Abstract
Anti-Müllerian hormone (AMH) is a glycoprotein produced by the granulosa cells of growing follicles and is considered a biomarker of ovarian reserve in humans,1-3 cattle,4 rodents,5 and non-human primates6. In males, AMH is produced by Sertoli cells in utero for sex differentiation, and in seasonal species such as the horse, serum concentrations of AMH in adult animals display annual variation.7 The present study aims to describe serum AMH patterns of male (99 samples from 10 males, age 0 to 37 y) and female (313 samples in 16 females, from ages 0 to 35) beluga to examine the influence of age, sexual maturation and reproductive season.
Females were grouped by age in 5-year increments (from 1 to 35 y) and by maturation status as follows: immature (< 6 y), mature (> 6 and < 30 y), and senescent (> 30 y). Mature females were also categorized into four groups according to season:8 breeding season (February–May), transition out of season (June–July), anestrus (August–November), and transition into season (December–January). For males, at each year of age, one sample from the breeding season (January–July) or non-breeding season (August–December) was analyzed for AMH and testosterone (T). Males at each year of age were categorized based on T, determined during the breeding season, as follows: immature (T < 0.5 ng/ml), pubescent (T ≥ 0.5 and ≤ 2.0 ng/ml), mature (T > 2.0 ng/ml) or aged (T ≤ 2.0 ng/ml and > 30 y). Results were analyzed using a mixed regression model with animal ID as the random variable. Significant (p < 0.05) differences in AMH concentrations between male (1917.1 ± 1082.0 ng/ml, n = 108) and female (5.5 ± 0.1 ng/ml, n = 310) beluga across all ages were detected, thus the sexes were analyzed separately. Across age group categories, the mean AMH concentration peaked at 6–10 y (6.71 ± 0.81 ng/ml) and continued to decrease (p < 0.05) after age 15. AMH concentrations did not differ between immature (5.2 ± 4.8 ng/ml) and mature females (5.8 ± 3.9 ng/ml). However, senescent females (2.5 ± 1.4 ng/ml) presented a lower (p < 0.05) concentration of AMH than mature females. There were no significant differences in AMH concentrations across reproductive seasons. For males, T values were higher (p < 0.05) in mature (8.2 ± 4.6 ng/ml) compared to immature (0.1 ± 0.1 ng/ml) animals. Significant effects of age and season on T concentrations were detected, but there were no effects of season on AMH values. However, there was a correlation between AMH and T (r = 0.28, p < 0.004). Aged males (509.4 ± 432.0 ng/ml) had reduced (p < 0.005) AMH concentrations when compared to immature (1832.3 ± 217.8 ng/ml), pubescent (2299.1 ± 343.1 ng/ml), and mature animals (2148.6 ± 214.6 ng/ml).
This is the first time AMH has been characterized in a cetacean species and the first potential hormonal evidence of senescence in beluga. Further research will be required to determine if this hormone can be used as a predictor of fertility for the species.
Acknowledgements
The authors wish to thank the staff at the SeaWorld & Busch Gardens Reproductive Research Center and the staff of the veterinary, animal care, training and laboratory departments at SeaWorld San Diego, San Antonio and Orlando. We thank the U.S. Navy Marine Mammal Program and Sam Ridgway, Celeste Parry, and Risa Daniels of the National Marine Mammal Foundation for providing serum samples from female belugas for analysis. We thank Brad Andrews (SEA) for his support of this project.
* Presenting author
Literature Cited
1. van Rooij IAJ, Broekmans FJM, Scheffer GJ, et al. Serum anti-müllerian hormone levels best reflect the reproductive decline with age in normal women with proven fertility: longitudinal study. Fertil Steril. 2005;83:979–987.
2. Leeners B, Geraedts K, Imthurn B, Stiller R. The relevance of age in female human reproduction - current situation in Switzerland and pathophysiological background from a comparative perspective. Gen Comp Endocrinol. 2013;188:166–174.
3. de Vet A, Laven JSE, de Jong FH, Themmen APN, Fauser BCJM. Anti-müllerian hormone serum levels: a putative marker for ovarian aging. Fertil Steril. 2002;77:357–362.
4. Ireland JJ, Smith GW, Scheetz D, et al. Does size matter in females? An overview of the impact of the high variation in the ovarian reserve on ovarian function and fertility, utility of anti-müllerian hormone as a diagnostic marker for fertility and causes of variation in the ovarian reserve in cattle. Reprod Fertil Dev. 2011;23:1–14.
5. Kevenaar ME, Meerasahib MF, Kramer P, et al. Serum anti-müllerian hormone levels reflect the size of the primordial follicle pool in mice. Endocrinology. 2006;147(7):3228–3234.
6. Appt SE, Clarkson TB, Chen H, et al. Serum anti-müllerian hormone predicts ovarian reserve in a monkey model. Menopause. 2009;16:597–601.
7. Claes A, Ball BA, Almeida J, Corbin CJ, Conley AJ. Serum anti-müllerian hormone concentration in stallions: developmental changes, seasonal variation, and differences between intact stallions, cryptorchid stallions, and geldings. Theriogenology. 2013;79:1229–1235.
8. Robeck TR, Monfort ST, Calle PP, et al. Reproduction, growth and development in captive beluga (Delphinapterus leucas). Zoo Biol. 2005;24:29–49.