Single-channel currents through transient-receptor-potential-like (TRPL) channels.
Single-channel current recordings were used to examine the properties and modulation of Drosophila transient-receptor-potential-like (TRPL) channels transiently expressed in HEK and COS cells. Recombinant TRPL channels were constitutively active and characterized by a conductance of 104 pS in on-cell membrane patches with 115 mM Na+ and 2 mM Mg2+ in the pipette solution. In inside-out membrane patches exposed to 115 mM Na+ plus 2 mM Mg2+, 115 mM Na+ plus 10 mM Mg2+, 90 mM Ca2+ and 90 mM Ba2+ on both sides, the single-channel conductances were 72 pS, 36 pS, 48 pS and 46 pS, respectively. The single TRPL channel currents reversed close to 0 mV and displayed a linear voltage dependence between -120 mV and +120 mV. Removal of cations from the pipette and bath solutions abolished inward and outward currents, respectively. Similar currents were not observed in mock-transfected and native cells. The opening probability of TRPL channels increased by depolarizing the membrane and accounted for the outward rectification of whole-cell TRPL currents. In on-cell membrane patches, the TRPL channel activity was enhanced by cell dialysis of 300 microM guanosine 5'-O-(3-thiotriphosphate) (GTP[gamma-S]) and by a rise of intracellular Ca2+ (>2 microM). Constitutively active TRPL channels depolarized the host cells to -10 mV and the membrane potential was restored by cell dialysis with 10 mM BAPTA. The present results suggest that TRPL forms non-selective cationic channels modulated by intracellular Ca2+ in mammalian cells.
Hambrecht
J
J
Institut für Pharmakologie und Toxikologie, Universität des Saarlandes, Homburg/Saar, Germany.
Zimmer
S
S
Flockerzi
V
V
Cavalié
A
A
D000818:Animals; D019556:COS Cells; D002118:Calcium / Q000493:pharmacokinetics*; D002148:Calmodulin-Binding Proteins / Q000235:genetics* / Q000378:metabolism*; D002614:Chelating Agents / Q000494:pharmacology; D004330:Drosophila; D029721:Drosophila Proteins; D004533:Egtazic Acid / Q000031:analogs & derivatives* / Q000494:pharmacology; D015870:Gene Expression / Q000502:physiology; D017930:Genes, Reporter; D005971:Glutamates / Q000494:pharmacology; D049452:Green Fluorescent Proteins; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000494:pharmacology; D006801:Humans; D007202:Indicators and Reagents / Q000378:metabolism; D007668:Kidney / Q000166:cytology; D008164:Luminescent Proteins / Q000235:genetics; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology; D008565:Membrane Proteins / Q000235:genetics* / Q000378:metabolism*; D018408:Patch-Clamp Techniques; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D050051:Transient Receptor Potential Channels; D014785:Vision, Ocular / Q000502:physiology
D002148:Calmodulin-Binding Proteins; D002614:Chelating Agents; D029721:Drosophila Proteins; D005971:Glutamates; D007202:Indicators and Reagents; D008164:Luminescent Proteins; D008565:Membrane Proteins; D011994:Recombinant Proteins; D050051:Transient Receptor Potential Channels; C074192:trpl protein, Drosophila; D049452:Green Fluorescent Proteins; C014574:gamma-glutamylmethylamide; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D004533:Egtazic Acid; C025603:1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid; D002118:Calcium