Bison seminal plasma factor affects sperm plasma membrane integrity

Abstract

ABSTRACT

The bison industry has been growing exponentially over the last few decades; with a 35 % increase in the number of bison within Canada from 2001 - 2006 and bison meat selling at a record high. This growth is important for the Canadian and American economy, especially in light of recent setbacks in the beef market. Presently, bison ranchers have not made use of reproductive technologies that are commonly practiced in the dairy and beef industries. In order to optimise financial gain it would be advantageous to make use of these technologies within the bison industry. One issue that has interfered with ranchers being able to make use of reproductive technologies is; bison are wild animals being kept in captivity. This means that bison become highly stressed while being handled in close contact with humans. When stressed, the bison become unpredictable and dangerous, possibly hurting themselves and the handlers. Moreover, stressed bison are difficult to electroejaculate and the collected semen is low quality.

The first objective of this study was to find a way to lower the stress in bison during handling in order to improve methods for semen collection. It was hypothesised that a long acting tranquilizer (Piportil®) would lower the stress levels in the bulls, and improved the quality of collected semen. The results of the study that make up this thesis demonstrated that the animals treated with 200 mg/bull of Piportil® were less stressed when moved through the hydraulic-controlled chute system (P < 0.05). Also, treated bulls had lowered endocrine stress indicators during handling and semen collection while testosterone production was increased for treated bulls compared to their control (corticosterone: 0.101 ± 0.01 vs. 0.145 ± 0.02 ng/mL; testosterone: 9.107 ± 1.68 vs. 5.327 ± 0.74 ng/mL) respectively (P < 0.05). Moreover, the quality of semen collected from bulls treated with 200 mg of Piportil® was significantly better when compared to untreated bulls (P < 0.05). No detrimental side effects were observed when using Piportil® on the bison bulls, allowing us to suggest that this drug provided a way of lowering level and increasing semen quality. It has been suggested, that there maybe a factor in bison seminal plasma that affects the sperm plasma membrane causing increased damage during cryopreservation; resulting in low post thaw motility. However, this factor appears to be captured in the low density lipoprotein (LDL) fraction of an animal based extender. The objective of this part of the study was to identify the bison seminal plasma proteins that were captured by the LDL in Triladyl®.

Bison seminal plasma proteins that were associated with either LDL from Triladyl® or the phospholipids from Andromed® were analysed using 2 dimensional electrophoreses (2DE) (47 and 21 protein spots respectively). From these spots, 17 protein spots of interest were identified by Matrix-Associated Laser Desorption Ionization Time of Flight (MALDI-TOF). Once the protein identities were known, the main functions of these proteins were identified. It was found that 6 of the proteins had functions directly involved with the spermatozoa. From these 6 proteins, only one type had a function that could possibly be associated with sperm being unable to survive the cryopreservation procedures. These proteins are called binder of sperm proteins (BSPs) and they are involved with cholesterol and phospholipid efflux from the sperm plasma membrane, initiating capacitation. However, a procedure must still be developed to prove whether BSP is the only interfering factor.

In conclusion, the use of LAN will help to reduce stress in bison bulls during handling and electroejaculation and may increase the quality of collected semen. This semen can then be used in a variety of reproductive technologies within the bison industry. The use of these reproductive technologies will be further enhanced once a method to successfully cryopreserve bison semen is developed. The first step in this process will be to identify the seminal plasma factor that is interfering with the sperm ability to survive cryopreservation and then to develop a method to neutralize its actions. The realization of these goals will be beneficial to bison producers in Canada and the United States to increase economic gain in the future.