Phthalates and sex steroid hormones across the perimenopausal period: A longitudinal analysis of the Midlife Women's Health Study.

BACKGROUND: The menopausal transition involves significant sex hormone changes. Environmental chemicals, such as urinary phthalate metabolites, are associated with sex hormone levels in cross-sectional studies. Few studies have assessed longitudinal associations between urinary phthalate metabolite concentrations and sex hormone levels during menopausal transition. METHODS: Pre- and perimenopausal women from the Midlife Women's Health Study (MWHS) (n = 751) contributed data at up to 4 annual study visits. We quantified 9 individual urinary phthalate metabolites and 5 summary measures (e.g., phthalates in plastics (∑Plastic)), using pooled annual urine samples. We measured serum estradiol, testosterone, and progesterone collected at each study visit, unrelated to menstrual cycling. Linear mixed-effects models and hierarchical Bayesian kernel machine regression analyses evaluated adjusted associations between individual and phthalate mixtures with sex steroid hormones longitudinally. RESULTS: We observed associations between increased concentrations of certain phthalate metabolites and lower testosterone and higher sub-ovulatory progesterone levels, e.g., doubling of monoethyl phthalate (MEP), monobenzyl phthalate (MBzP), di-2-ethylhexyl phthalate (∑DEHP) metabolites, ∑Plastic, and ∑Phthalates concentrations were associated with lower testosterone (e.g., for ∑DEHP: -4.51%; 95% CI: -6.72%, -2.26%). For each doubling of MEP, certain DEHP metabolites, and summary measures, we observed higher mean sub-ovulatory progesterone (e.g., ∑AA (metabolites with anti-androgenic activity): 6.88%; 95% CI: 1.94%, 12.1%). Higher levels of the overall time-varying phthalate mixture were associated with lower estradiol and higher progesterone levels, especially for 2nd year exposures. CONCLUSIONS: Phthalates were longitudinally associated with sex hormone levels during the menopausal transition. Future research should assess such associations and potential health impacts during this understudied period.

Using Quantitative Hormonal Fertility Monitors to Evaluate the Luteal Phase: Proof of Concept Case Study.

Several new quantitative fertility monitors are now available for at-home use that measure estrogen, luteinizing hormone (LH), and progesterone (PDG) in urine. This case report compares the Mira and Inito quantitative fertility monitors with the well-established qualitative ClearBlue fertility monitor. Three clinical scenarios were evaluated: a normal cycle, a prolonged luteinization cycle, and an anovulatory cycle. The identification of the luteal phase (or lack thereof in the case of anovulation) and the transition through the three processes of luteinization, progestation, and luteolysis were clearly demarcated with the help of quantitative LH and PDG. Quantitative fertility monitors have the potential to identify details of the luteal phase to help women with regular cycles and abnormal luteal phases to help target interventions for optimizing fertility.

CYP3A7*1C allele: linking premenopausal oestrone and progesterone levels with risk of hormone receptor-positive breast cancers.

BACKGROUND: Epidemiological studies provide strong evidence for a role of endogenous sex hormones in the aetiology of breast cancer. The aim of this analysis was to identify genetic variants that are associated with urinary sex-hormone levels and breast cancer risk. METHODS: We carried out a genome-wide association study of urinary oestrone-3-glucuronide and pregnanediol-3-glucuronide levels in 560 premenopausal women, with additional analysis of progesterone levels in 298 premenopausal women. To test for the association with breast cancer risk, we carried out follow-up genotyping in 90,916 cases and 89,893 controls from the Breast Cancer Association Consortium. All women were of European ancestry. RESULTS: For pregnanediol-3-glucuronide, there were no genome-wide significant associations; for oestrone-3-glucuronide, we identified a single peak mapping to the CYP3A locus, annotated by rs45446698. The minor rs45446698-C allele was associated with lower oestrone-3-glucuronide (-49.2%, 95% CI -56.1% to -41.1%, P = 3.1 × 10-18); in follow-up analyses, rs45446698-C was also associated with lower progesterone (-26.7%, 95% CI -39.4% to -11.6%, P = 0.001) and reduced risk of oestrogen and progesterone receptor-positive breast cancer (OR = 0.86, 95% CI 0.82-0.91, P = 6.9 × 10-8). CONCLUSIONS: The CYP3A7*1C allele is associated with reduced risk of hormone receptor-positive breast cancer possibly mediated via an effect on the metabolism of endogenous sex hormones in premenopausal women.

Reproductive hormone measurement from minimally invasive sample types: Methodological considerations and anthropological importance.

Energetic investment in human reproduction has long been recognized as costly, influencing developmental, physiological, and behavioral patterns in males and females. These effects are largely coordinated through the actions of reproductive hormones (eg, testosterone, estradiol, and progesterone). Here, the utility and limitations of minimally invasive sampling techniques are explored, providing a novel perspective on how reproductive hormone measurements can enhance reproductive endocrinology research. Salivary steroid measures are most commonly used, although several dried blood spot and urine assays are also available, and researchers continue to explore the efficacy of other sample types. These relatively simple measures have facilitated the collection of multiple samples from a single participant, allowing researchers to more accurately track the diurnal and cyclical variation exhibited by many reproductive hormones. Ultimately, the ability to collect fine-grained participant data allows biological anthropologists to better test questions central to human reproductive ecology, life history theory, and public health. For example, fieldwork using these techniques suggests that testosterone profile variation across populations is influenced by energetic constraints and reproductive status. Moreover, hormone concentrations shape the development of sex characteristics, with implications for evolutionary questions related to sexual selection. Hormone levels also can be used to identify a range of medical concerns (eg, suppressed hormone production levels linked with psychosocial stress). These findings highlight how minimally invasive collection techniques can be applied to test diverse evolutionary hypotheses and identify important health concerns. Still, more work is needed to standardize collection and laboratory analysis procedures, thereby enabling more direct data comparisons between researchers.

How reproductive hormonal changes affect relationship dynamics for women and men: A 15-day diary study.

Research suggests that women's sexual psychology and behavior change across the ovulatory cycle, but very little is known about how fluctuations in estradiol and progesterone - two hormones that systematically vary across the ovulatory cycle - affect romantic relationship dynamics. We present the first dyadic study to assess daily hormonal fluctuations and personal and relationship well-being from both partners' perspectives. Specifically, we recruited women who were not using hormonal contraception and their partners for a 15-day diary study. Participants collected daily urine samples to assess estradiol, progesterone, and testosterone, and they responded to daily questions about their relationship. Results revealed that increases in estradiol negatively affected women's relationship evaluations. Men perceived these changes, which in turn, affected men's well-being. The present findings highlight the importance of women's hormonal fluctuations in shaping relationship dynamics and provide, for the first time, information about how such fluctuations affect male partners.

Daily luteal serum and urinary hormone profiles in the menopause transition: Study of Women's Health Across the Nation.

OBJECTIVE: To further characterize the endocrinology of the menopause transition, we sought to determine: whether relationships between urine and serum hormones are maintained as women enter their sixth decade; whether a single luteal phase serum progesterone (P) is reflective of integrated-luteal urinary pregnanediol glucuronide (uPdg); and whether serum P, like luteal uPdg, declines as women approach their final menses (FMP). METHODS: The Study of Women's Health Across the Nation (SWAN) Daily Hormone Study's (DHS) is a community-based observational study. A subset of participants underwent a timed, luteal blood draw planned for cycle days 16 to 24 during the same month of DHS collection. Serum-luteinizing hormone (LH), follicle-stimulating hormone (FSH), estradiol and P, and urine LH, FSH, estrone conjugates (E1c), and daily and integrated luteal uPdg were measured in 268 samples from 170 women. Serum/urine hormone associations were determined using Pearson's correlation and linear regression, adjusted for concurrent age, body mass index, smoking status, and race/ethnicity. RESULTS: Pearson's r ranged from 0.573 (for LH) to 0.843 (for FSH) for serum/urine correlations. Integrated luteal uPdg weakly correlated with serum P (Pearson's r = 0.26, P = 0.004) and explained 7% of the variability in serum P in adjusted linear regression (total R 0.09, P = 0.002). Serum P demonstrated a marginally significant decline with approaching FMP in adjusted analysis (P = 0.04). CONCLUSIONS: Urine and serum hormones maintain a close relationship in women into their sixth decade of life. Serum luteal P was weakly reflective of luteal Pdg excretion.

A multichannel system integrating molecularly imprinted conductive polymers for ultrasensitive voltammetric determination of four steroid hormones in urine.

This work reports on a modularized electrochemical method for the determination of the hormones cortisol, progesterone, testosterone and 17ß-estradiol in urine. These hormones were employed as templates when generating molecular imprints from aniline and metanilic acid by electropolymerization on the surface of screen-printed electrodes. The electrically conductive imprint was characterized by SEM, AFM and cyclic voltammetry. A four-channel system was then established to enable simultaneous determination of the hormones by cyclic voltammetry. The detection limits for cortisol, progesterone, testosterone and 17ß-estradiol are as low as 2, 2.5, 10 and 9 ag·mL-1 (for S/N = 3). Graphical abstract A four-channel system was established to enable simultaneous determination of 4 steroid hormones by cyclic voltammetry and by using moleculalry imprinted polymers.

In vivo study on biotransformation of pentacyclic analogs of progesterone: Identification of their metabolites by HPLC-MS method.

Progesterone derivatives containing the D' additional cyclohexane ring in the 16α,17α-positions of steroid core (pregna-D'-pentaranes) exhibited high in vitro and in vivo selective progestogenic activity. The assessment of their biotransformation in the body, and the identification of possible metabolites are integral parts of a potential drug studies. Here we describe the results of in vivo metabolic transformation of 6α-methyl-16α,17α-cyclohexanopregn-4-ene-3,20-dione 1 and its 6-demethylated analog 2 and identification of their metabolites in rat urine. We synthesized and fully characterized (1D and 2D NMR, HRMS) 11 possible metabolites as the standards. Then we developed the LC-MS/MS assay including sample preparation and chromatography conditions for identification of the detected metabolites of 1 and 2. The 5α- and 5ß-3,20-diketo-, 3ß-hydroxy-20-keto-5α-, 3-keto-20(S)-hydroxy-5α-, 3ß,20(S)-dihydroxy-5α-metabolites of compounds 1 and 2 were found in rat urine samples. The starting steroids 1 and 2, as well as both 3ß,20(R)-dihydroxy metabolites were not detected in the examined biological urine samples. Thus, we demonstrate for the first time that exogenous progestines - pregna-D'-pentaranes - and endogenous progesterone follow similar metabolic pathways. Therefore, despite the presence of an additional ring D' and the methyl group in position 6, the main enzymatic transformations are similar to those of the natural hormone.

How high-resolution techniques enable reliable steroid identification and quantification.

Due to possible matrix interferences and artefact generation during sample preparation, careful method validation is required for quantitative bioanalytical methods, especially for analytes that are only present in low concentrations. Using the identification and quantification of progesterone metabolites in the urine of newborns as an example, we show how modern high-resolution instruments can be used to verify analyte assignment and avoid pitfalls commonly encountered by the use of low-resolution instruments.

Variable selection in rank regression for analyzing longitudinal data.

In this paper, we consider variable selection in rank regression models for longitudinal data. To obtain both robustness and effective selection of important covariates, we propose incorporating shrinkage by adaptive lasso or SCAD in the Wilcoxon dispersion function and establishing the oracle properties of the new method. The new method can be conveniently implemented with the statistical software R. The performance of the proposed method is demonstrated via simulation studies. Finally, two datasets are analyzed for illustration. Some interesting findings are reported and discussed.

Urinary specific gravity as an alternative for the normalisation of endocrine metabolite concentrations in giant panda (Ailuropoda melanoleuca) reproductive monitoring.

Reproductive monitoring for captive breeding in giant pandas is based on behavioural observation and non-invasive hormone analysis. In urine, interpretation of results requires normalisation due to an animal's changing hydration. Correction of urinary concentrations based on creatinine is the gold standard. In this study, a largely unexplored, easy-to-perform normalisation technique, based on urinary specific gravity (USpG), was examined and compared to creatinine. To this extent, six cycles from two female pandas (SB741(1) and SB569(5)) were monitored through urine analysis for oestrogen, progesterone, ceruloplasmin and 13,14-dihydro-15-keto-PGF2a (PGFM). The Pearson's correlation between creatinine and USpG was high (r = 0.805-0.894; p < 0.01), indicative for a similar performance of both normalisation methods. However, generally lower values were observed during pro-oestrus and primary (progesterone) rise. This could be associated with huge shifts in appetite, monitored by faecal output (kg) with an averaged > 50% decrease during oestrus and >50% increase during primary progesterone rise. In parallel, respectively highest and lowest creatinine and USpG levels, were measured, with creatinine obviously more affected as a result of linkage with muscle tissue metabolism affected by reproductive hormones. As a consequence, metabolite levels were significantly different between both corrected datasets with significantly higher oestrogen peak levels during oestrus ranging from 2.13-86.93 and 31.61-306.45 ng/mL (USpG correction) versus 2.33-31.20 and 36.36-249.05 ng/mL Cr (creatinine correction) for SB569 and SB741 respectively, and significant lower progesterone levels during primary progesterone rise ranging from 0.35-3.21 and 0.85-6.80 ng/mL (USpG correction) versus 0.52-10.31 and 2.10-272.74 ng/mL Cr (creatinine correction) for SB569 and SB741 respectively. Consequently, USpG correction rendered unbiased profiles, less subject to variation and metabolic artefacts and therefore allowed a more straightforward identification of peak oestrogen and onset of secondary progesterone rise, being potentially advantageous for future studies unravelling key giant panda reproductive events, including (delayed) implantation. The alternative application of USpG as a normalisation factor was further supported by its easy application and environmental and technical robustness.

Reproductive biology of captive southern hairy-nosed wombats (Lasiorhinus latifrons). Part 1: oestrous cycle characterisation.

Southern hairy-nosed wombats (SHNWs: Lasiorhinus latifrons) do not breed well in captivity. To better understand their reproduction, daily urine samples were collected from nine captive females and analysed for volume (mL), specific gravity and a qualitative index of the number of epithelial cells, then stored at -20°C until samples could be analysed for progesterone metabolites (P4M). The mean oestrous cycle length was 35.1±2.4 days; however, individual cycle length ranged from 23 to 47 days. The mean luteal phase length was 20.8±1.3 days (range: 12 to 33 days). Urinary P4M was divided into four oestrous cycle stages: (1) early follicular phase, (2) late follicular phase, (3) early luteal phase, (4) late luteal phase, and analysed against urinary characteristics. During the late follicular phase, urine volume decreased (P=0.002) while urine specific gravity (P=0.001) and concentration of epithelial cells (P=0.004) both increased. The level of variability in oestrous cycle length suggests that some captive females may exhibit abnormal cycles; however, the changes in the urinary characteristics associated with the different stages of the oestrous cycle appear to offer a possible non-invasive means of monitoring the reproductive status of captive SHNWs.

Reproductive biology of captive female southern hairy-nosed wombats (Lasiorhinus latifrons). Part 2: oestrous behaviour.

The poor captive breeding success of southern hairy-nosed wombats (SHNWs; Lasiorhinus latifrons) has been attributed to the difficulty in accurately characterising oestrous behaviour and their relationship to circulating reproductive hormones. Over two wombat breeding seasons, the use of infrared cameras for 24-h remote behavioural monitoring and the analysis of urine samples collected from seven captive females, were investigated to determine the relationship between behaviour and changes in urinary progesterone metabolites (P4M). Urinary P4M was divided into two concentrations: (1) ≤ baseline P4M values and (2)>baseline P4M values and evaluated against urine volume (mL) and the duration (s) and frequency of 23 behaviours recorded for 8 days surrounding D0 of the luteal phase (D0: a sustained increase in P4M for three or more consecutive days). When P4M was ≤ baseline, the duration of urination and volume both decreased, whereas the duration and frequency of both pacing and rump bites by the female towards the male increased. These results suggest that there were detectable behavioural changes that can be mapped to the changes in the SHNW oestrous cycle, which may be used as behavioural indicators to identify the reproductive status of females.

Interrelationship among annual cycles of sex steroids, corticosterone and body condition in Nyctibatrachus humayuni.

Synergism between extrinsic and intrinsic factors is crucial for the seasonality of reproduction. Environmental factors such as photoperiod and temperature activate the hypothalamus-pituitary-gonadal axis leading to the secretion of steroid hormones that are crucial for reproduction. Sex steroids are not only essential for the maturation of gonads, but also for development of secondary sexual characters in males and reproductive behaviour of both the sexes. In the present study, we quantified the urinary testosterone (UTM) and corticosterone (UCM) metabolites in males and urinary estradiol metabolites (UEM) and UCM in females of Nyctibatrachus humayuni for two consecutive years to determine annual and seasonal variation in the levels of sex steroids, corticosterone and body condition index (BCI). The results show that sex steroids were highest during the breeding season and lowest during the non-breeding season in both the sexes. An increase in UTM and UEM was observed in males and females respectively during the breeding season. Testicular histology showed the presence of all stages of spermatogenesis throughout the year indicating that spermatogenesis is potentially continuous. Ovarian histology showed the presence of vitellogenic follicles only during the breeding season indicating that oogenesis is strictly seasonal. In males, UCM levels were highest during the breeding season, while in females their levels were highest just prior to the breeding season. In males, BCI was highest during the pre-breeding season, declined during the breeding season to increase again during the post-breeding season. In females, BCI was comparable throughout the year. In males, UTM levels were positively correlated with UCM levels but negatively correlated with BCI. Interestingly, UEM, UCM and BCI were not correlated in females. These results indicate that N. humayuni exhibits an associated pattern of reproduction. Quantification of urinary progesterone metabolites (UPM) during the breeding season showed UPM levels were higher in post-spawning females, suggesting the significance of progesterone in ovulation. Further, non-invasive enzyme immunoassay has been successfully standardized in N. humayuni for the quantification of urinary metabolites of steroid hormones.

FPCA-based method to select optimal sampling schedules that capture between-subject variability in longitudinal studies.

A critical component of longitudinal study design involves determining the sampling schedule. Criteria for optimal design often focus on accurate estimation of the mean profile, although capturing the between-subject variance of the longitudinal process is also important since variance patterns may be associated with covariates of interest or predict future outcomes. Existing design approaches have limited applicability when one wishes to optimize sampling schedules to capture between-individual variability. We propose an approach to derive optimal sampling schedules based on functional principal component analysis (FPCA), which separately characterizes the mean and the variability of longitudinal profiles and leads to a parsimonious representation of the temporal pattern of the variability. Simulation studies show that the new design approach performs equally well compared to an existing approach based on parametric mixed model (PMM) when a PMM is adequate for the data, and outperforms the PMM-based approach otherwise. We use the methods to design studies aiming to characterize daily salivary cortisol profiles and identify the optimal days within the menstrual cycle when urinary progesterone should be measured.

Ovarian hormones, age and pubertal development and their association with days of headache onset in girls with migraine: An observational cohort study.

Background Fifty-three percent of adolescent girls report headaches at the onset of menses, suggesting fluctuations of ovarian hormones trigger migraine during puberty. Aims To determine if urinary metabolites of estrogen and progesterone are associated with days of headache onset (HO) or severity in girls with migraine. Methods This was a pilot study and included 34 girls with migraine balanced across three age strata (pre-pubertal (8-11), pubertal (12-15), and post-pubertal (16-17) years of age). They collected daily urine samples and recorded the occurrence and severity of headache in a daily diary. Urine samples were assayed for estrone glucuronide (E1G) and pregnandiol glucuronide (PdG) and the daily change was calculated (ΔE1G, ΔPdG). Pubertal development was assessed by age, pubertal development score (PDS), and menstrual cycle variance. The primary outcome measures were HO days and headache severity. Generalized linear mixed models were used, and included the hormonal variables and three different representations of pubertal development as covariates. Results Models of HO days demonstrate a significant age*PdG interaction (OR 0.85 [95% CI 0.75, 0.97]) for a 1 standard deviation increase in PdG and three-year increase in age. A separate model showed a significant PDS*PdG interaction (OR -0.85 [95% CI; 0.76, 0.95]). ΔPDG was associated with headache severity in unadjusted models ( p < 0.017). Conclusion Age and pubertal development could moderate the effect of ovarian hormones on days of headache onset in girls with migraine.

Diagnostic relevance of urinary steroid profiles on ovarian granulosa cell tumors: two case reports.

BACKGROUND: Granulosa cell tumor of the ovary is the most frequent sex cord stromal tumor and represents 2 to 5% of all primary ovarian cancers. Ovarian granulosa cell tumor is a malignant tumor with slow progression and in some cases this tumor is hormonally active. The recurrence of granulosa cell tumor often happens after 5 years. CASE PRESENTATION: We describe two cases of postmenopausal women with adult-type granulosa cell tumors of the ovary. Patient 1 is a 49-year-old European woman with a recurrent tumor; patient 2 is a 55-year-old European woman without recurrence of tumor. Urinary steroid profiles of patient 1 were monitored during a 5-year period starting from before an operation (13 samples). In patient 2, the urinary steroid profiles were monitored during a 3-year period starting from after an operation (six samples). The 24-hour urinary samples were examined and the urinary concentration of 20 androgen, progesterone, and corticoid metabolites was quantitatively determined by gas chromatography-mass spectrometry with selected ion-monitoring mode. CONCLUSIONS: Based on these cases a correlation could be observed between increased levels of the urinary steroids and the recurrence of ovarian granulosa cell tumor; therefore, we concluded that a urinary steroid profile could be a more effective method to follow-up such patients compared to the traditional serum hormones determinations supplemented with conventional tumor markers.

Monitoring menstrual cycle, gestation and lactation by measuring urinary oestradiol and progesterone in the captive golden snub-nosed monkey (Rhinopithecus roxellanae).

The golden snub-nosed monkey is an endangered species and study of its reproductive physiology is crucial for the species' breeding programs. Urine samples (770) from 5 mature female golden snub-nosed monkeys were collected in the Shengnongjia Nature Reserve between October 2013 and December 2014 to monitor their menstrual cycle, gestation, and lactation. The concentrations of oestradiol (E2) and progesterone (P4) in the samples were measured by Chemiluminescent Microparticle Immunoassay (CMIA), and the hormone concentrations were indexed to creatinine levels to compensate for differences in water content. The results showed that the E2 and P4 levels during the breeding season were significantly higher than those during the non-breeding season (P<0.01). The length of the menstrual cycle during the breeding season was 24.29±0.71days (mean±SEM) with a follicular cycle of 8.33±0.62days and luteal cycle of 15.27±0.83days. In addition, the levels of E2 and P4 began to rise on day 14 and day 10 after conception and remained at a high level until parturition. However, the E2 and P4 levels during lactation were lower than those during gestation (P<0.01). In summary, this study extends our knowledge regarding the basic reproductive physiology of golden snub-nosed monkeys, which could play an important role in the expansion of this species' population.

Quantifying endogenous androgens, estrogens, pregnenolone and progesterone metabolites in human urine by gas chromatography tandem mass spectrometry.

A method for the quantitation of 22 urinary steroids (androgens, estrogens and the main pregnenolone and progesterone metabolites) by means of gas chromatography tandem mass spectrometry using a triple quadrupole analyzer has been developed. Two different enzymatic hydrolysis protocols were investigated; one capable of releasing steroids present as both sulfates and glucuronides (total fraction), and another with ß-glucuronidase activity only. After selecting adequate internal standards and choosing the optimal instrumental parameters, i.e. chromatographic separation and ion transition conditions, the method was fully validated using both hydrolysis protocols. The method was shown to be linear (r >0.99) in the range of endogenous concentrations for all studied steroids with extraction recoveries higher than 80%. The use of labeled internal standards allowed for both a correct quantification and the evaluation of the rate of deconjugation for sulfates and glucuronides in every sample. In general, the sensitivity of the method was suitable for the detection of the endogenous levels, with limits of quantification ranging from 0.1 to 20ng/mL. Accuracies ranging from 80% to 120%, and relative standard deviations below 25% in intra- and inter- assay experiments were found for most of the analytes. The applicability of the validated method was tested by quantifying twenty-two metabolites in 24-h urine samples collected from healthy individuals. The ranges for the excretion of steroids in the total and glucuronide fractions obtained with the new method were compared with those available in the literature. By comparing the figures in both fractions, an estimation of the percentage that the sulfation represents for each steroid was also calculated. The presence of side enzymatic activities and the utility of the method for clinical studies as well as for doping control analysis is discussed.

Progesterone transfer among cohabitating female big brown bats (Eptesicus fuscus).

Experiments using female mice and bats have demonstrated that tritium-labeled 17ß-estradiol (3H-E2) can be absorbed via cutaneous and intranasal routes and distributed to reproductive and neural tissues. Radioactivity has also been measured in tissues of untreated females after 48h cohabitation with 3H-E2 injected males. The present study was designed to quantify steroid transfer among female bats. Radioactive quantification via liquid scintillation counting revealed absorption of tritium-labeled progesterone (3H-P4) in adult females 1h after cutaneous and intranasal application (10µCi). Subsequently, pairs of mature females were each housed for 48h with a single mature female that had been administered 3H-P4 (50µCi) via intraperitoneal injection. Radioactivity was observed in all collected tissues of all non-injected females at levels significantly greater than the control group. Following the same paradigm, radioactivity was not observed in the tissues of untreated female bats that were housed with stimulus females treated with 3H-E2 (50µCi). Enzyme immunoassays revealed measurable levels of unconjugated progesterone and estradiol in the urine of female bats, suggesting urine as a vector for steroid transfer. Given that bats of this species live in predominantly female roosts in very close contact, progesterone transfer among individuals is likely to occur in natural roosts.