title: Physiological pathway of magnesium influx in rat ventricular myocytes.
abstract: Cytoplasmic free Mg(2+) concentration ([Mg(2+)]i) was measured in rat ventricular myocytes with a fluorescent indicator furaptra (mag-fura-2) introduced by AM-loading. By incubation of the cells in a high-K(+) (Ca(2+)- and Mg(2+)-free) solution, [Mg(2+)]i decreased from ? 0.9 mM to 0.2 to 0.5 mM. The lowered [Mg(2+)]i was recovered by perfusion with Ca(2+)-free Tyrode's solution containing 1 mM Mg(2+). The time course of the [Mg(2+)]i recovery was fitted by a single exponential function, and the first derivative at time 0 was analyzed as being proportional to the initial Mg(2+) influx rate. The Mg(2+) influx rate was inversely related to [Mg(2+)]i, being higher at low [Mg(2+)]i. The Mg(2+) influx rate was augmented by the high extracellular Mg(2+) concentration (5 mM), whereas it was greatly reduced by cell membrane depolarization caused by high K(+). Known inhibitors of TRPM7 channels, 2-aminoethoxydiphenyl borate (2-APB), NS8593, and spermine reduced the Mg(2+) influx rate with half inhibitory concentrations (IC50) of, respectively, 17 ?M, 2.0 ?M, and 22 ?M. We also studied Ni(2+) influx by fluorescence quenching of intracellular furaptra by Ni(2+). The Ni(2+) influx was activated by lowering intra- and extracellular Mg(2+) concentrations, and it was inhibited by 2-APB and NS8593 with IC50 values comparable with those for the Mg(2+) influx. Intracellular alkalization (caused by pulse application of NH4Cl) enhanced, whereas intracellular acidification (induced after the removal of NH4Cl) slowed the Mg(2+) influx. Under the whole-cell patch-clamp configuration, the removal of intracellular and extracellular divalent cations induced large inward and outward currents, MIC (Mg-inhibited cation) currents or IMIC, carried by monovalent cations likely via TRPM7 channels. IMIC measured at -120 mV was diminished to ? 50% by 100 ?M 2-APB or 10 ?M NS8593. These results suggest that TRPM7/MIC channels serve as a major physiological pathway of Mg(2+) influx in rat ventricular myocytes.
authors.affiliation: Department of Physiology, Tokyo Medical University, Tokyo 160-8402, Japan. Electronic address: tashiro@tokyo-med.ac.jp.
authors.forename: Michiko
authors.initials: M
authors.lastname: Tashiro
authors.affiliation: Department of Physiology, Tokyo Medical University, Tokyo 160-8402, Japan.
authors.forename: Hana
authors.initials: H
authors.lastname: Inoue
authors.affiliation: Department of Physiology, Tokyo Medical University, Tokyo 160-8402, Japan.
authors.forename: Masato
authors.initials: M
authors.lastname: Konishi
mesh_terms: D015057:1-Naphthylamine / Q000031:analogs & derivatives / Q000494:pharmacology; D000818:Animals; D001896:Boron Compounds / Q000494:pharmacology; D002412:Cations / Q000378:metabolism; D005110:Extracellular Space / Q000378:metabolism; D016257:Fura-2 / Q000031:analogs & derivatives; D006863:Hydrogen-Ion Concentration; D042541:Intracellular Space / Q000378:metabolism; D008274:Magnesium / Q000378:metabolism*; D008297:Male; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology; D032383:Myocytes, Cardiac / Q000187:drug effects / Q000378:metabolism*; D009465:Neuromuscular Agents / Q000494:pharmacology; D009532:Nickel / Q000378:metabolism; D018408:Patch-Clamp Techniques; D011188:Potassium / Q000378:metabolism; D017208:Rats, Wistar; D013096:Spermine / Q000494:pharmacology; D050053:TRPM Cation Channels / Q000037:antagonists & inhibitors / Q000378:metabolism
chemical_list: C515166:(R)-N-(benzimidazol-2-yl)-1,2,3,4-tetrahydro-1-naphthylamine; D001896:Boron Compounds; D002412:Cations; D009465:Neuromuscular Agents; D050053:TRPM Cation Channels; C058776:2-(2-(5-carboxy)oxazole)-5-hydroxy-6-aminobenzofuran-N,N,O-triacetic acid; D013096:Spermine; D009532:Nickel; D015057:1-Naphthylamine; C109986:2-aminoethoxydiphenyl borate; C486584:Trpm7 protein, rat; D008274:Magnesium; D011188:Potassium; D016257:Fura-2