Mitochondrial membrane potential changes in osteoblasts treated with parathyroid hormone and estradiol

Exp Cell Res. 1997 Jun 15;233(2):274-80. doi: 10.1006/excr.1997.3570.

Abstract

This study assessed mitochondrial membrane potential changes in cultured osteoblasts treated with hormones known to regulate osteoblasts. A fluorescent carbocyanine dye, 5,5',6,6'-tetrachloro-1,1',3,3'-tetraethylbenzimidazolocarbocyanine++ + iodide, also called JC-1, was used as a probe. JC-1 emits photons at 585 nm (orange-red) when the membrane potential in mitochondria is highly negative, but when the potential becomes reduced emission occurs at 527 nm (green). Osteoblasts were rinsed in serum-free medium for 5 min, then loaded with 1 x 10(-6) M JC-1 for 10 min. The distribution and intensity of JC-1 fluorescence were evaluated with a laser-scanning confocal microscope system. Hormone treatments included parathyroid hormone (PTH; 10(-8) M), 17beta-estradiol (10(-8) M), and thyroxine (T4; 10(-8) M). The potassium ionophore valinomycin (10(-6) M) was used as a control since it is known to disrupt the electrochemical gradient of mitochondria without interfering with the pH gradient. Valinomycin caused a profound, rapid increase (22.5% above untreated values) in the green/red ratio, which indicated a lowering of the mitochondrial membrane potential in all samples evaluated. PTH caused a less pronounced, but significant (7-14%), reduction in membrane potential in all cells examined. PTH is known to affect osteoblasts in a number of ways and is inhibitory to mitochondrial respiration; the results confirm this effect. For estradiol, half of the cells responded at a significant level, with a membrane potential reduction of 6 to 13% being recorded; the other half did not respond. Thyroxine did not alter mitochondrial membrane potential. Responses were detectable 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+.

Publication types

  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Animals
  • Cells, Cultured
  • Chickens
  • Estradiol / pharmacology*
  • Fluorescent Dyes
  • Intracellular Membranes / physiology
  • Male
  • Membrane Potentials / drug effects
  • Microscopy, Fluorescence
  • Mitochondria / physiology*
  • Osteoblasts / drug effects
  • Osteoblasts / physiology*
  • Parathyroid Hormone / pharmacology*
  • Potassium / physiology
  • Thyroxine / pharmacology
  • Valinomycin / pharmacology

Substances

  • Fluorescent Dyes
  • Parathyroid Hormone
  • Valinomycin
  • Estradiol
  • Thyroxine
  • Potassium