Abstract: Adult male elephants periodically show the phenomenon of musth, a condition associated with increased aggressiveness, restlessness, significant weight reduction and markedly elevated androgen levels. It has been suggested that musth-related behaviours are costly and that therefore musth may represent a form of physiological stress. In order to provide data on this largely unanswered question, the first aim of this study was to evaluate different assays for non-invasive assessment of adrenocortical function in the male African elephant by (i) characterizing the metabolism and excretion of [3H]cortisol (3H-C) and [14C]testosterone (14C-T) and (ii) using this information to evaluate the specificity of four antibodies for determination of excreted cortisol metabolites, particularly with respect to possible cross-reactions with androgen metabolites, and to assess their biological validity using an ACTH challenge test. Based on the methodology established, the second objective was to provide data on fecal cortisol metabolite concentrations in bulls during the musth and non-musth condition. 3H-C (1 mCi) and 14C-T (100 microCi) were injected simultaneously into a 16 year old male and all urine and feces collected for 30 and 86 h, respectively. The majority (82%) of cortisol metabolites was excreted into the urine, whereas testosterone metabolites were mainly (57%) excreted into the feces. Almost all radioactive metabolites recovered from urine were conjugated (86% 3H-C and 97% 14C-T). In contrast, 86% and >99% of the 3H-C and 14C-T metabolites recovered from feces consisted of unconjugated forms. HPLC separations indicated the presence of various metabolites of cortisol in both urine and feces, with cortisol being abundant in hydrolysed urine, but virtually absent in feces. Although all antibodies measured substantial amounts of immunoreactivity after HPLC separation of peak radioactive samples and detected an increase in glucocorticoid output following the ACTH challenge, only two (in feces against 3alpha,11-oxo-cortisol metabolites, measured by an 11-oxo-etiocholanolone-EIA and in urine against cortisol, measured by a cortisol-EIA) did not show substantial cross-reactivity with excreted 14C-T metabolites and could provide an acceptable degree of specificity for reliable assessment of glucocorticoid output from urine and feces. Based on these findings, concentrations of immunoreactive 3alpha,11-oxo-cortisol metabolites were determined in weekly fecal samples collected from four adult bulls over periods of 11-20 months to examine whether musth is associated with increased adrenal activity. Results showed that in each male levels of these cortisol metabolites were not elevated during periods of musth, suggesting that in the African elephant musth is generally not associated with marked elevations in glucocorticoid output. Given the complex nature of musth and the variety of factors that are likely to influence its manifestation, it is clear, however, that further studies, particularly on free-ranging animals, are needed before a possible relationship between musth and adrenal function can be resolved. This study also clearly illustrates the potential problems associated with cross-reacting metabolites of gonadal steroids in EIAs measuring glucocorticoid metabolites. This has to be taken into account when selecting assays and interpreting results of glucocorticoid metabolite analysis, not only for studies in the elephant but also in other species.

Abstract: Jugular vein blood samples were collected from 23 young and sexual mature feral stallions to examine the relationship between plasma testosterone concentration and age, breeding season or harem size. Testosterone concentration increased with the age of the stallions until they formed their own harems, at about 4 to 6 years old. Seasonal variations in testosterone concentrations were observed, and found to be significantly higher (P<0.001) throughout the breeding season than non-breeding season, from 3 years of age. Testosterone levels were correlated with harem size for individual stallions. It can be inferred from these results that there is a relationship between plasma testosterone concentration and age, breeding season and harem size.

Abstract: Clinical investigators have been forced primarily to use experiments of nature (e.g., cloacal exstrophy; androgen insensitivity, congenital adrenal hyperplasia) to assess the contribution of fetal sex hormone exposure to the development of male- and female-typical profiles of gender identity and role behavior as well as sexual orientation. In this review, I summarize the results of numerous correlative as well as mechanistic animal experiments that shed significant light on general neuroendocrine mechanisms controlling the differentiation of neural circuits controlling sexual partner preference (sexual orientation) in mammalian species including man. I also argue, however, that results of animal studies can, at best, provide only indirect insights into the neuroendocrine determinants of human gender identity and role behaviors.

Abstract: Summary The aim of this study was to investigate the effect of gonadotrophin-releasing hormone (GnRH) immunisation on mature stallions that had been used for breeding. Four Standardbred stallions were used in the study: 3 experimental animals and 1 control animal. Semen was collected regularly, i.e. twice/week, during the 4 months prior to the experimental period. The stallions were immunised against GnRH with a GnRH-BSA conjugate. Equimune was used as the adjuvant. The stallions were immunised on 5 occasions, 4 at 2 week intervals, and the fifth 4 weeks after the fourth. Blood samples were taken once a week for analysis of GnRH antibody titre and every third week for testosterone and oestrone sulphate analyses. Semen was collected once a week, and libido and sexual behaviour were observed. Ejaculate volume, sperm concentration, total number of sperm in the ejaculate, sperm motility and sperm morphology were evaluated. Testicular size was measured once a week. At the end of the study, the stallions were castrated, and a histological examination of the testes performed. All immunised stallions produced antibodies against GnRH, and plasma testosterone concentration decreased. However, the effect of immunisation varied between stallions. In 2 of the stallions, high levels of antibodies were found, while in the third, the level was moderate. Four weeks after the first immunisation, a decrease in libido was observed. Two months after the first immunisation, marked changes in semen quality were observed in the 2 stallions with high antibody titres. Fourteen weeks after the first immunisation, the total number of sperm/ejaculate had decreased from >8.6×109 to<2.7×109, sperm motility from >59 to<10% and the frequency of morphological normal spermatozoa had decreased from >60 to<14%. The dominating abnormalities were abnormal head shapes, proximal cytoplasmic droplets and detached heads. In the third stallion, only slight changes in semen quality were found. No changes were observed in the control stallion. Decreases in testicular size were noted in all of the experimental stallions. Pronounced histological alterations in the testes were observed in 2 of the stallions. It is concluded that the vaccine was effective in stimulating production of GnRH antibodies and in suppressing testicular function and androgen secretion. However, there was an individual variation in the responses among the stallions and, further, libido was not totally suppressed.