116 Role of GnRH-II and its receptor in porcine sperm function

Pigs are the only livestock species encoding a functional protein for the second form of gonadotrophin-releasing hormone (GnRH-II) and its receptor (GnRHR-II), which are both abundantly produced in porcine testes. Moreover, GnRHR-II is localised to the connecting piece of ejaculated sperm, whereas GnRH-II is detected in seminal plasma, suggesting an interaction influencing sperm function. Therefore, our objective was to determine effects of GnRH-II analogues on extended boar semen. Ejaculates were collected (Day 0) from white crossbred boars (n = 3–4) and extended (37.5 × 10⁶ spermatozoa mL⁻¹) in Beltsville Thaw Solution. Following extension, doses were treated with increasing concentrations of the GnRH-II agonist, D-ala⁶ GnRH-II (0, 0.01, 0.1, 1.0, 10, or 100 µM; Exp. 1) or the GnRHR-I/GnRHR-II antagonist, SB-75 (0, 0.01, 0.1, 1.0, or 10 µM; Exp. 2), diluted in dimethyl sulfoxide (DMSO). In a rescue experiment (Exp. 3), semen doses were treated with 10 µM SB-75 in the presence or absence of 100 µM D-ala⁶ GnRH-II or with vehicle alone. On Day 5 of storage at 17°C, computer-assisted sperm analysis (CASA; IVOS II Animal) was performed to determine measures of sperm motion (motility (MOT), progressive motility (PROG), slow, static), morphology (normal morphology (NM), bent tail (BT), coiled tail (CT), distal droplet (DD), proximal droplet (PD), distal midpiece reflex (DMR), area), and kinematics (length of average path (DAP), length of straight line path (DSL), length of curvilinear path (DCL), average path velocity (VAP), straight line velocity (VSL), curvilinear velocity (VCL), straightness (STR), linearity (LIN), amplitude of lateral head displacement (ALH), beat-cross frequency (BCF), and wobble (WOB)). Data were analysed using the Mixed procedure of SAS (SAS Institute Inc.), with treatments compared to vehicle controls using Dunnett’s test. In Exp. 1, only two sperm parameters were affected by any concentration of D-ala⁶ GnRH-II; PROG was higher in samples treated with 100 µM D-ala⁶ GnRH-II and fewer BT were present in doses exposed to 0.01, 0.1, and 100 µM D-ala⁶ GnRH-II compared with DMSO-treated controls (P < 0.05). Conversely, samples treated with 10 µM SB-75 (Exp. 2) possessed sperm that were more static; had lower MOT, PROG, NM, DAP, DSL, DCL, VAP, VSL, VCL, STR, and LIN; exhibited more BT, DD, and PD; had higher BCF (P < 0.05); and tended to have more CT (P < 0.10) compared with vehicle controls. No other SB-75 concentration influenced any CASA measurement (P > 0.10). In Exp. 3, 10 µM SB-75 altered the same sperm characteristics as in Exp. 2, except STR, LIN, and CT were not different (P > 0.10) and ALH tended to be higher (P < 0.10) than in vehicle-treated doses. However, 10 µM SB-75 in the presence of 100 µM D-ala⁶ GnRH-II was only able to rescue the sperm traits of BT and BCF because they did not differ from DMSO-treated controls (P > 0.10). In conclusion, we inferred that GnRH-II and its receptor are important in regulating sperm motion, morphology, and kinematics, representing a potential avenue to improve commercially extended boar semen.

This research was funded by USDA/NIFA AFRI (2017-67015-26508).