Fecal androgen metabolite analysis for noninvasive monitoring of testicular steroidogenic activity in felids

Limited data are available on long-term, seasonal changes in testicular steroidogenic activity in nondomestic felids, primarily because of the difficulties associated with longitudinal blood sampling (e.g., handling, restraint, anesthesia). Therefore, a noninvasive approach for assessing testicular androgen production was developed using the domestic cat (Felis catus) as a model. Two adult males were injected i.m. with 4 μCi¹⁴-testosterone to determine the time course and relative proportions of androgen metabolites excreted in urine and feces. Peak urinary radioactivity was detected 13 and 19 hr postinjection and accounted for ∼8% of the total radioactivity recovered. High performance liquid chromatography (HPLC) analysis detected multiple polar urinary metabolites, none of which eluted with the ³H-testosterone reference tracer. The majority of urinary testosterone metabolites consisted of nonenzyme-hydrolyzable, water-soluble (presumably conjugated) forms. In feces, radioactivity was detected in the first sample collected at 22 hr postinjection for both males, although peak metabolite excretion in one male was not observed until 61 hr postinjection. HPLC analysis detected several fecal metabolites consisting primarily of nonhydrolyzable, water-soluble forms (84.4 ± 0.9%) with some ether-soluble forms (15.6 ± 0.9%). None of the fecal androgen metabolites were associated with free testosterone. However, one or more of the water-soluble fecal metabolites was quantifiable using a commercially available testosterone radioimmunoassay. The biological relevance of this immunoactivity was confirmed in the domestic cat; concentrations were high in adult, intact males and nondetectable in intact females and castrated males and females. In addition, fecal androgen concentrations in a male Pallas' cat (Felis manul) exhibited seasonal fluctuations that corresponded with parallel changes in serum testosterone and ejaculate quality. These data indicate that testicular steroidogenic activity can be monitored non invasively in felids, providing a potentially valuable tool for endangered felid management to: (1) assess pubertal status, (2) determine the influence of season on reproduction, and (3) diagnose possible causes of sub- or infertility. © 1996 Wiley-Liss, Inc.     

Annual variation in plasma cortisol levels in common marmosets, Callithrix jacchus

In contrast to numerous studies showing breeding seasonality for animals living in temperate zones, few have addressed the effect of annual environmental changes on hormones and behavior related to reproduction for animals living in tropical and equatorial regions. In order to expand information on other primates living near the equator, we used 13 adult common marmosets, eight males and five females living under natural climatic conditions in a colony located in North-Eastern Brazil. Blood samples were collected monthly from July 2002 to June 2004. Cortisol assays were performed by ELISA. Cortisol levels varied according to environmental temperature and rainfall but were not significantly affected by photoperiod. After separating out these environmental effects through an analysis of covariance for repeated measurements, a marginal significant difference in cortisol levels according to the seasons of the year was detected. A significant difference in cortisol levels between females and males was also observed. The results suggest that annual variation in cortisol in common marmosets is influenced by temperature and rainfall and might be associated with the mating season.     

Effects of zeranol upon bovine testicular function

Effects of zeranol upon testicular function in bulls were studied. In Experiment I, young bulls (104+/-1.7 days of age) were either implanted with 36 mg zeranol or served as controls with reimplantation on days 56 and 112 after first implant. After a 168-day treatment period, half the bulls were orchidectomized with the remaining bulls assigned to a 168-day recovery period, after which they were also orchidectomized. Zeranol inhibited testicular development during the treatment period. Testosterone (T) concentrations were lower (P<0.01) in treated bulls. Spermatozoa were absent in testes of treated bulls while control bulls averaged 27x10(6) sperm per gram of testis. At the end of the 168-day recovery period, testes weights and sperm concentrations were similar (P>0.10) between bulls previously treated with zeranol and controls. Implantation of older bulls with either 36 mg (267+/-3.4 days of age: Experiment II) or 72 mg (16 months of age: Experiment III) of zeranol had no effect (P>0.10) upon spermatogenesis or T production over a 168-day treatment period. The results indicate that the inhibitory effects of zeranol upon bovine testicular function are age-dependent and suggest that there may be an increased sensitivity of the hypothalmic-pituitary-testicular axis to zeranol in the prepuberal bull.     

Energetic reserves, leptin and testosterone: a refinement of the immunocompetence handicap hypothesis

The immunocompetence handicap hypothesis (ICHH) assumes that testosterone (T), required for the expression of sexual traits, can also incur a cost due to its immunosuppressive properties. However, T-dependent immunosuppression could also arise as an indirect consequence of energy reallocation from the immune system to other metabolic demands. Leptin is mostly produced in lipogenic tissues and its circulating level is positively correlated with the amount of lipid reserves. Leptin also has an important role as immunoenhancer and we suggest that this hormone could play a role as a mediator of the immunosuppressive effect of testosterone. In particular, we propose that only the individuals able to maintain large lipid reserves (with high leptin levels), while sustaining high testosterone levels, might be able to develop sexual displays without an impairment of their immune defences. Here, we tested one of the assumptions underlying this extension of the ICHH: leptin administration should attenuate testosterone-induced immunosuppression. T-implanted and control male zebra finches (Taeniopygia guttata) received daily injections of leptin or phosphate buffered saline. T-implants initially depressed the phytohaemagglutinin-induced immune response. However, T-birds injected with leptin enhanced their immune response to the level of control birds. These results open a new perspective on the study of the ICHH.     

Seasonal changes in serum dehydroepiandrosterone,androstenedione, and testosterone levels in the squirrel monkey (Saimiri boliviensis boliviensis)

The squirrel monkey (Saimiri boliviensis boliviensis) has a well-defined breeding season during which adult males undergo androgen-dependent morphological changes, with acquisition of active spermatogenesis. To assess the hormonal events of this annual cycle, blood samples were obtained weekly from ten adult males, and serum was assayed for testosterone (T), androstenedione (ΔA), and dehydroepiandrosterone (DHEA). A significant seasonal variation was noted in mean serum T (P < 0.02), ΔA (P < 0.02), and DHEA (P < 0.001) concentrations. Mean ΔA concentrations increased from a nonbreeding season nadir of 91.4 ± 12.9 ng/ml (mean ± standard error) to a prebreeding concentration of 139 ± 10.5 ng/ml and breeding season peak of 167.5 ± 15.4 ng/ml (P < 0.05). Mean DHEA concentrations increased from a nonbreeding season nadir of 8.3 ± 0.8 to a breeding season peak of 14.3 ± 1.2 (P < 0.001). Mean T levels in the nonbreeding (52.2 ± 11.6 ng/ ml) and prebreeding season (48.6 ± 7.4) were similar. However, T significantly increased during the breeding season to 103.5 ± 12.8 ng/ml (P < 0.05). Progressive changes in body weight and morphology paralleled the rise in serum ΔA levels. The pattern of peripheral serum androgen concentrations throughout the year would suggest annual activation of the hypothalamic-pituitary-adrenal and/or hypothalamic-pituitary-gonadal axes.     

A review of hormonal factors influencing the sexual and aggressive behavior of macaques

The effects of gonadal hormones on the sexual and aggressive behavior of adult macaques are reviewed. Similarities among findings from field, colony, and laboratory studies strengthen the view that testosterone facilitates the sexual and aggressive behavior of males, while sexual and perhaps aggressive behavior by the female is mainly dependent on estradiol, which increases both the sexual motivation of the female and her attractiveness to males. Differences between results from different settings help to emphasize the role of environmental and social factors in modulating the effects of hormones. © 1993 Wiley-Liss, Inc.     

Biochemical and hormonal correlates of dominance and social behavior in all-male groups of squirrel monkeys (Saimiri sciureus)

This study examined the relationship between dominance rank and several physiological and behavioral measures in stable, captive, all-male squirrel monkey groups. Four groups, each containing three males, were observed for 12 weeks during the breeding season. Relative dominance ranking among males in each group was based on the direction of agonistic behaviors displayed. For each subject, whole blood serotonin (WBS), plasma testosterone (T), and cortisol (C) were sampled 4 to 6 times over the course of the study. Samples were separated by 1- to 2-week intervals. Each group had a stable linear dominance hierarchy. WBS had small intraanimal variance and was positively related to dominance rank. In contrast, T and C were highly variable within subjects and were unrelated to dominance rank. Among alpha males, concentrations of T and C were positively correlated, and WBS and C were negatively correlated. The effect of dominance rank on the relationship between within-animal fluctuations in WBS, T, and C and behavior was also assessed. In dominant, but not subordinate subjects, intraanimal fluctuations in WBS correlated with agonistic behavior initiated, and fluctuations in C and T correlated with huddling. In beta and gamma males, C was related to agonism received, and in gamma males to food stealing received. Dominance status also affected endocrine response to the stress of capture but not the rate of sneezing. Sneezing was positively correlated with T concentrations irrespective of dominance rank. These results extend the association between WBS and dominance rank previously reported in Old World monkeys to a New World monkey species, support previous suggestions that mean T and C titers are not reflective of dominance rank in well-established groups, and indicate that dominance rank affects adrenocortical response to the stress associated with capture and anesthesia.     

Individual and seasonal variation in fecal testosterone and cortisol levels of wild male tufted capuchin monkeys, Cebus apella nigritus

This study tested the "challenge hypothesis" and rank-based predictions for temporal steroid production in male tufted capuchin monkeys, Cebus apella. Fecal samples (n = 209) collected from six wild males were analyzed for testosterone and cortisol concentration by enzyme immunoassay. The temporal pattern in male steroid production was compared to female sexual activity and rates of male aggression. The top-ranking adult male did not differ from other adult males in testosterone or cortisol concentration. Mean adult testosterone was significantly higher than mean subadult testosterone throughout the year. There was a clear elevation of testosterone and cortisol in both adult and subadult males during the peak of adult female sexual activity after the birth season. In fact, the magnitude of increase in testosterone was higher than predicted for a species with low male-male aggression. However, there was no difference between nonbreeding baseline testosterone levels during the birth season, and the "breeding" baseline of testosterone in males found during asynchronous female sexual activity. Of all behavioral indices examined, the distribution of female-maintained consortships was the best predictor of mean adult male testosterone concentrations. Although in many species, elevated testosterone coincides with increased male-male aggression, in the present study, the sustained high-magnitude increase in steroids during the peak of adult female sexual activity was associated with a relatively low rate of male-male intragroup aggression.     

Reproductive synchrony in captive macaques

Captive bonnet macaques housed at the California Primate Research Center reproduce seasonally. The chances of producing surviving infants were substantially reduced among females who conceived at the peak of the mating season compared with females whose conceptions were more isolated in time. Primiparous females and low ranking females were most consistently affected by the extent of reproductive synchrony. Behavioral data suggest that harassment of conceiving and pregnant females may have contributed to the correlation between the extent of reproductive synchrony and infant mortality as the levels of aggression toward females were highest during the months in which conceptions were most common.     

Androgen control of vocal control region volumes in a wild migratory songbird (Junco hyemalis) is region and possibly age dependent

Previous laboratory studies have shown that photoperiodic adult songbirds experience seasonal variations in singing frequency that correlate with plasma androgen levels, as well as changes in the brain regions that control singing (vocal control regions). The present study investigates naturally occurring seasonal changes in the sizes of these regions in a wild migratory species (dark-eyed junco, Junco hyemalis), with samples from adolescence to post-breeding fall migration. In adult males, the volumes of the vocal control regions area X and the higher vocal center (HVC) were large during the breeding season when birds were singing and androgen levels were high, and decreased in size after the breeding season when singing had stopped and androgen levels were low. HVC volume in adolescent males caught in the fall (no singing), when plasma androgen levels were low, was smaller than in breeding adults, thereby following the seasonal pattern of change in plasma androgen levels. In adolescent males, however, area X volume was the same as in breeding adults. Thus, area X size in adolescent male juncos may be testosterone independent. The seasonal pattern of robust nucleus of the archistriatum volume was similar to that of the HVC. The volumes of neither the magnocellular nucleus of the anterior neostriatum nor the nucleus rotundus, a control region, differed seasonally. Castration of breeding adult males caused both area X and HVC volumes to decrease compared to castrated controls with testosterone replacement, indicating that maintenance of these two region volumes is testosterone dependent in adults. © 1997 John Wiley & Sons, Inc. J Neurobiol 32: 391–402, 1997.