Prioritization of social information by the basolateral amygdala in rats.

The amygdala is a collection of nuclei that support adaptive social behavior and are implicated in disorders such as autism. The basolateral complex of the amygdala (BLA), a main subdivision of the amygdala, influences fear responses, motivated behavior, and memory of emotional events via its communication with other amygdalar nuclei and with other brain regions such as the prefrontal cortex, striatum, and hippocampus. The specific role of the BLA in responses to social stimuli is less clear. The present study of female rats investigated the role of the BLA in responding to socially-relevant information by asking how inactivation of the BLA with bilateral infusions of the GABA receptor agonist muscimol would affect spontaneous exploration of wood blocks scented either with conspecific male or female urine or with nonsocial odorants. Conspecific urine samples were used because urine conveys information about sex, health, social status, and reproductive state in rodents. The results revealed that BLA inactivation reduced female rats' spontaneous preference for social odors over nonsocial odors, specifically for female urine. However, BLA inactivation did not generally impair rats' ability to distinguish two odors from the same category (e.g., urine odors from two different male rats). The results indicate that the BLA is important for responding to salience of social stimuli but not for discriminating between different individuals, a result that has important implications for amygdalar modulation of downstream attention, motivation, and memory processes for social stimuli.

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.

Urinary profiles of luteinizing hormone, estrogen and progestagen during the estrous and gestational periods in giant pandas (Ailuropda melanoleuca).

Luteinizing hormone (LH) is one of the main pituitary hormones that regulate ovulation, however its role has not been studied in giant panda. In this study, we developed an ELISA method for the detection of panda urinary LH. We analyzed urinary hormones of 24 female pandas during 36 breeding periods, we found females could easily be impregnated if the first mating occurred within 10 hours after LH peak. We also found the patterns of the ratios of urinary LH and progestagen in pandas that bred and successfully gave birth were significantly different from those that bred but failed to give birth. These data was the first to provide the urinary LH profiles during the estrous and gestational periods in pandas, and demonstrated that the appearance of the urinary LH peak indicated the timing of ovulation. The LH detection together with estrogen analysis makes the window for successful mating narrower than previously reported. Moreover, detection of urinary LH and progestagen can be used to discriminate between pregnancies and pseudopregnancies/miscarriages in the species. Thus, our findings suggest that LH not only plays a critical role in regulating ovulation but also plays an important role in maintaining pregnancy in the giant panda.

Either main or accessory olfactory system signaling can mediate the rewarding effects of estrous female chemosignals in sexually naive male mice.

A long-held view has been that interest of male mice in female body odors reflects an activation of reward circuits in the male brain following their detection by the vomeronasal organ (VNO) and processing via the accessory olfactory system. We found that adult, sexually naive male mice acquired a conditioned place preference (CPP) after repeatedly receiving estrous female urine on the nose and being placed in an initially nonpreferred chamber with soiled estrous bedding on the floor. CPP was not acquired in control mice that received saline on the nose before being placed in a nonpreferred chamber with clean bedding. Robust acquisition of a CPP using estrous female odors as the reward persisted in separate groups of mice in which VNO-accessory olfactory function was disrupted by bilateral lesioning of the accessory olfactory bulb (AOB) or in which main olfactory function was disrupted by zinc sulfate lesions of the main olfactory epithelium (MOE). By contrast, no CPP was acquired for estrous odors in males that received combined AOB and MOE lesions. Either the main or the accessory olfactory system suffices to mediate the rewarding effects of estrous female odors in the male mouse, even in the absence of prior mating experience. The main olfactory system is part of the circuitry that responds to chemosignals involved in motivated behavior, a role that may be particularly important for humans who lack a functional accessory olfactory system.

Gas chromatographic-mass spectrometric analysis of chemical volatiles in buffalo (Bubalus bubalis) urine.

Isolation of active fraction and characterization of chemosignals from urine have been attempted in several mammalian species in the recent years. The objective of this study was to identify the urinary volatiles across various reproductive stages of buffalo cow, namely, estrus, diestrus, and pregnancy, and in bull, by chemical extraction followed by gas chromatography-linked mass spectrometry (GC-MS). Urine samples were collected from six buffalo cows at two different phases of estrous cycle, namely, estrus and diestrus. Besides, urinary samples were collected from five pregnant buffalo cows (60-75 days after artificial insemination (AI)) and six adult bulls. Thin-layer chromatography was performed as a preliminary test for qualitative comparison of different compounds extracted by organic solvents. Identification of the urinary compounds was carried out in a gas chromatograph (Perkin Elmer, Autosystem XL) linked to a mass spectrometer (Turbomass). The results of GC-MS analysis indicated the presence of 21 compounds with varying molecular weights and retention time, which were further categorized as diestrus-specific, pregnancy-specific, and bull-specific urinary compounds. No compound, however, could be identified as estrus-specific. We concluded that qualitative differences do exist in estrus, diestrus, and pregnant buffalo cow urine and in bull urine, as evidenced by GC-MS.

Biochemical analysis of female mice urine with reference to endocrine function: a key tool for estrus detection.

Species-specific chemical signals released through urine, sweat, saliva and feces are involved in communication between animals. Urinary biochemical constituents along with pheromones may contribute to variation across reproductive cycles and facilitate to estrus detection. Hence, the present study was designed to analyze such biochemical profiles, such as proteins, carbohydrates, lipids, fatty acids, in response with steroid hormones such as estradiol and progesterone. The experimental groups were normal, prepubertal, ovariectomized, and ovariectomized with estrogentreated female mice. In normal mice, the protein and lipid concentrations in urine were significantly higher in proestrus and estrus phases and the quantity of fatty acids was also comparatively higher in estrus. Furthermore, certain fatty acids, namely tridecanoic, palmitic and oleic acids, were present during proestrus and estrus phases, but were exclusively absent in ovariectomized mice. However, the carbohydrate level was equally maintained throughout the four phases of estrous cycle. For successful communication, higher concentrations of protein and specific fatty acids in estrus are directly involved. The significant increase in estradiol at estrus and progesterone at metestrus seems to be of greater importance in the expression pattern of biochemical constituents and may play a notable role in estrous cycle regulation. Thus, we conclude that the variations observed in the concentration of the biochemical constituents depend on the phase of the reproductive cycle as well as hormonal status of animals. The appearance of protein and specific fatty acids during estrus phase raises the possibility to use these as a urinary indicators for estrus detection.

Reproductive life history traits of female orangutans (Pongo spp.).

Data from wild populations demonstrate that orangutans have the slowest life history of all the great apes. In this chapter, we provide an overview of reproduction and life history traits of female orangutans in the wild and captivity. This comparison of wild and captive data illustrates the variability that exists for orangutans. Wild orangutan females first reproduce at a mean age of 15.4 years, with an age range of 13-18 years, and they have a mean interbirth interval of 9.3 years. Wild male orangutans are conservatively estimated to live at least 58 years, and 53 years for females [1], and to date, there is no evidence to suggest that wild orangutans experience reproductive senescence. We use captive data from 2,566 individuals to show that in captivity orangutan females regularly begin reproducing at the age of 7 and have interbirth intervals that can be shorter than 1 year. We provide additional data that describe the onset and normalization of menses in a young adolescent orangutan as well as the reproductive cycles of three adult females of different ages. Although captive females routinely cycle and reproduce throughout much of their lifespan, age at last reproduction in captivity is 41, which is well before maximum female lifespan. To date, longevity in the wild and in captivity appears equivalent [2]. The reasons for the presence of a postreproductive lifespan in captivity as opposed to its absence in wild populations may be related to management issues. The above results indicate a need for more detailed comparisons between wild and captive orangutans using similar methodologies.

Development and evaluation of a rapid enzyme-immunoassay system for measurement of the urinary concentration of estrone-3-glucuronide in a female giant panda (Ailuropoda melanoleuca).

To detect estrus for reproductive management, and to determine the relationship between urinary estrogen and estrous behavior, in a female giant panda, we developed and evaluated a rapid enzyme immunoassay (EIA) system for urinary Estrone-3-glucuronide (E1G) using commercial reagents. The developed EIA system took only around 3 hours, including all procedures to obtain a result. It indicated good reproducibility (intra-assay CV of 5.16%, interassay CV of 15.4%) and sensitivity (lowest standard concentration was 0.0156 ng/ml) for measurement of the urinary concentrations of E1G in the giant panda. There was a positive correlation (r=0.934) with the data for estrone (E1) in the same samples, as measured by radioimmunoassay (RIA) performed in a commercial laboratory. The changes in the E1G concentrations were almost synchronous with the changes in E1 assayed by RIA in urine collected during 4 consecutive estrous seasons. The dynamics of urinary E1G measured by this system highly correlated with the occurrence of the presenting estrous behavior in the giant panda. The above results indicate that this assay system may be normally, rapidly and practically used for measurement of the urinary concentration of E1G in the giant panda.

The advertisement role of major urinary proteins in mice.

The variation of expression of major urinary proteins was studied in laboratory mice to further the understanding of the role of these proteins in social and reproductive contexts. Mouse major urinary proteins (MUPs) are known to carry volatile substances and protect them during their passage from the liver, through the kidneys into the urine. However, most studies on the role of MUPs were carried out on males. Using densitometry analysis of total MUP concentration in the urine, our present study clearly demonstrates that (i) individuals of both sex up-regulate MUPs during social contact, and that (ii) females use these proteins to advertise their reproductive state by varying the concentration of MUPs during the oestrous cycle. As the concentration of MUPs was normalized by the concentration of creatinine -- a marker of glomerular filtration -- the corrected concentration of MUPs represents instantaneous expression on the level of proteins. Cross-correlation analysis between oestrus quantification and MUP expression revealed that the oestrous curve is delayed by 1 day behind the MUP curve so that the expression of MUPs is up-regulated immediately at the beginning of oestrus. To conclude, the regulation of pheromone-carrying MUPs is directly linked to reproduction and, thus, enables female honest signalling.

Do dispersing non-reproductive female Damaraland mole-rats, Cryptomys damarensis (Rodentia: Bathyergidae) exhibit spontaneous or induced ovulation?

The Damaraland mole-rat is a eusocial, subterranean rodent that exhibits a seasonal breeding. Non-reproductive females show physiological suppression of reproduction whilst in the confines of the natal colony. This study set out to investigate whether dispersing female Damaraland mole-rats exhibit induced or spontaneous ovulation. Fifteen non-reproductive females were removed from their natal colonies and housed individually for a period of 6 weeks. During this period urine was collected from all animals every second day. After this initial period the animals were divided into 3 groups. Females were subjected to 1 of 3 trials: a control group housed separately without a male, allowed non-physical contact, or placed in direct physical contact with vasectomized males. Urine was collected for a further 5 weeks, and urinary progesterone profiles established. All three groups showed a significant difference in urinary progesterone concentrations between the two treatment periods indicating initiation of follicular development in all animals following removal from the colony. Histological results further revealed that at least one animal in the control group and five of the six animals in the vasectomized group had corpora lutea present in the ovaries showing that ovulation has taken place. All groups had similar numbers of Graafian follicles, indicating that all females were likely to ovulate in the near future. This finding suggests that females are capable of spontaneous ovulation, but the act of coitus may advance the onset of ovulation in this arid dwelling mole-rat.

Non-invasive repeated measurement of urinary progesterone, 17beta-estradiol, and testosterone in developing, cycling, pregnant, and postpartum female mice.

Excretory samples from adult female mice were collected non-invasively during development, estrous cycling, pregnancy, and postpartum. In initial studies, urinary measures were statistically more dynamic over days than were fecal measures; thus subsequent studies focused on urine. Higher 17beta-estradiol levels were present in isolated females than in those exposed to males. In cycling females, urinary 17beta-estradiol was more variable than were measures of testosterone or progesterone, showing peaks with an approximate 5-day periodicity. When urinary estradiol and progesterone were monitored in conjunction with vaginal smear cell counts, patterns were idiosyncratic; most females showed distinct peaks in urinary steroids, not in clear synchrony with vaginal cell cornification. Levels of progesterone rose markedly during the first 10 days of pregnancy, then declined before birth. Estradiol showed a substantial peak on days 7-8 of gestation in all females measured. Urinary testosterone was not dynamic during pregnancy, but rose in immediate prenatal and postpartum measures. During post-weaning, pre-pubertal development, urinary levels of progesterone remained constant but levels of estradiol rose substantially over time.

Annual changes of urinary progesterone and estradiol-17beta of the Dugong (Dugong dugon) in captivity.

Levels of urinary progesterone and estradiol-17 beta were measured twice a week in a female dugong, Dugong dugon, in captivity for two years from April 1996 to April 1998. The dugong showed 14 ovarian cycles during the period of study. Concentrations of progesterone ranged from 0.01 ng/mg creatinine (Cr) to 1.94 ng/mg Cr and the length of estrous cycle was 53.6+/-8.6 (mean+/-SEM) days based on intervals of urinary progesterone peak-to-peak measurements. Concentrations of urinary estradiol-17 beta ranged from 0.9pg/mgCr to 23.7pg/mgCr, and tended to peak just prior to elevations of progesterone during the first year of study. This is the first report demonstrates that the ovulatory cycle of the dugong is about 50 days. The present findings suggest that measurement of urinary progesterone is a useful method to detect ovarian cycle of the dugong in captivity.

Female odors lead to rapid activation of mitogen-activated protein kinase (MAPK) in neurons of the vomeronasal system.

Pheromonal mediation of reproductive function proceeds along a neuroanatomical pathway that connects the vomeronasal organ (VNO) at the periphery with downstream target-sites in the amygdala and hypothalamus. The MAPK pathway is a prominent cascade linking receptor activation to induction of effectors such as c-Fos. We addressed the question: Does a specific pheromone stimulus lead to activation (phosphorylation, P) of MAPK in the VN system of the male mouse? Phosphorylation of MAPK in the VN system was evaluated 15-30 min and 1.5-2 h after exposure to female odors, using immunocytochemical techniques. A rapid and transient cytoplasmic expression of PMAPK was noted in the VNO with a unique distribution of the expressing neurons in columns extending over the entire basal to apical axis. A rapid and sustained expression was noted in most amygdaloid and hypothalamic VN target-sites and also in a few amygdaloid and hypothalamic sites outside the traditional VN system. The extent of expression and the subcellular compartmentalization (nucleus, cytoplasm, processes) of PMAPK were region-dependent. Of the VN target-sites, the accessory olfactory bulb (AOB) stood out in the lack of expression of PMAPK, in the high expression of the MAPK enzyme itself and in the massive of expression of c-Fos. This expression profile implicates another pathway(s) in mediating VNO signaling to the AOB. Our results are the first to demonstrate the use of PMAPK to trace functional pathways. Based on the wide cellular and intracellular expression of phosphorylated MAPK in the VN system, we propose that the MAPK pathway plays an important role in mediating female pheromone signaling in the male mouse.