Characterization of Mg2+-regulated TRPM7-like current in human atrial myocytes

Journal of Biomedical Science, Aug 2012

Background TRPM7 (Transient Receptor Potential of the Melastatin subfamily) proteins are highly expressed in the heart, however, electrophysiological studies, demonstrating and characterizing these channels in human cardiomyocytes, are missing. Methods We have used the patch clamp technique to characterize the biophysical properties of TRPM7 channel in human myocytes isolated from right atria small chunks obtained from 116 patients in sinus rhythm during coronary artery and valvular surgery. Under whole-cell voltage-clamp, with Ca2+ and K+ channels blocked, currents were generated by symmetrical voltage ramp commands to potentials between -120 and +80 mV, from a holding potential of -80 mV. Results We demonstrate that activated native current has dual control by intracellular Mg2+ (free-Mg2+ or ATP-bound form), and shows up- or down-regulation by its low or high levels, respectively, displaying outward rectification in physiological extracellular medium. High extracellular Mg2+ and Ca2+ block the outward current, while Gd3+, SpM4+, 2-APB, and carvacrol inhibit both (inward and outward) currents. Besides, divalents also permeate the channel, and the efficacy sequence, at 20 mM, was Mg2+>Ni2+>Ca2+>Ba2+>Cd2+ for decreasing outward and Ni2+>Mg2+>Ba2+≥Ca2+>Cd2+ for increasing inward currents. The defined current bears many characteristics of heterologously expressed or native TRPM7 current, and allowed us to propose that current under study is TRPM7-like. However, the time of beginning and time to peak as well steady state magnitude (range from 1.21 to 11.63 pA/pF, ncells/patients = 136/77) of induced TRPM7-like current in atrial myocytes from different patients showed a large variability, while from the same sample of human atria all these parameters were very homogenous. We present new information that TRPM7-like current in human myocytes is less sensitive to Mg2+. In addition, in some myocytes (from 24 out of 77 patients) that current was already up-regulated at membrane rupture. Conclusions This study provides the first electrophysiological description of TRPM7-like current in native human atrial myocytes. Less sensitivity to intracellular Mg2+ suggests for channel operation under physiological conditions. The TRPM7-like current up-regulation indicates the pathophysiological evidence of that current in human heart.

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Characterization of Mg2+-regulated TRPM7-like current in human atrial myocytes

Journal of Biomedical Science 2+ Characterization of Mg -regulated TRPM7-like current in human atrial myocytes Regina Macianskiene 0 Irma Martisiene 0 Danguole Zablockaite 0 Vida Gendviliene 0 0 Institute of Cardiology, Lithuanian University of Health Sciences , Sukileliu 17, LT-50009 Kaunas , Lithuania Background: TRPM7 (Transient Receptor Potential of the Melastatin subfamily) proteins are highly expressed in the heart, however, electrophysiological studies, demonstrating and characterizing these channels in human cardiomyocytes, are missing. Methods: We have used the patch clamp technique to characterize the biophysical properties of TRPM7 channel in human myocytes isolated from right atria small chunks obtained from 116 patients in sinus rhythm during coronary artery and valvular surgery. Under whole-cell voltage-clamp, with Ca2+ and K+ channels blocked, currents were generated by symmetrical voltage ramp commands to potentials between -120 and +80 mV, from a holding potential of -80 mV. Results: We demonstrate that activated native current has dual control by intracellular Mg2+ (free-Mg2+ or ATP-bound form), and shows up- or down-regulation by its low or high levels, respectively, displaying outward rectification in physiological extracellular medium. High extracellular Mg2+ and Ca2+ block the outward current, while Gd3+, SpM4+, 2-APB, and carvacrol inhibit both (inward and outward) currents. Besides, divalents also permeate the channel, and the efficacy sequence, at 20 mM, was Mg2+>Ni2+>Ca2+>Ba2+>Cd2+ for decreasing outward and Ni2+>Mg2+>Ba2+≥Ca2+>Cd2+ for increasing inward currents. The defined current bears many characteristics of heterologously expressed or native TRPM7 current, and allowed us to propose that current under study is TRPM7-like. However, the time of beginning and time to peak as well steady state magnitude (range from 1.21 to 11.63 pA/pF, ncells/patients = 136/77) of induced TRPM7-like current in atrial myocytes from different patients showed a large variability, while from the same sample of human atria all these parameters were very homogenous. We present new information that TRPM7-like current in human myocytes is less sensitive to Mg2+. In addition, in some myocytes (from 24 out of 77 patients) that current was already up-regulated at membrane rupture. Conclusions: This study provides the first electrophysiological description of TRPM7-like current in native human atrial myocytes. Less sensitivity to intracellular Mg2+ suggests for channel operation under physiological conditions. The TRPM7like current up-regulation indicates the pathophysiological evidence of that current in human heart. Atria; Human; Myocyte; Mg2+; TRPM7 Background Within the past decade, TRPM7 (Transient Receptor Potential of the Melastatin subfamily) channels [ 1 ] (also named MagNuM, for Magnesium Nucleotide-regulated Metal ion channel [ 2 ], or MIC, for Magnesium Inhibited Channel [ 3 ]) have been detected in a large number of tissues, including heart, using molecular approaches [ 2,4,5 ]. TRPM7 channels are important in human physiology, since they have been implicated in the regulation of transmembrane movement of Mg2+ in the cell [ 2,4,6-8 ] and of the cellular Mg2+ homeostasis [ 9,10 ]. Moreover, TRPM7 is required for cell viability [ 1,3,9 ], since both knockout and/ or overexpression of these channels cause loss of cell adhesion, growth arrest, and rapid cell death [ 2,9,11 ]. Certain features, including the implication in the entry of extracellular Mg2+ (Mg2o+) and other divalent cations [ 12,13 ], as well as a bifunctional property with ion channel and kinase activities [ 2,4,6 ] discriminate TRPM7 from a variety of other known ion channels [ 7,8,14,15 ]. These channels are constitutively open [2]. However, under the physiological conditions they pass very weak inward current due to substantial down-regulation by high levels of intracellular Mg2+ (Mgi2+) and MgATP. By contraries, decreased Mgi2+ (in free or ATP-bound form) up-regulates the current carried by TRPM7 channels [ 2,8,15,16 ]. Data from different groups suggest that the altered function and/or regulation of TRPM7 channels could be implicated in pathological conditions such as anoxic cell death and ischaemia-reperfusion injury [ 5 ], arrhythmias [ 17 ], atria fibrillation [ 18 ], etc. However, it is important to note that TRPM7 channels in native cells have been detected mainly by using molecular biology and immunostaining methods [ 4,6 ]. The majority of electrophysiological studies so far were carried out either on heterologously expressed or on native RBL (rat basophilic leukemia cells), human Jurkat T-cells, and fibroblasts [ 2,4,7,12,16,18 ]. Although TRPM7 proteins are highly expressed in the heart [ 4,6 ], only few studies have been performed to detect or characterize the channels in native cardiomyocytes [ 15,19,20 ]. A non-selective channel, which is blocked by extracellular divalent ca (...truncated)


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Regina Macianskiene, Irma Martisiene, Danguole Zablockaite, Vida Gendviliene. Characterization of Mg2+-regulated TRPM7-like current in human atrial myocytes, Journal of Biomedical Science, 2012, pp. 75, 19, DOI: 10.1186/1423-0127-19-75