Mouse Placental Cells Secrete Immunoreactive Growth Hormone-Releasing Factor1

Abstract

The initial objective of this study was to establish a placental cell culture system in which the secretion of mouse growth hormone-releasing factor (mGHRF) could be examined during a several-day period. To determine when during pregnancy placental cells begin to express mGHRF, Northern blot analysis was carried out on total RNA from placentas collected on Days 6, 9, 11, 13, 15, 17, and 18 of pregnancy. Mouse GHRF mRNA could be detected as early as Day 11 of pregnancy. Its steady-state levels increased to maximum values on Days 15-17 and then declined slightly on Day 18. Placentas from Day 12 of pregnancy were selected for cell culture. The basal zone and labyrinth were dispersed in collagenase, and the cells were fractionated on a Percoll gradient. Two bands of cells were selected for further study. Both released significant amounts of immunoreactive mGHRF during a 5-day culture period. Effects of prolonged exposure of the cells to 8-bromo-cAMP and to agents that elevate intracellular cAMP concentration were then examined. Treatment of the cells with 0.5 mM 8-bromo-cAMP resulted in a significant decrease in the mGHRF concentration of the medium by the second day of culture. Mouse GHRF secretion was also inhibited by treatment of the cells with 100 ng/ml cholera toxin or 0.1 mM forskolin. The effect of 8-bromo-cAMP was concentration-dependent, with 0.1 mM being the lowest concentration that was active. 8-Bromo-cAMP treatment also reduced the steady-state level of mGHRF mRNA in the cells.(ABSTRACT TRUNCATED AT 250 WORDS)

Mouse Growth Hormone-Releasing Factor Secretion is Activated by Inhibin and Inhibited by Activin in Placenta

“…These findings suggest that endogenous activin regulates hCG or mGHRF expression from cultured cells derived from human or mouse placenta. We recently found that treatment of placental cells with 8-bromo cAMP, interleukin-6 (IL-6), IL-11, oncostatin M, leukemia inhibitory factor, or TGFi1 results in inhibition of mGHRF secretion [7,24]. As we demonstrated here, activin also inhibited mGHRF secretion.…”

“…Placenta was collected from mice at the 12th day of pregnancy, and placental cell cultures were performed as described previously [7,8]. Briefly, the fetuses and decidua basalis were discarded from the conceptuses, and the remaining tissue was dispersed with collagenase.…”

“…The mGHRF concentrations in the media were measured by a highly specific RIA [7,8]. The rat FSH (rFSH) concentration in the media was also determined by RIA, using the NIDDK rFSH-RP-2 materials and protocols supplied by the National Institutes of Health (Bethesda, MD).…”

“…Human, rat, and mouse placenta express growth hormone-releasing factor (GHRF) [1][2][3][4][5][6][7]. We recently established a primary culture system to study the expression of placental mouse GHRF (mGHRF) secretion and found that 8-bromo cAMP and transforming growth factor 31 (TGF31) inhibit mGHRF secretion in vitro [7,8].…”

Identification of Growth Hormone-Releasing Hormone and Somatostatin Neurons Projecting to the Median Eminence in Normal and Growth Hormone-Deficient Ames Dwarf Mice

“…Sections were then incubated for 48 h at 4°C in a rabbit polyclonal antiserum (a gift from Dr. The mGHRH antibody has been demonstrated in RIA not to cross-react with several GHRH-related peptides [35]. In the present study, GHRH and SRIH) .…”