title: Optopharmacological control of TRPC channels by coumarin-caged lipids is associated with a phototoxic membrane effect.
abstract: Photouncaging of second messengers has been successfully employed to gain mechanistic insight of cellular signaling pathways. One of the most enigmatic processes of ion channel regulation is lipid recognition and lipid-gating of TRPC channels, which represents pivotal mechanisms of cellular Ca(2+) homeostasis. Recently, optopharmacological tools including caged lipid mediators became available, enabling an unprecedented level of temporal and spatial control of the activating lipid species within a cellular environment. Here we tested a commonly used caged ligand approach for suitability to investigate TRPC signaling at the level of membrane conductance and cellular Ca(2+) handling. We report a specific photouncaging artifact that is triggered by the cage structure coumarin at UV illumination. Electrophysiological characterization identified a light-dependent membrane effect of coumarin. UV light (340 nm) as used for photouncaging, initiated a membrane conductance specifically in the presence of coumarin as low as 30 μmol L(-1) concentrations. This conductance masked the TRPC3 conductance evoked by photouncaging, while TRPC-mediated cellular Ca(2+) responses were largely preserved. The observed light-induced membrane effects of the released caging moiety may well interfere with certain cellular functions, and prompt caution in using coumarin-caged second messengers in cellular studies.
authors.affiliation: Institute of Biophysics, Medical University of Graz, BioTechMed-Graz, Harrachgasse 21, Graz, 8010, Austria.
authors.forename: Oleksandra
authors.initials: O
authors.lastname: Tiapko
authors.affiliation: Institute of Biophysics, Medical University of Graz, BioTechMed-Graz, Harrachgasse 21, Graz, 8010, Austria. bernadett.bacsa@medunigraz.at.
authors.forename: Bernadett
authors.initials: B
authors.lastname: Bacsa
authors.affiliation: Institute of Chemistry, University of Graz, NAWI Graz, BioTechMed-Graz, Heinrichstrasse 28, Graz, 8010, Austria.
authors.forename: Gema Guedes
authors.initials: GG
authors.lastname: de la Cruz
authors.affiliation: Institute of Chemistry, University of Graz, NAWI Graz, BioTechMed-Graz, Heinrichstrasse 28, Graz, 8010, Austria.
authors.forename: Toma
authors.initials: T
authors.lastname: Glasnov
authors.affiliation: Institute of Biophysics, Medical University of Graz, BioTechMed-Graz, Harrachgasse 21, Graz, 8010, Austria. klaus.groschner@medunigraz.at.
authors.forename: Klaus
authors.initials: K
authors.lastname: Groschner
mesh_terms: D002118:Calcium / Q000378:metabolism*; D002121:Calcium Channel Blockers / Q000494:pharmacology; D020013:Calcium Signaling / Q000187:drug effects / Q000528:radiation effects; D003374:Coumarins / Q000737:chemistry*; D004075:Diglycerides / Q000737:chemistry / Q000494:pharmacology; D057809:HEK293 Cells; D006801:Humans; D007093:Imidazoles / Q000494:pharmacology; D008055:Lipids / Q000737:chemistry / Q000494:pharmacology*; D008164:Luminescent Proteins / Q000235:genetics / Q000378:metabolism; D008564:Membrane Potentials / Q000187:drug effects / Q000528:radiation effects; D008856:Microscopy, Fluorescence; D018408:Patch-Clamp Techniques; D010782:Photolysis / Q000528:radiation effects; D015398:Signal Transduction / Q000187:drug effects* / Q000528:radiation effects; D050052:TRPC Cation Channels / Q000235:genetics / Q000378:metabolism*; D014466:Ultraviolet Rays
chemical_list: D002121:Calcium Channel Blockers; D003374:Coumarins; D004075:Diglycerides; D007093:Imidazoles; D008055:Lipids; D008164:Luminescent Proteins; D050052:TRPC Cation Channels; C104603:TRPC3 cation channel; C063159:1-(2-(3-(4-methoxyphenyl)propoxy)-4-methoxyphenylethyl)-1H-imidazole; D002118:Calcium
keywords: Ca2+ signaling; TRPC channels; caged diacylglycerols; optopharmacology