165 EFFECTS OF AGE, HORMONES AND HIBERNATION ON BREEDING SUCCESS IN BOREAL TOADS (BUFO BOREALIS)

The global amphibian extinction crisis has stimulated interest in captive breeding programs for this diverse taxa. The goals of this study were to test the effects of exogenous hormone injections (LHRH and hCG) and hibernation on breeding behavior and gamete release by the boreal toad, a temperate species. The 5-year study was initiated with 77 toadlets. Each year, a subset of female and/or male toads was hibernated for 5 months at 4°C. The remaining toads were maintained in a lab receiving 12 h of light daily and ample food and water. Within 48 h of removal from hibernation, toads were paired with hibernated or non-hibernated mates. All toads were weighed and pairs were randomly assigned to treatments. Toads were injected IP with 6 μg LHRH, 250 IU hCG, or saline control and observed for amplexus and egg production. Some treatment groups received a second hormone injection if eggs were not produced within 48 h. Eggs were counted and transferred to aquariums for tadpole and toadlet development. During years 1 to 3, sperm production was assessed in male toads by sampling tank water or urine samples after injections. Data were analyzed using unpaired t-tests and chi-square analyses. In year 1, only 19% of pairs overall exhibited amplexus, none of the males produced sperm and none of the females produced eggs. In year 2, only male toads injected with LHRH exhibited amplexus, and it was more prevalent (P < 0.05) in those that were hibernated than non-hibernated (54.0% and 33.3%, respectively). Percentage of pairs in amplexus in years 2 and 3 (56.9% and 100%) was greater (P < 0.05) than in year 1. Only one and two females produced eggs in years 2 and 3, respectively. In contrast, most hibernated (87.5%) and non-hibernated (75%) males produced sperm. In year 4, eight females produced eggs, but two of these died egg-bound. Interestingly, more (P < 0.05) non-hibernated (7/10) than hibernated (1/10) females produced eggs despite no differences (P ≥ 0.05) in proportion of pairs amplexing or time spent in amplexus between groups. Mean (±SD) weight of female toads producing eggs (58.9 ± 11.9 g) was greater (P < 0.05) than that for non-producing females (43.6 ± 7.0 g). Similarly, 4/7 non-hibernated females produced eggs in year 5, and their weights averaged (58.8 ± 8.3 g). All eggs were produced by females injected a single time with LHRH. Number of eggs per female varied (141–3307), and percentage of eggs developing to tadpoles was low (0–36.5%), but tadpoles were capable of developing into adult toads. In conclusion, two-year-old male boreal toads are mature, exhibiting amplexus and releasing sperm in response to LHRH injections. Females appear to mature at four years of age, and eggs are more likely to be produced by heavier females. Therefore, to enhance breeding success, males should be hibernated and injected with LHRH. In contrast, better productivity in females may be achieved by improving their body condition instead of subjecting them to winter hibernation prior to LHRH injection. Supported by: AZA Conservation Endowment Fund and Colorado Division of Wildlife.