Fluorescent monitoring of Jurkatt cell intracellular magnesium during metabolic poisoning.
Divalent cation movement characterizes the final common pathway of cellular death from ischemic or metabolic injury. The influx of calcium is an essential step in cellular death. We hypothesized that intracellular magnesium levels may change during the progression to cellular death. Verapamil-sensitive changes in free ionized intracellular Mg2+ ([Mg2+[i) and Ca2+ ([Ca2+]i) levels were estimated in transformed T-lymphocytes exposed to metabolic inhibitors. Separate experiments used a Mg(2+)-sensitive fluoroprobe, fura-2 (Ex 1,344, Ex 2,376, Em 500), and a Ca(2+)-sensitive fluoroprobe, fura-2 (Ex 1,340, Ex 2,380, Em 510). Chemical anoxia (sodium cyanide 1 mM, iodoacetic acid 10 mM) caused a gradual increase in [Ca2+]i (control 126 +/- 13 nM) to > 1 mM by 10 min. This increase in [Ca2+]i was not affected by verapamil treatment. In separate experiments, [Mg2+]i levels were monitored during chemical anoxia. The specificity of mag-fura for Mg2+ over Ca2+ was reflected in the absence of a response to the lymphocyte Ca2+ mobilizer OKT-3. Uncorrected control [Mg2+]i levels (.4 +/- .1 mM) were not affected by the combined cyanide-iodoacetate treatment. A small increase in mag-fura-2 fluorescence was noted, probably due to binding of Ca2+ to the fluoroprobe when [Ca2]i exceeded 1 mM. Elimination of Ca2+ from the extracellular buffer increased the resting estimate of intracellular [Mg2+] to 1.6 + .1 mM. These results indicate that 1) extracellular Ca2+ can interfere with the fluorescent determination of intracellular magnesium concentration, and 2) intracellular free Mg2+ concentrations do not change in this cell line during chemical anoxia.
Carroll
R G
RG
Department of Physiology, East Carolina University School of Medicine, Greenville, North Carolina 27858-4354, USA.
Thomas
J M
JM
D000255:Adenosine Triphosphate / Q000378:metabolism; D002118:Calcium / Q000494:pharmacology; D002412:Cations; D016923:Cell Death / Q000502:physiology; D002460:Cell Line; D005456:Fluorescent Dyes / Q000737:chemistry* / Q000378:metabolism; D006801:Humans; D007461:Iodoacetates / Q000506:poisoning*; D019807:Iodoacetic Acid; D008274:Magnesium / Q000378:metabolism*; D011190:Potassium Cyanide / Q000506:poisoning*; D013601:T-Lymphocytes / Q000187:drug effects / Q000378:metabolism* / Q000473:pathology
D002412:Cations; D005456:Fluorescent Dyes; D007461:Iodoacetates; D000255:Adenosine Triphosphate; D008274:Magnesium; D011190:Potassium Cyanide; D002118:Calcium; D019807:Iodoacetic Acid