Down-regulation of mt1 melatonin receptors in rat ovary following estrogen exposure

Life Sci. 2001 May 25;69(1):27-35. doi: 10.1016/s0024-3205(01)01097-9.

Abstract

In this study, we have demonstrated that 2-[125I]-iodomelatonin binds specifically to rat ovarian granulosa cell (GC) membranes with high affinity (KD=83 pM; Bmax=3.28 fmol/mg protein). Using immunoblot analysis and an anti-mt1 melatonin receptor antibody, we have also detected mt1 melatonin receptors in rat ovary. Because melatonin has been reported to alter the steroidogenic responses of ovarian tissues to gonadotropins, a physiological role for intra-ovarian melatonin may exist. Thus, in order to investigate a possible intra-ovarian role for melatonin, we have used both an in vivo and in vitro model of follicular development. Treatment of immature (day 21) female rats with estradiol (E; 0.2 mg/d x 3 d; subcutaneous) was used to induce follicular growth. Membranes from both untreated (U) and E-treated animals' ovaries contained high-affinity 2-[125I]-iodomelatonin (I-MEL) binding sites (Kd=83 and 23 pM, respectively). Estradiol treatment in vivo caused a significant decrease (P<0.05) in binding of 2-[125I]-iodomelatonin to ovarian membranes with untreated animals' ovaries having a Bmax=3.28 fmol/mg protein vs. estradiol-treated animals' ovaries having a Bmax=0.92 fmol/mg protein. In addition, following Estradiol treatment, mt1 melatonin receptors in rat ovary were down-regulated (approximately 95%) using immunoblot analysis. Granulosa cells isolated from E-treated rats were further matured in vitro with testosterone (T) and the pituitary gonadotropin follicle-stimulating hormone (FSH). Granulosa cells were cultured with either T (10 ng/ml) or FSH (5.71 ng ovine FSH-20/ml) alone, or both FSH and T for 48 h. There was no statistically significant specific binding of 2-[125I]-iodomelatonin to GC membranes cultured with T or FSH alone. However, following a 48-h exposure to FSH and T in vitro specific 2-[125I]-iodomelatonin binding occurred with total 2-[125I]-iodomelatonin binding =3.15 [corrected] fmol/mg protein. Therefore, the existence of hormonally-regulated expression of high-affinity melatonin binding sites suggests that melatonin may have an important intra-ovarian physiological role.

Publication types

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

MeSH terms

  • Animals
  • Cell Membrane / physiology
  • Down-Regulation / drug effects
  • Estradiol / pharmacology*
  • Female
  • Follicle Stimulating Hormone / pharmacology
  • Granulosa Cells / drug effects
  • Granulosa Cells / physiology
  • Iodine Radioisotopes
  • Kinetics
  • Melatonin / analogs & derivatives
  • Melatonin / pharmacokinetics
  • Melatonin / physiology
  • Ovarian Follicle / drug effects
  • Ovarian Follicle / physiology
  • Ovary / drug effects
  • Ovary / physiology*
  • Rats
  • Rats, Sprague-Dawley
  • Receptors, Cell Surface / drug effects
  • Receptors, Cell Surface / metabolism*
  • Receptors, Cytoplasmic and Nuclear / drug effects
  • Receptors, Cytoplasmic and Nuclear / metabolism*
  • Receptors, Melatonin
  • Testosterone / pharmacology

Substances

  • Iodine Radioisotopes
  • Receptors, Cell Surface
  • Receptors, Cytoplasmic and Nuclear
  • Receptors, Melatonin
  • Testosterone
  • Estradiol
  • Follicle Stimulating Hormone
  • 2-iodomelatonin
  • Melatonin