery endothelium (CPAE) cells, which do not express CFTR, were transfected transiently with three hybrid CFTR constructs. The functional interaction between CaCC and CFTR was assessed using the patch-clamp technique in the whole-cell configuration. CaCC was stimulated by application of adenosine 5'-triphosphate (ATP) to the bath solution. CFTR currents were evoked by application of a forskolin/3-isobutyl-l-methylxanthine (IBMX) cocktail. The inhibitory effect of CFTR was conserved when the PDZ (PSD-95/Discs large/ZO-1) binding motif was deleted (CFTR-delta PDZ). In contrast, both the CFTR activity-independent and -dependent inhibition of CaCC were abolished when the C-terminal part of the regulatory (R)-domain of CFTR was deleted (CFTR-delta R780-830). The activity-dependent inhibition of CaCC, but not the activity-independent inhibition, could be rescued by introducing the multiple drug resistance (MDR)-1 mini-linker in place of the deletion (CFTR-delta R-linker). It is concluded that the C-terminal part of the R-domain is an important determinant for CFTR-CaCC interaction.

Amino Acid Motifs↗

PubMed2001

Differential activation of the volume-sensitive cation channel TRP12 (OTRPC4) and volume-regulated anion currents in HEK-293 cells.

The detection of changes in volume and osmolality is an essential function in vertebrate cells. A novel member of the transient receptor potential (trp) family of ion channels, which is sensitive to changes in cell volume, has been described recently. Heterologous expression of TRP12 in HEK cells resulted in the appearance of a swelling-activated cation current. The permeability sequence of this cation current for various monovalent cations, as determined from shifts in reversal potential upon extracellular cation substitution, was PK>PCs>PNa>PLi, corresponding to an Eisenman-IV sequence characteristic for a weak-field-strength site. Surprisingly, over-expression of this channel in HEK cells was accompanied by a dramatic down-regulation of the volume-regulated anion channel (VRAC), which is activated by cell swelling in non-transfected cells. In contrast to VRAC, TRP12 could not be activated at constant volume by a reduction of intracellular ionic strength or by intracellular perfusion with guanosine 5'-O-(3-thiotriphosphate (GTPgammaS). The kinetic and pharmacological profile of VRAC and TRP12 currents were also different.

Anions↗

PubMed2001

Lack of an endothelial store-operated Ca2+ current impairs agonist-dependent vasorelaxation in TRP4-/- mice.

Agonist-induced Ca2+ entry into cells by both store-operated channels and channels activated independently of Ca2+-store depletion has been described in various cell types. The molecular structures of these channels are unknown as is, in most cases, their impact on various cellular functions. Here we describe a store-operated Ca2+ current in vascular endothelium and show that endothelial cells of mice deficient in TRP4 (also known as CCE1) lack this current. As a consequence, agonist-induced Ca2+ entry and vasorelaxation is reduced markedly, showing that TRP4 is an indispensable component of store-operated channels in native endothelial cells and that these channels directly provide an Ca2+-entry pathway essentially contributing to the regulation of blood vessel tone.

Acetylcholine↗