223 THE EFFECT OF IN VITRO FERTILIZATION MEDIA AND THE CAPACITATION AGENT ON EPIDIDYMAL WHITE-TAILED DEER SPERMATOZOA

Abstract

To determine the requirements for in vitro production of white-tailed deer embryos, media components that induce sperm capacitation and an acrosome reaction (AR) should first be identified. Typically, in vitro production protocols are species specific with regard to media composition and capacitating agents (CA). Comparison of the bovine and red deer systems revealed a distinct difference in the CA utilised [heparin v. 20% sheep serum (SS), respectively] and in the amount of bicarbonate and Ca²⁺ necessary for fertilization to occur. The aim of this study was to evaluate the ability of 2 CA (heparin and SS) to induce the capacitation of white-tailed deer epididymal sperm in either a standard bovine (BIVF) or red deer-formulated (DSOF) fertilization medium. Epididymal spermatozoa were collected from fair-chase bucks (n = 15), extended with Triladyl (48 to 147 × 10⁶ sperm mL^–1), and frozen in LN₂ vapor. Live spermatozoa were selected from frozen–thawed semen samples from each buck and reconstituted (55 × 10⁶ sperm mL^–1) in either BIVF or DSOF and supplemented with either heparin (10 µg mL^–1) or SS, or were nontreated controls. Samples were then incubated at 37°C for either zero [treatment (T) 0], 2 (T2), or 4 (T4) hours and the AR was artificially induced using lysophosphatidylcholine (100 µg mL^–1). After AR induction, treatments were labelled with PI/PNA and M540/YoPro-1 so that live cells, acrosomal status, and membrane lipid disorders could be analysed by flow cytometry after each incubation period (T0, T2, and T4). The media (P = 0.42), CA (P = 0.38), or their interaction (P = 0.68) did not affect the percentage of live acrosome-intact spermatozoa across time (P = 0.73). The percentage of lipid disruption was higher (P < 0.001) in SS and changed (P = 0.005) across time. Data obtained from this study showed that unlike for red deer, increased bicarbonate and Ca²⁺ are not necessary to induce capacitation of white-tailed deer epididymal spermatozoa. Although SS did not appear to affect the number of live acrosome-intact spermatozoa, its presence did alter the lipid architecture of the sperm plasma membrane.