MITOCHONDRIAL-MEMBRANE POTENTIAL CHANGES IN OSTEOBLASTS TREATED WITH PARATHYROID-HORMONE AND ESTRADIOL

This study assessed mitochondrial membrane potential changes in cultur ed osteoblasts treated with hormones known to regulate osteoblasts., A fluorescent carbocyanine dye, 5,5', 6,6'-tetrachloro-1,1', 3,3'-tetra ethylbenzimidazolocarbocyanine iodide, also called JC-l, was used as a probe. JC-l emits photons at 585 nm (orange-red) when the membrane po tential in mitochondria is highly negative, but when the potential bec omes reduced emission occurs at 527 nm (green). Osteoblasts were rinse d in serum-free medium for 5 min, then loaded with 1 x 10(-6) M JC-l f ar 10 min, The distribution and intensity of JC-l fluorescence were ev aluated with a laser-scanning confocal microscope system. Hormone trea tments included parathyroid hormone (PTH; 10(-8) M), 17 beta-estradiol (10(-8) M), and thyroxine (T-4; 10(-8) M), The potassium ionophore va linomycin (10(-6) M) was used as a control since it is known to disrup t the electrochemical gradient of mitochondria without interfering wit h the pH gradient. Valinomycin caused a profound, rapid increase (22.5 % above untreated values) in the green/red ratio, which indicated a lo wering of the mitochondrial membrane potential in all samples evaluate d. PTH caused a less pronounced, but significant (7-14%), reduction in membrane potential in all cells examined, PTH is known to affect oste oblasts in a number of ways and is inhibitory to mitochondrial respira tion; the results confirm this effect. For estradiol, half of the cell s responded at a significant level, with a membrane potential reductio n of 6 to 13% being recorded; the other half did not respond. Thyroxin e did not alter mitochondrial membrane potential. Responses were detec table within 20 s for valinomycin, but occurred at a slower rate, over 200 to 300 s, following PTH and estradiol treatment. Responses to PTH and estradiol could be due to mitochondrial uptake of cytosolic Ca2+. (C) 1997 Academic Press.