221 PREPUBERTAL MOUSE BIOASSAY FOR OVULATION-INDUCING FACTOR IN SEMINAL PLASMA

Abstract

A substance in the seminal plasma of llamas and alpacas has been discovered that induces ovulation and growth of the corpus luteum (CL) in the female of the same species. The ovarian effects of the ovulation-inducing factor (OIF) are associated with a surge release of LH into circulation. Ultrasonographic detection of ovulation and CL development is currently the only method available for testing the bioactive effects of OIF. The purpose of this study was to determine if a superstimulatory prepubertal mouse model could be developed as an in vivo bioassay for OIF. Prepubertal female CD1 mice (n = 144), 20 days of age and weighing 20–25 g, were housed at 24°C with lights on from 0500 to 1900 h and free access to food and water. An intramuscular dose of 5 IU of eCG (Novormon, Bioniche Animal Health, Belleville, ON, Canada) was given (Day 0) for ovarian superstimulation. On Day 2, mice were assigned randomly to 4 groups (n = 36 per group) and given a single 0.1 mL intraperitoneal dose of (1) 5 IU of hCG (Chorulon, Intervet Canada, Ltd., Whitby, ON, Canada), (2) 5 µg GnRH (gonadotropin-releasing hormone: Cystorelin, Merial, Ltd., Iselin, NJ, USA), (3) llama seminal plasma, or (4) phosphate-buffered saline (negative control). On Day 3, females were euthanized by an overdose of inhaled halothane. Oviducts were collected and oocytes were counted using trans-illumination stereomicroscopy. The proportion of mice that ovulated did not differ among groups treated with hCG, GnRH, and seminal plasma (31/36, 31/36, 28/36, respectively); however, the proportion of mice that ovulated in each treatment group was greater than that in the saline-treated group (9/36) (P < 0.001). The number of oocytes counted (mean ± SEM) was also similar among groups treated with hCG (25.8 ± 2.9), GnRH (27.4 ± 2.7), and seminal plasma (19.2 ± 2.8), all of which were greater (P < 0.01) than in the saline-treated group (6.2 ± 2.1). We conclude that the superstimulated prepubertal CD1 mouse model is effective as an in vivo bioassay for OIF in seminal plasma. Whether the bioassay may be used for quantitative estimates of OIF activity will require dose-response trials using serial dilutions of seminal plasma.