TRPV1 regulators mediate gentamicin penetration of cultured kidney cells.
Transient receptor potential (TRP) receptors are, typically, calcium-permeant cation channels that transduce environmental stimuli. Both kidney epithelial and inner ear sensory cells express TRPV1, are mechanosensors and accumulate the aminoglycoside antibiotic gentamicin. Recently, we showed that Texas Red-conjugated gentamicin (GTTR) enters kidney cells via an endosome-independent pathway. Here, we used GTTR to investigate this non-endocytotic mechanism of gentamicin uptake. In serum-free buffers, GTTR penetrated MDCK cells within 30 s and uptake was modulated by extracellular, multivalent cations (Ca2+, La3+, Gd3+) or protons. We verified the La3+ modulation of GTTR uptake using immunocytochemical detection of unconjugated gentamicin. Membrane depolarization, induced by high extracellular K+ or valinomycin, also reduced GTTR uptake, suggesting electrophoretic permeation through ion channels. GTTR uptake was enhanced by the TRPV1 agonists, resiniferatoxin and anandamide, in Ca2+-free media. Competitive antagonists of the TRPV1 cation current, iodo-resiniferatoxin and SB366791, also enhanced GTTR uptake independently of Ca2+, reinforcing these antagonists' potential as latent agonists in specific situations. Ruthenium Red blocked GTTR uptake in the presence or absence of these TRPV1-agonists and antagonists. In addition, GTTR uptake was blocked by RTX in the presence of more physiological levels (2 mM) of Ca2+. Thus gentamicin enters cells via cation channels, and gentamicin uptake can be modulated by regulators of the TRPV1 channel.
Myrdal
Sigrid E
SE
Oregon Hearing Research Center, Oregon Health & Science University, 3181 SW Sam Jackson Park Road, Portland, OR 97239, USA.
Steyger
Peter S
PS
D000813:Anilides / Q000494:pharmacology; D000818:Animals; D000900:Anti-Bacterial Agents / Q000009:adverse effects / Q000378:metabolism*; D001095:Arachidonic Acids / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism; D002121:Calcium Channel Blockers / Q000494:pharmacology; D002460:Cell Line; D002934:Cinnamates / Q000494:pharmacology; D004224:Diterpenes / Q000494:pharmacology; D004285:Dogs; D063388:Endocannabinoids; D005456:Fluorescent Dyes; D005682:Gadolinium / Q000494:pharmacology; D005839:Gentamicins / Q000009:adverse effects / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D007202:Indicators and Reagents; D007476:Ionophores / Q000494:pharmacology; D007686:Kidney Tubules, Distal / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D007811:Lanthanum / Q000494:pharmacology; D008564:Membrane Potentials / Q000187:drug effects; D053284:Polyunsaturated Alkamides; D012430:Ruthenium Red; D050916:TRPV Cation Channels / Q000819:agonists / Q000037:antagonists & inhibitors / Q000502:physiology*; D014634:Valinomycin / Q000494:pharmacology; D014966:Xanthenes
D000813:Anilides; D000900:Anti-Bacterial Agents; D001095:Arachidonic Acids; D002121:Calcium Channel Blockers; D002934:Cinnamates; D004224:Diterpenes; D063388:Endocannabinoids; D005456:Fluorescent Dyes; D005839:Gentamicins; D007202:Indicators and Reagents; D007476:Ionophores; C477659:N-(3-methoxyphenyl)-4-chlorocinnamanilide; D053284:Polyunsaturated Alkamides; D050916:TRPV Cation Channels; C494484:TRPV1 receptor; D014966:Xanthenes; C468130:iodoresiniferatoxin; D012430:Ruthenium Red; D014634:Valinomycin; D007811:Lanthanum; C034657:Texas red; C024353:resiniferatoxin; D005682:Gadolinium; D002118:Calcium; C078814:anandamide