Evidence that the arcuate nucleus is an important site of progesterone negative feedback in the ewe

Abstract

There is now considerable evidence that dynorphin neurons mediate the negative feedback actions of progesterone to inhibit GnRH and LH pulse frequency, but the specific neurons have yet to be identified. In ewes, dynorphin neurons in the arcuate nucleus (ARC) and preoptic area (POA) are likely candidates based on colocalization with progesterone receptors. These studies tested the hypothesis that progesterone negative feedback occurs in either the ARC or POA by determining whether microimplants of progesterone into either site would inhibit LH pulse frequency (study 1) and whether microimplants of the progesterone receptor antagonist, RU486, would disrupt the inhibitory effects of peripheral progesterone (study 2). Both studies were done in ovariectomized (OVX) and estradiol-treated OVX ewes. In study 1, no inhibitory effects of progesterone were observed during treatment in either area. In study 2, microimplants of RU486 into the ARC disrupted the negative-feedback actions of peripheral progesterone treatments on LH pulse frequency in both OVX and OVX+estradiol ewes. In contrast, microimplants of RU486 into the POA had no effect on the ability of systemic progesterone to inhibit LH pulse frequency. We thus conclude that the ARC is one important site of progesterone-negative feedback in the ewe. These data, which are the first evidence on the neural sites in which progesterone inhibits GnRH pulse frequency in any species, are consistent with the hypothesis that ARC dynorphin neurons mediate this action of progesterone.

Fig. 1.

Crossover experimental designs for study 1 (A) and study 2 (B). Treatments with peripheral progesterone (CIDR) and E₂ implants (starting at time of OVX and neurosurgeries) are indicated by gray and stippled bars, respectively. Frequent blood sampling (Bl) and microimplant treatments (study 1: black/open bars, progesterone/control; study 2: striped/open bars, RU486/control) are shown above peripheral implants. The treatment protocol for the first half of each study (in OVX+E ewes) is depicted; identical protocols were used for the second half of each study (OVX ewes), except that E₂ implants were removed and two vaginal progesterone implants were inserted during RU486/control microimplant treatments in study 2. See text for more details.

Fig. 2.

Sites of microimplants in study 1. The top three panels depict bilateral microimplants in the ARC; the bottom three panels are sites in the POA. Bilateral sites in the same ewe are connected by a line. The shaded area depicts distribution of PR-containing dynorphin neurons based on previous data (25). Ac, Anterior commissure; fx, fornix; ME, median eminence; OCh, optic chiasm; Pit, pituitary stalk; VMH, ventromedial hypothalamus.

Fig. 3.

The top two panels depict representative LH pulse patterns in OVX ewes before (d 0) and during (d 3) treatment with progesterone-containing microimplants in the ARC or POA. Solid circles depict peaks of LH pulses. The bottom two panels present mean (±sem) LH pulse frequency before (open bars) and during treatment with empty (C) (shaded bars) or progesterone-filled (solid bars) microimplants in the ARC or POA. Results from OVX+E ewes are shown on the left, and results from OVX ewes are shown on the right. There were no significant effects of progesterone. P, Progesterone.

Fig. 4.

Effects of ARC RU486 microimplants. Left panels depict representative episodic LH secretion in OVX+E (top panels) and OVX (bottom panels) ewes receiving empty (Cont) or RU486-containing (RU486) microimplants in the ARC. LH pulse patterns before (d 0) and during (d 2) one vaginal progesterone implant are shown. Solid circles depict peaks of LH pulses. Right panels present corresponding changes in the LH pulse frequency (FREQ) and amplitude (AMPL) when ewes received empty (C) or RU486-containing (R) microimplants. Open bars depict values before and solid/shaded bars values during vaginal progesterone treatment. *, P < 0.05 vs. values before progesterone treatment.

Fig. 5.

Effects of POA RU486 microimplants. Left panels depict representative episodic LH secretion in OVX+E (top panels) and OVX (bottom panels) ewes receiving empty (Cont) or RU486-containing (RU486) microimplants in the POA. LH pulse patterns before (d 0) and during (d 2) treatment with one (OVX+E ewes) or two (OVX ewes) vaginal progesterone implants are shown. Solid circles depict peaks of LH pulses. Right panels present corresponding changes in LH pulse frequency (FREQ) and amplitude (AMPL) when ewes received empty (C) or RU486-containing (R) microimplants. Open bars depict values before and solid/shaded bars values during vaginal progesterone treatment. *, P < 0.05 vs. values before progesterone treatment.