119 hormone-assisted reproduction in a captive mountain yellow-legged frog population

Severe declines of the mountain yellow-legged frog (MYLF, Rana muscosa) led to establishment of a captive population at the San Diego Zoo Institute for Conservation Research. With less than 200 adults estimated to remain in the wild, the Institute’s MYLF colony currently holds approximately 30% of the entire gene pool. Simulating natural seasonality by artificial brumation and manipulation of atmospheric temperature and light cycles is an integral part of this species’ management in captivity. However, over 5 years, the number of females that have oviposited has decreased from 80% in 2011 to 28% in 2014. It is unclear if changes in reproduction are related to husbandry or a lack of information regarding the natural reproductive cycles of these animals. Therefore, we examined the effects of hormone treatments on reproduction in the MYLF captive population. Prior to breeding, 21 females and 18 males were evenly assigned to either a control group (IP injection of PBS) or hormone-treated group [IP injection of gonadotropic releasing hormone (GnRH, D-Ala6, des-GLy10 ethylamide LHRH derivative) at 0.4µgg-1 of body weight and metoclopramide (dopamine inhibitor) at 10µgg-1 of body weight]. Males and females were housed in groups of 3 males and 3 females and allowed to choose their mate. Once a male had amplexed a female they were housed in pairs until oviposition occurred or until males ceased amplexus. Responses recorded during this study included amplexus, spermiation, oviposition, and embryonic cleavage. There was no significant difference in the number of eggs deposited by females treated with hormones and untreated controls (P=0.1949) nor were there any differences between groups in the number of embryos that cleaved (P=0.673) or survived to tadpole stage (P=0.629). No significant differences were detected between the numbers of males that amplexed in the control or treated groups (P=0.1120). Urine collected from 10/18 amplexed males (7 hormone-treated and 3 controls) indicated that 57% percent of hormone-treated males and 67% of control males were spermiating at the time of collection. Therefore, hormone treatments did not increase the number of eggs oviposited or the number of males spermiating or amplexing. Closer analysis of individual female reproductive histories indicated that 48% of captive female MYLF oviposited just once in 5 years, 40% in 2 consecutive years, 8% oviposited eggs in 2 consecutive years, skipped a year, and then oviposited again in 2 consecutive years, and 4% oviposited in 3 consecutive years. The number of females ovipositing in 2015 was significantly higher than 2012 (P=0.0002), 2013 (P=0.0001), and 2014 (P=0.0026), but not 2011 (P=0.0885), suggesting breeding may not occur annually in females. Understanding the breeding cycles of MYLF females will enable managers to determine if and when hormone administration is efficacious in captive amphibian breeding populations.