Intracellular calibration of the fluorescent Mg2+ indicator furaptra in rat ventricular myocytes.
Single ventricular myocytes enzymatically isolated from rat hearts were loaded with the Mg2+ indicator furaptra, and the relationship between the fluorescence ratio signal (R) and the intracellular free concentration of Mg2+ ([Mg2+]i) was studied in situ at 25 degrees C. After the application of ionophores (ionomycin, monensin, nigericin and valinomycin), an immediate change in furaptra R was noted, followed by a slow change in R that reached a steady level in 2-4 h. The direction of the early change in R that accompanied rigor contraction was independent of the extracellular Mg2+ concentration ([Mg2+]o), and was consistent with the breakdown of ATP and release of bound Mg2+. The intracellular calibration curve was constructed from the steady levels of R obtained at various [Mg2+]o between 0 and 47 mM. The dissociation constant of intracellular furaptra was estimated to be 5.3 mM, which was 44% higher than that determined in salt solutions (3.7 mM). The basal [Mg2+]i of rat ventricular myocytes calculated with the intracellular curve averaged 0.91 mM.
Watanabe
M
M
Department of Physiology, Tokyo Medical University, 6-1-1 Shinjuku, Shinjuku-ku, Tokyo 160-8402, Japan.
Konishi
M
M
D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D002138:Calibration; D005453:Fluorescence; D016257:Fura-2 / Q000031:analogs & derivatives* / Q000032:analysis* / Q000737:chemistry; D006352:Heart Ventricles; D015759:Ionomycin / Q000494:pharmacology; D007476:Ionophores / Q000494:pharmacology; D008274:Magnesium / Q000032:analysis* / Q000378:metabolism; D008297:Male; D008839:Microelectrodes; D008985:Monensin / Q000494:pharmacology; D009206:Myocardium / Q000737:chemistry* / Q000378:metabolism; D009550:Nigericin / Q000494:pharmacology; D051381:Rats; D017208:Rats, Wistar; D013050:Spectrometry, Fluorescence; D014634:Valinomycin / Q000494:pharmacology
D007476:Ionophores; C058776:2-(2-(5-carboxy)oxazole)-5-hydroxy-6-aminobenzofuran-N,N,O-triacetic acid; D014634:Valinomycin; D015759:Ionomycin; D000255:Adenosine Triphosphate; D008985:Monensin; D008274:Magnesium; D009550:Nigericin; D016257:Fura-2