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The objective of the present study was to determine the effects of soya and whey milk protein, α-lactalbumin (α-LA), on mammary gland morphology and the structural support of the gland, in pre-pubertal mice after 7 d of treatment. In Expt 1, weaned (day 21) CD1 mice were given one of the four treatments, three included dietary supplements: (1) control diet, casein, (2) soya, (3) α-LA and (4) subcutaneous injection of 2·5 μg oestradiol benzoate in 20 μl maize oil and fed the control diet. All diets were isoenergetic with equal protein concentrations. All groups that were not treated with oestradiol received the vehicle. Whole-mount analyses were performed to determine longitudinal ductal growth and terminal end bud development. DNA was extracted from the gland and assessed by spectrophotometry (260/280 nm). Tissue extracts for extracellular matrix (ECM) proteins, matrix metalloproteinase-2 (MMP(2)), tissue inhibitor of MMP(2) (TIMP(2)), and serum oestradiol and mammary tissue epidermal growth factors (EGF) were measured by immunoassays. Expt 2 utilised the Her2/neu transgenic strain, with the same protocols. Statistical significance was determined by one-way ANOVA. From Expt 1 and 2, soya and α-LA significantly increased ductal elongation when compared with the oestrogen and control groups. These results were corroborated by data on total DNA and the ratio of MMP(2):TIMP(2). The ratio of MMP(2):TIMP(2) was affected by α-LA. Serum oestradiol was decreased only in the oestradiol-treated groups in both experiments. Soya is known to be oestrogenic and can act on epithelia directly. The mechanism by which α-LA affects glandular development is by modulating the ECM or by promoting the synthesis/activity of EGF.

The preputial glands of house mice express the gene for the fifth melanocortin receptor (MC5-R) and are a primary source of urinary pheromones involved in inter-male aggression. A 'resident-intruder' behavioral model was used to compare the responses of resident males to urine from mice with an engineered disruption of the fifth melanocortin receptor (MC5-RKO) with residents' responses to urine from wild-type mice (WT). Each type of urine was presented in combination with a castrated intruder male to provide the appropriate biological context. Resident males responded with a longer latency to bite when the urine was from gonadally intact WT males compared with urine from MC5-RKO mice. These results are consistent with the hypothesis that activation of the fifth melanocortin receptor in the preputial glands of male house mice causes excretion of urinary pheromones that delay aggressive responses by other males.

Several short-duration tests have demonstrated that the surgical removal of the vomeronasal organ (VNX) from sexually-inexperienced male rodents results in a reduction in copulatory behavior, compared to the effects of sham surgery (SHAM). We extended these studies to adult male prairie voles, Microtus ochrogaster, and substantially increased the duration of the tests. During the initial interactions with females, VNX males spent significantly less time with their noses in close proximity to the females than did SHAM males. Moreover, only two of nine VNX males sired offspring after having been paired with females for 8 weeks, whereas nine of 12 SHAM males sired offspring in that interval. We also found that VNX and SHAM males were equivalently non-aggressive to an anesthetized stimulus-male prior to being paired with females. However, after spending 2 weeks paired with a female, the VNX males were significantly less aggressive than were the SHAM males, possibly as a result of having copulated less often. In a later test, nearly all of the VNX and SHAM males that sired offspring were vigorously aggressive to a stimulus male. We conclude that the stimulation of the vomeronasal system in sexually-inexperienced male prairie voles is important for maximal reproductive performance and that the VNX-induced impairment in reproduction is associated with a decrease in inter-male aggression. The possible sensory effects of the vomeronasal system on the neural and endocrine control of reproduction and behavior are discussed.

Cranial tissue from adult and neonatal voles was examined with immunocytochemical techniques to determine the distribution of luteinizing hormone-releasing hormone-immunoreactive (LHRH-ir) neurons in extracerebral structures. The overall distribution of the LHRH-ir portion of the nervus terminalis in the nasal and extracranial cavities was comparable to that of other rodents. However, we observed a unique association of LHRH-ir neurons with the pterygopalatine ganglia of neonatal and adult voles. We also found LHRH-ir fibers in nasopalatine nerves, and trigeminal nerves and ganglia of neonatal voles. We speculate that these neurons may influence the autonomic control of the vascular pump in the vomeronasal organ.

Chemical cues from male voles activate reproduction in female prairie voles (Microtus ochrogaster). Twelve hours of contact with a male, followed by exposure to his soiled bedding for 2 days, is sufficient to initiate follicular maturation and induce uterine hypertrophy. Our recent work indicates that the chemosensory vomeronasal organ (VNO) can mediate this response. Here, we examined whether other sensory systems can acquire the ability to activate female reproduction as a result of learning or experience. To explore this issue, the VNO was removed (VNX) from nulliparous and primiparous females who were then exposed to cues from males. In Experiment 1, we found that nulliparous VNX females had lower uterine and ovarian weights than did sham-operated females. In Experiment 2, we determined that sexual experience did not ameliorate the reproductive deficits normally induced by VNX. The present results contrast with those of previous studies suggesting that males of some rodent species, when allowed reproductive experience prior to VNX, can utilize other sensory systems to mediate subsequent reproductive responses. We conclude that the role of the VNO in transducing chemosensory information is crucial for coordinating the reproductive efforts of male and female prairie voles.

In the last decade, research in our laboratories has focused on the effects of deafferentation of the mammalian chemosensory vomeronasal organ (VNX). Many different assays have been conducted and the results of some are briefly reviewed in this contribution, including the effects of VNX on neuroanatomical assessments using histochemistry (lectin binding) and immunohistochemistry (LHRH), male mouse and prairie vole ultrasonic vocalizations and hormone surges in response to cues from females, male mouse courtship and sexual behavior, territorial marking and inter-male aggression, the production of a puberty-altering substance found in mice, activation of reproduction in female voles (who generally do not exhibit estrous cycles) and maternal behaviors by female mice, including aggression directed toward intruder males. In some instances, the otherwise detrimental effects of VNX can be overcome by experience prior to deafferentation, especially in assays that are dependent upon expressions of behavior. In other situations, experience may have little impact on amelioration of the effects of VNX. The essential conclusions of this work focus our attention on reproductive physiology and behavior and a role for the vomeronasal organ in the perception of pheromones that modulate these functions.

We investigated the reproductive biology of an induced ovulator, the pine vole (Microtus pinetorum). Male puberty, measured as age at first impregnation, was found to occur as early as 44 days of age. Female puberty measured as age at first conception, was found to occur as early as 32 days of age, considerably earlier than previously reported. Females paired with stud males exhibited a doubling of uterine weight within 12 h, and vaginal sperm were present after 48 h. This indicates that although behavioral responses to males--including mating--require prolonged contact, physiological responses to males occur rapidly. Chemosignals from males slightly increased uterine and ovarian weights of females, but chemosignals from other females did not. Young females paired with stud males for 48 h in the presence of soiled bedding from the female's family had significantly smaller increases in ovarian and uterine weights than similar females paired on clean bedding. Suppression of reproduction in female offspring while they remain with the extended family unit is discussed as a life-history tactic and as a possible mechanism for inbreeding avoidance.

We investigated the vocalizations produced by adult pine voles during various social interactions by presenting an experimental animal with either an anesthetized or awake (unanesthetized) conspecific. Ultrasonic vocalizations (USVs) occurred frequently during tests in which an awake male was present, but were rarely detected in tests involving only awake females, or when a female was presented with an anesthetized conspecific. Higher rates of USVs were produced when males were tested with a familiar female than when tested with an unfamiliar male or female. Equivalent rates were produced when males were presented with either anesthetized or awake animals, but female-soiled bedding failed to elicit USVs from males. Sonic vocalizations (SVs) were produced by both sexes and were associated with aggressive behavior, but occurred only in tests between awake, unfamiliar animals. Castration greatly reduced and testosterone therapy restored USVs emitted by males in response to anesthetized conspecifics. Our results suggest that (i) USVs are emitted predominantly by males; (ii) familiarity enhances USV response; (iii) SVs are produced during aggressive interactions; and (iv) androgens regulate the production of USVs by males. Possible roles for pine vole vocalizations are discussed.

The reproductive system of female prairie voles remains quiescent in the absence of stimulation from males; however, chemosignals from males are capable of at least partially activating female reproduction. In other species, the vomeronasal system mediates some of the reproductive responses of females to males. We found that surgical removal of the vomeronasal organ (VNX) from adult female prairie voles impeded reproductive activation in response to pairing with stud males: ovarian and uterine weights of VNX females paired with stud males for 24 or 60 hours were significantly less than those of normal (NORMAL) or sham-operated (SHAM) females. Furthermore, 8 of 9 NORMAL, 10 of 13 SHAM, but only 4 of 9 VNX females paired with stud males for 60 hours mated. VNX females, however, were still able to use chemosensory cues to locate food. Behavioral observations of females encountering stud males were similar for VNX, SHAM and NORMAL females. We conclude that vomeronasal chemoreception may be a primary component of reproductive activation in female prairie voles, presumably by mediating neuroendocrine responses to chemosignals.

Male and female Microtus ochrogaster were presented with anesthetized and awake conspecifics while ultrasonic vocalizations (USVs) were monitored. Males produced significantly more USVs than females during 5-min testing sessions. Males tended to produce more USVs to unfamiliar females than to familiar female siblings. Sexual experience led to increased USV scores by males. These results suggest that USVs by male prairie voles communicate to females the male's gender and his availability for reproductive behavior.

Adult male mice excrete a urinary chemosignal that accelerates puberty in females, whereas group-housed female mice excrete a urinary chemosignal that delays puberty in young females. We found that: (1) the excretion of the puberty-acceleration chemosignal by males persisted in the absence of the vomeronasal organs and (2) the puberty-delay chemosignal was not present in the urine of group-housed females whose vomeronasal organs had been surgically removed (VNX), but was present in the urine of group-housed females subjected to sham surgery (SHAM). These results suggest that in males, vomeronasal chemoreception does not affect the excretion of the puberty-acceleration chemosignal, but that in females, the vomeronasal organ receives chemosignals that influence the excretion of the puberty-delay chemosignal. Additionally, we found no difference between SHAM and VNX females in rates of conception, litter size, pup growth, pup recognition, or maternal behavior, indicating that normal maternal processes are expressed in the absence of an intact accessory olfactory system.

Detection of reproductive status from chemical signals at sandbathing loci was examined in Merriam's kangaroo rat. Castrated and intact males were attracted similarly to the sandbathing loci of four classes of scent donors: intact males, castrated males, estrous females, and nonestrous females. This suggested that reproductive status was not communicated to males at sandbathing loci. In a control test, castrated males were attracted to disturbed sand that contained no olfactory cues whereas intact males were not. Castration caused a significant decrease in the size of a specialized sebaceous gland on the dorsum. Castration did not affect total pelage lipids, however, indicating that lipid-producing cells have a baseline level of secretion that is necessary for pelage maintenance.