Progestin exposure before gonadotropin stimulation improves embryo development after in vitro fertilization in the domestic cat

Abstract

This study investigated the influence of progestin priming and ovarian quiescence on response to exogenous gonadotropin stimulation in the cat. Because a subpopulation of cats routinely ovulated spontaneously, there also was the opportunity to examine the ovary's reaction to the added impact of endogenously secreted progestagen. Queens were given 1) equine chorionic gonadotropin (eCG) plus human chorionic gonadotropin (hCG) only (control; n = 9 cats), 2) GnRH antagonist (antide) injections followed by eCG and hCG (n = 9), and 3) a progestin implant (levonorgestrel) followed by eCG and hCG (n = 9). Laparoscopy was used to assess ovarian activity and aspirate follicular oocytes that were graded on the basis of morphology. In five cats per treatment, half of the high-quality oocytes were assessed for glucose, pyruvate, and lactate metabolism as well as nuclear maturation. Remaining oocytes were inseminated in vitro, cultured, and examined at 72 h after insemination for cleavage. In the remaining four cats per treatment, all oocytes were inseminated in vitro and assessed at 72, 120, and 168 h after insemination for embryo developmental stage. Cats pretreated with progestin had more follicles and produced more embryos per donor (including at the combined morula/blastocyst stage) than controls or females treated with GnRH antagonist (P < 0.05). There were no differences among groups (P > 0.05) in oocyte carbohydrate metabolism, nuclear maturation metrics, or fertilization success, although there was a tendency toward improvements in all three (P < 0.2) in progestin-treated females. Interestingly, cats that spontaneously ovulated within 60 days of treatment onset also produced more embryos per cat than induced-ovulation counterparts (P < 0.05). Results indicate that prior exposure to exogenous progestin (via implant) or endogenous progestagen (via spontaneous ovulation) improves ovarian responsiveness to gonadotropins in the cat through a mechanism that is independent of the induction of ovarian quiescence.

FIG. 1.

Ovarian grade (based on follicular response) as assessed by laparoscopy at 25–28 h post-hCG injection. A) Grade 1 (excellent) response in a cat exposed to progestin-induced ovarian suppression followed by eCG/hCG. Note enlarged ovary with multiple, antral (4–5 mm diameter), well-vascularized follicles. B) Grade 2 (good) response in a cat exposed to GnRH antagonist-induced ovarian suppression followed by eCG/hCG. C, D) Grade 3 (moderate; C) and Grade 4 (poor; D) response in cats exposed to eCG/hCG stimulation alone (control). Black arrows indicate the ovary. Each white arrowhead indicates a mature, vascularized follicle.

FIG. 2.

Kinetics of embryonic development in vitro following oocyte aspiration and IVF (0 h = time of insemination) in cats (n = 4 females per treatment) treated with A) gonadotropins alone (control), B) GnRH antagonist (antide), or C) progestin (levonorgestrel) prior to gonadotropin stimulation. Data (mean ± SEM) are for fertilized (cleaved) oocytes only. Within each time period, developmental stage, and across the three treatments, means with different letter superscripts indicate differences (P < 0.05) among treatments. Within inhibition treatments and developmental stage, different symbols indicate differences (P < 0.05) among time periods.

FIG. 3.

Variation in fecal progesterone metabolite (closed squares) response to elevated estradiol concentrations (open circles) in representative domestic cats. A) An induced ovulator exhibiting estradiol peaks with no elevations in progesterone prior to progestin (levonorgestrel) treatment (observed in 3 cats). B) A spontaneous ovulator exhibiting an estradiol peak followed by a full-length luteal phase (>3 wk) prior to progestin (levonorgestrel) treatment (observed in 4 cats). C) An induced ovulator prior to GnRH antagonist (antide) treatment (observed in three cats). D) A spontaneous ovulator exhibiting an estradiol peak followed by a full-length luteal phase (>3 wk) prior to GnRH antagonist treatment (observed in four cats). Open stars indicate estradiol peaks associated with a follicular phase. Closed stars indicate estradiol peaks associated with laparoscopically observed follicles. Light gray horizontal bars represent spontaneous ovulation events. Black horizontal bar represents the 37-day levonorgestrel implant period. Each arrow indicates an antide injection. Vertical bar represents the day of eCG injection.