Gonadal hormones, but not sex, affect the acquisition and maintenance of a Go/No-Go odor discrimination task in mice.

In mice, olfaction is crucial for identifying social odors (pheromones) that signal the presence of suitable mates. We used a custom-built olfactometer and a thirst-motivated olfactory discrimination Go/No-Go (GNG) task to ask whether discrimination of volatile odors is sexually dimorphic and modulated in mice by adult sex hormones. Males and females gonadectomized prior to training failed to learn even the initial phase of the task, which involved nose poking at a port in one location obtaining water at an adjacent port. Gonadally intact males and females readily learned to seek water when male urine (S+) was present but not when female urine (S-) was present; they also learned the task when non-social odorants (amyl acetate, S+; peppermint, S-) were used. When mice were gonadectomized after training the ability of both sexes to discriminate urinary as well as non-social odors was reduced; however, after receiving testosterone propionate (castrated males) or estradiol benzoate (ovariectomized females), task performance was restored to pre-gonadectomy levels. There were no overall sex differences in performance across gonadal conditions in tests with either set of odors; however, ovariectomized females performed more poorly than castrated males in tests with non-social odors. Our results show that circulating sex hormones enable mice of both sexes to learn a GNG task and that gonadectomy reduces, while hormone replacement restores, their ability to discriminate between odors irrespective of the saliency of the odors used. Thus, gonadal hormones were essential for both learning and maintenance of task performance across sex and odor type.

A sex comparison of the anatomy and function of the main olfactory bulb-medial amygdala projection in mice.

We previously reported that some main olfactory bulb (MOB) mitral/tufted (M/T) cells send a direct projection to the "vomeronasal" amygdala in female mice and selectively respond to volatile male mouse urinary odors. We asked whether MOB M/T cells that project to the vomeronasal amygdala exist in male mice and whether there is a sexually dimorphic response of these neurons to volatile male urinary pheromones. Gonadectomized male and female mice received bilateral injections of the retrograde tracer, Cholera toxin-B (CTb) into the medial amygdala (Me), which is part of the vomeronasal amygdala. All subjects were then treated with estradiol benzoate and progesterone before being exposed to volatile male urinary odors whereupon they were sacrificed 90 min later. Sections of the MOB were immunostained for Fos protein and/or CTb. Male mice, like females, displayed a small population of MOB M/T cells that project to the Me. While the general localization of these cells was similar in the two sexes, there were statistically significant sex differences in the percentage of MOB M/T cells in the anterior and posterior medial segments of the MOB that were retrogradely labeled by CTb. Male urinary volatiles stimulated equivalent, significant increases in Fos expression by MOB M/T neurons projecting to the Me in the two sexes. By contrast, in the same mice exposure to male urinary volatiles stimulated a significant increase in Fos expression by mitral cells in the accessory olfactory bulb (AOB) only in female subjects. Thus any sexually dimorphic behavioral or neuroendocrine responses to male urinary volatiles likely depend on the differential processing of these odor inputs in the AOB and/or other downstream forebrain structures after their detection by the main olfactory system.

Comparison of urinary odor-induced glomerular activation in the main olfactory bulb of aromatase knock-out and wild type female mice.

Previously [D.W. Wesson, M. Keller, Q. Douhard, M.J. Baum, J. Bakker, Enhanced urinary odor discrimination in female aromatase knockout mice, Horm. Behav. 49 (2006) 580-586] female aromatase knock out mice successfully learned to discriminate in a food-motivated go/no-go task between urinary volatiles from ovariectomized female mice treated with estradiol as opposed to estradiol plus progesterone whereas wild type females failed to learn this odor discrimination. We asked whether this behavioral difference is reflected in the ability of these two types of urinary volatiles to differentially stimulate Fos expression in juxtaglomerular cells (an index of glomerular activation) of the main olfactory bulb (MOB) in wild type versus ArKO female mice. Statistically significant differences in the profiles of MOB glomerular activation were seen in ovariectomized, estrogen-treated ArKO as well as WT female subjects following exposure to urinary volatiles from ovariectomized females given estradiol alone as opposed to estradiol plus progesterone. Therefore, previously observed differences between females of the two genotypes in their behavioral responses to these odors must reflect differential processing in more central segments of the olfactory pathway instead of in the MOB.

Sex and gonadal steroid modulation of pheromone receptor gene expression in the mouse vomeronasal organ.

Non-volatile chemosignals in rodents are detected by unique receptors in the vomeronasal organ of the accessory olfactory system. Although the vomeronasal organ has been implicated in the regulation of sexually dimorphic behavioral and neuroendocrine functions, the underlying cellular mechanisms are undetermined. In previous studies we showed that exposure to soiled male bedding augmented immediate early gene immunoreactivity in neurons of the basal zone of the vomeronasal organ, an effect that depended on gender and sex steroid expression. To determine whether this effect could be due to differences in vomeronasal organ receptor expression, we examined two representatives (VR1 and VR4) from different subfamilies of the V2R family of receptors that are expressed in the basal zone of the vomeronasal organ. Adult Swiss-Webster male and female mice were gonadectomized and implanted with capsules containing 17beta-estradiol, testosterone or neither steroid (control). Two weeks later vomeronasal organs were processed for in situ hybridization using probes from the N-terminal extracellular domains of VR1 and VR4. Expression of both VR1 and VR4 was significantly higher in males than in females. Estradiol, but not testosterone-treated, males had significantly lower levels of VR1 expression in the caudal vomeronasal organ compared with untreated gonadectomized males. In contrast, testosterone enhanced VR4 expression in males relative to similarly treated females. Despite these effects, we found no evidence that vomeronasal organ neurons express either androgen or estrogen receptors. These data show that expression of vomeronasal organ receptors in mice is sexually dimorphic and regulated by sex steroids. Thus, gonadal hormones may affect the response of vomeronasal organ neurons to chemosignals by altering levels of the receptors to which they bind.

Comparison of odor and mating-induced glomerular activation in the main olfactory bulb of estrous female ferrets.

Previously [S.K. Woodley, M.J. Baum, Differential activation of glomeruli in the ferret's main olfactory bulb by anal scent gland odors from males and females: an early step in mate identification, Eur. J. Neurosci. 20 (2004) 1025-1032], the receipt of intromission from a male activated glomeruli (indexed by Fos immunoreactivity in juxtaglomerular cells) in the main olfactory bulb (MOB) of estrous female ferrets which exceeded the activation seen after exposure to male anal scent gland odorants alone. We asked whether centrifugal inputs (e.g., from the locus coeruleus to the MOB) generated by the receipt of vaginal-cervical stimulation influence odor-induced MOB glomerular activation. We compared the activation of MOB glomeruli in estrous female ferrets which received a unilateral naris occlusion prior to exposure to: unscented air, volatile odorants from an anesthetized male, volatile + non-volatile odorants from direct physical contact with an anesthetized male, or mating stimulation. Little glomerular activation was observed in the MOB ipsilateral to an occluded naris, including females which received intromission. An equivalent distribution of activated glomeruli was observed in the ventral MOB of estrous females which either received mating stimulation or had direct physical contact with an anesthetized male. Considerably less glomerular activation occurred in females exposed only to volatile male odors. The MOB of female ferrets responded to body odorants from the opposite sex; however, there was no evidence that mating-induced centrifugal inputs directly activated MOB glomeruli or modified odor-induced glomerular activation.

Putative chemosignals of the ferret (Mustela furo) associated with individual and gender recognition.

Quantitative stir bar sorptive extraction methods, both in the aqueous and headspace modes, followed by thermal desorption gas chromatography-mass spectrometry were used to investigate individual variations in the volatile components of male and female ferret (Mustela furo) urine. The urinary profiles were further compared with volatile profiles of anal gland secretions of breeding male and female ferrets. Thirty volatile compounds were quantified in male and female urine. Among them, 2-methylquinoline was unique to male urine. Four ketones (4-heptanone, 2-heptanone, o-aminoacetophenone, and a dimethoxyacetophenone) and several nitrogen compounds (e.g., 2,5-dimethylpyrazine, quinoline, 4-methylquinazoline) and low levels of three unidentified nonsulfur compounds were significantly more abundant in males than in females. Quantitative comparison of 30 volatile urinary compounds showed several statistically significant differences between the sexes and individuals of the same sex. These findings suggest that ferrets may use urine marking for sex and individual recognitions. Ten of the 26 compounds identified in anal gland secretions from females and males were also found in urine. However, most of the major compounds (thietanes, dithiolanes, and indole) in anal glands were not present in urine. This suggests that urine may convey specific signals that differ from those of anal glands. Additionally, 10 volatiles (two aldehydes, five ketones, benzothiazole, 2-methylquinoline, and 4-methylquinazoline), not previously identified, were found in ferret anal gland secretions. Among the new compounds, o-aminoacetophenone was found only in males, while only traces of this compound were found in females. Similar results were previously obtained in anal glands of three other Mustela species. These findings provide new information about the constituents of urine and volatile components of anal gland secretions in ferrets.

Effects of vomeronasal organ removal on olfactory sex discrimination and odor preferences of female ferrets.

Previous research suggests that body odorants, including anal scents and urinary odors, contribute to sex discrimination and mate identification in European ferrets of both sexes. We assessed the possible role of the vomeronasal organ (VNO) in these functions by surgically removing the organ bilaterally in sexually experienced female ferrets. Lesioned (VNOx) and sham-operated control (VNOi) females reliably discriminated between male- and female-derived anal scent gland as well as fresh urinary odors in habituation/dishabituation tests. However, VNOi females spent significantly more time than VNOx subjects investigating male urinary odors in these tests. Also, VNOi females, but not VNOx subjects, preferred to investigate day-old male versus female urine spots as well as wooden blocks that had previously been soiled by male versus female ferrets. Both groups of female ferrets preferred to approach volatile odors from a breeding male instead of an estrous female in Y-maze tests and both groups showed similar levels of receptive sexual behavior in response to a male's neck grip. The VNO is apparently not required for olfactory sex discrimination or mate recognition in this carnivore, but instead may play a role in promoting continued contact with nonvolatile body odors previously deposited by opposite-sex conspecifics during territorial scent marking.

Contribution of anal scent gland and urinary odorants to mate recognition in the ferret.

Previous research [J. Neurosci. 21 (2001) 5832-5840] showed that ferrets of both sexes require olfactory signals to identify opposite-sex mating partners at a distance. The present experiments assessed the contributions of anal scent gland and urinary odorants to these preferences. Sexually experienced, ovohysterectomized female and castrated male ferrets were injected daily with estradiol benzoate and testosterone propionate, respectively. When tested in an airtight Y-maze, subjects of both sexes preferred to approach volatile odors emitted from opposite- versus same-sex stimulus ferrets that were anesthetized and placed in the goal boxes, regardless of whether the anal scent glands of stimulus ferrets had been surgically removed or left intact. Subjects of each sex showed an equal preference to approach volatile odors emitted from anesthetized opposite-sex ferrets that were scent-gland intact as opposed to descented. Female subjects preferred to approach volatile anal scent gland odorants, as well as urinary odorants from male, as opposed to female conspecifics. Male subjects preferred to approach volatile anal scents from females versus males; however, males showed no preference for female over male urinary odorants. Our results suggest that anal scent gland odorants are sufficient, but not required, for mate recognition in the ferret. Instead, a combination of body odorants including, but not restricted to, those derived from anal scent gland secretions apparently underlie olfactory sex discrimination and partner preference in this carnivore.

Interactive effects of testosterone and superior cervical ganglionectomy on attraction thresholds to volatile urinary odors in gonadectomized mice.

Volatile urinary odors contribute to mate recognition in mice after their detection by the main olfactory epithelium (MOE). We used a habituation/dishabitution task to ask whether the capacity of gonadectomized mice of both sexes to detect and investigate decreasing concentrations of volatile urinary odors from either breeding males or estrous females is modulated by administering androgen or estrogen and if so, whether any effects of these sex steroids are altered by disrupting the sympathetic innervation of the MOE via bilateral superior cervical ganglionectomy (SCGx). In tests given, beginning 51 days after gonadectomy without steroid treatment both male and female subjects detected even the lowest concentrations (1:120 and 1:160 dilutions by volume) of male urinary odors, provided they were SCGx as opposed to sham operated. In subsequent tests given after estradiol benzoate (EB) followed later by 5alpha-dihydrotestosterone (DHT) treatments, neither male nor female subjects detected low concentrations of male urinary odors regardless of whether or not their SCG's were intact. Administration of testosterone (T) prior to a final series of tests restored the ability of gonadectomized subjects of both sexes to detect low concentrations of male urinary odors regardless of their SCG status. This suggests that T, but not its neural metabolites estradiol, or DHT, facilitates responsiveness to low concentrations of male odors in mice of both sexes. In tests given 51 days after gonadectomy without steroid treatment most male and female subjects readily detected the three highest concentrations of estrous female urinary odors whereas SCGx males and females failed to detect the lowest concentrations of these odors. After treatment with EB and then with DHT, gonadectomized mice of both sexes generally failed to detect the three lowest concentrations of estrous female urinary odors regardless of their SCG status. After T treatment; however, subjects of both sexes again detected most dilutions of estrous female urine, provided their SCG's were intact. Again, these results suggest that T, but not estradiol or DHT, facilitates responsiveness to estrous female urinary odors. Provided such an activational effect of T is present, sympathetic, noradrenergic inputs to the MOE may enhance odorant contrast, as previously suggested [Nat. Neurosci. 2 (1999) 106], by reducing the responsiveness of olfactory neurons to low (1:120 and 1:160 dilutions) concentrations of some biologically significant odorants (e.g. male urinary odors) while facilitating their responsiveness to low to moderate (1:80 dilution) concentrations of others (e.g. estrous female urinary odors).

Removal of the superior cervical ganglia fails to block Fos induction in the accessory olfactory system of male mice after exposure to female odors.

When exposed to female odors, testosterone-primed male mice show a robust expression of immediate early genes in the vomeronasal organ (VNO) and associated accessory olfactory structures. We asked whether the superior cervical ganglia (SCG), which provide autonomic inputs to the VNO, are required for odor induction of Fos. Gonadally intact male mice received sham, unilateral, or bilateral SCG lesions and were exposed to odors from estrous females. In comparison to clean bedding, female odors significantly increased neuronal Fos immunoreactivity in sites throughout the VNO projection pathway, but these responses were not reliably modified by SCG removal. Thus, noradrenergic inputs to the VNO, which regulate a pumping mechanism thought to facilitate entrance of chemosignals into the VNO lumen, are not required for odors to induce Fos in the mouse accessory olfactory system.

Induction of Fos in the accessory olfactory system by male odors persists in female mice with a null mutation of the aromatase (cyp19) gene.

The ability of odors from soiled male bedding to induce neuronal Fos-immunoreactivity (IR) in sensory neurons located in both the apical and basal zones of the vomeronasal organ (VNO) and in two segments of the VNO-projection pathway, the anterior nucleus of the medial amygdala and the bed nucleus of the stria terminalis (BNST), was significantly reduced in adult, ovariectomized, estrogen-treated female mice with a homozygous null mutation of the cyp19 gene (ArKO) which encodes the estrogen biosynthetic P450 enzyme, aromatase. However, a significant odor-induced activation of Fos-IR was seen in other segments of the VNO-projection pathway of ArKO females, including the accessory olfactory bulb (AOB) granule cell layer, the posterior-dorsal medial amygdala (MePD), and the medial preoptic area (MPA). These results suggest that the VNO/accessory olfactory pathway to the hypothalamus was functional in ArKO females even though they had presumably been exposed to less estrogenic stimulation than wild-type (WT) control females throughout development and until the time that estrogen treatment was begun in adulthood. Thus, the hypothesis of Toran-Allerand [Prog. Brain Res. 61 (1984) 63] that female-typical features of neuroendocrine and behavioral function require perinatal exposure to estrogen was not supported, at least for the VNO/accessory olfactory system.

Sexually dimorphic activation of galanin neurones in the ferret's dorsomedial preoptic area/anterior hypothalamus after mating.

Male ferrets in breeding condition possess three times as many galanin-immunoreactive (IR) neurones as oestrous females in the sexually dimorphic dorsomedial preoptic area/anterior hypothalamus (dmPOA/AH). Using Fos-IR as a marker of activation, we investigated whether mating with intromission differentially activates this sexually dimorphic group of galanin-IR neurones in male and female ferrets. Male ferrets that intromitted had a significantly greater percentage of galanin-IR neurones in the dmPOA/AH that were colabelled with nuclear Fos-IR than oestrous females that received an intromission. Intromissive stimulation augmented Fos-IR in an equal percentage of galanin-IR neurones in both sexes in the medial amygdala (MA) and bed nucleus of the stria terminalis (BNST). Peripheral anosmia induced by bilateral occlusion of males' nares did not reduce the mating-induced activation of galanin-IR neurones in the dmPOA/AH, and there was a significant correlation among individual males between intromission duration and the percentage of dmPOA/AH galanin-IR neurones colabelled with Fos-IR. Exposure of castrated, testosterone propionate-treated male ferrets to either soiled bedding or to volatile odours from oestrous females failed to induce nuclear Fos-IR in galanin-IR neurones located in the dmPOA/AH, BNST or MA, suggesting that the mating-induced activation of galanin-IR forebrain neurones in male ferrets depends more on genital-somatosensory than on olfactory inputs. The observed sex dimorphism in the mating-induced activation of galanin-IR neurones in the dmPOA/AH raises the possibility that these neurones perform a mating-dependent function that occurs only in males.

Sex difference in attraction thresholds for volatile odors from male and estrous female mouse urine.

Volatile urinary odors from opposite sex conspecifics contribute to mate recognition in numerous mammalian species, including mice. We used a simple habituation/dishabituation testing procedure to ask whether the capacity to detect and investigate decreasing concentrations of volatile urinary odors is sexually differentiated in mice. Beginning 2 months after gonadectomy and in the absence of any sex steroid treatment, adult, sexually naive male and female CBA x C57Bl/6 F1 hybrid mice received two series of daily tests that involved the presentation of different dilutions of urine from C57Bl/6 males followed by urine from estrous females. Each test session began with three consecutive presentations of deionized water (10 microl on filter paper for 2 min, behind a mesh barrier which prevented direct physical access, in the home cage at 1-min intervals) followed by three presentations of one of five different dilutions of urine (a different dilution on each test day). Males and females showed equivalent, significant habituation/dishabituation responses (low investigation times for successive water presentations; increased investigation of the first urine stimulus, followed by a decline in successive urine investigation times) to both male and female urine/water dilutions of 1:1, 1:10, and 1:20. However, only female mice responded reliably to 1:40 and 1:80 dilutions of both types of urine, pointing to a sex dimorphism in the detection and/or processing of biologically relevant, volatile urinary odors by the main olfactory system.

The ferret's vomeronasal organ and accessory olfactory bulb: effect of hormone manipulation in adult males and females.

The male ferret, a carnivore, was recently shown to possess a vomeronasal organ (VNO). We compared the morphology of the VNO and its associated accessory olfactory bulb (AOB) in male and female ferrets that were killed in adulthood. The volume and surface area of the VNO neuroepithelium were similar in adult gonadectomized male and female ferrets regardless of whether they were treated with testosterone propionate (TP) or oil vehicle. An AOB was localized bilaterally in the medial caudal part of the olfactory bulbs of adult ferrets using soybean agglutin binding and immunostaining for luteinizing hormone-releasing hormone and tyrosine hydroxylase as well as Nissl staining of coronal, horizontal, and sagittal brain sections. There was no effect of sex or TP treatment on AOB cell layer volume in adult gonadectomized animals. We found the ferret's AOB to be more medially located and much smaller than previously reported in this species, thus highlighting the importance of using several histochemical markers to characterize this structure in any previously unexamined species. Adult male and female ferrets both have a VNO and an associated AOB. More research is needed to determine what role, if any, this accessory olfactory system plays in mediating behavioral and neuroendocrine responses to pheromones in ferrets of either sex.

Nares occlusion eliminates heterosexual partner selection without disrupting coitus in ferrets of both sexes.

Using an airtight Y maze and a new method to induce peripheral anosmia in ferrets, we assessed the contribution of conspecific odors, either alone or in combination with visual and auditory signals, to heterosexual partner preference. Sexually naive ferrets were gonadectomized and treated with sex steroids, after which their nares were either bilaterally occluded using dental impression material or were sham-occluded. Behavioral and histological evidence suggested that nares occlusion blocked access of odors to the main olfactory epithelium for the duration of the study. Sham-occluded females and males preferred to approach odor only or odor plus visual plus auditory cues from opposite-sex conspecifics, whereas nares-occluded ferrets approached opposite- and same-sex cues equally. All ferrets subsequently mated successfully in tests conducted in a small chamber. When retested in the Y maze, sham-occluded females and males again preferred to approach odor-only or odor plus visual plus auditory cues from opposite-sex ferrets, whereas nares-occluded subjects showed no such preference even in tests when a brief physical interaction with tethered stimulus ferrets was allowed after each trial. Our results show that in the ferret, a carnivore, the detection and processing of volatile odors from conspecifics by the main olfactory system is required for heterosexual mate choice.

Maternal odours induce Fos in the main but not the accessory olfactory bulbs of neonatal male and female ferrets.

Previous research demonstrated that exposing gonadectomized adult ferrets to odours in oestrous female bedding induced nuclear Fos-immunoreactivity (Fos-IR; a marker of neuronal activity) in the main as opposed to the accessory olfactory system in a sexually dimorphic fashion, which was further augmented in both sexes by treatment with testosterone propionate. Ferrets are born in an altricial state and presumably use maternal odour cues to locate the nipples until the eyes open after postnatal (P) day 23. We investigated whether maternal odours augment neuronal Fos preferentially in the main versus accessory olfactory system of neonatal male and female ferret kits. Circulating testosterone levels peak in male ferrets on postnatal day P15, and mothers provide maximal anogenital stimulation (AGS) to males at this same age. Therefore, we assessed the ability of maternal odours to augment Fos-IR in the accessory olfactory bulb (AOB), the main olfactory bulb (MOB) and other forebrain regions of male and female ferret kits on P15 and investigated whether artificial AGS (provided with a paintbrush) would further enhance any effects of maternal odours. After separation from their mothers for 4 h, groups of male and female kits that were placed for 1.5 h with their anaesthetized mother had significantly more Fos-IR cells in the MOB granule cell layer and in the anterior-cortical amygdala, but not in the AOB cell layer, compared to control kits that were left on the heating pad. Artificial AGS failed to amplify these effects of maternal odours. Maternal odours (with or without concurrent AGS) failed to augment neuronal Fos-IR in medial amygdaloid and hypothalamic regions that are activated in adult ferrets by social odours. In neonatal ferrets of both sexes, as in adults, socially relevant odours are detected by the main olfactory epithelium and initially processed by the MOB and the anterior-cortical amygdala. In neonates, unlike adults, medial amygdaloid and hypothalamic neurones either do not respond to these inputs or respond in a manner that fails to induce Fos expression.

Sex difference and steroid modulation of pheromone-induced immediate early genes in the two zones of the mouse accessory olfactory system.

Two anatomically and neurochemically distinct zones within the vomeronasal organ (VNO) and accessory olfactory bulb (AOB) have been identified that are responsible for the detection of pheromones. Using markers to distinguish between apical and basal neurons of the VNO neuroepithelium and rostral versus caudal AOB glomeruli, we examined immediate early gene immunoreactivity (IEG-IR) in gonadectomized, steroid-treated mice in response to pheromones of male and female conspecifics. After exposure of estradiol-treated females to soiled male bedding, more VNO neurons in the basal than the apical layer exhibited IEG-IR compared with VNO neurons of estradiol-treated males. Conversely, whereas soiled female bedding failed to induce IEG-IR in VNO neurons of estradiol-treated males or females, both apical and basal neurons were activated in testosterone-treated males. Male and female pheromones also activated mitral and granule cells in the AOBs of all subjects, but responses to different pheromones were distributed across the boundary of the rostral and caudal regions. These data show that differences in the response of males and females to the same pheromonal stimulus are found in the sensory neurons of the VNO. We propose that centrifugal, noradrenergic inputs to VNO neurons, which may differ in the two sexes and respond differently to adult sex steroids, modulate sensitivity to pheromonal stimulation.

Mating induces gonadotropin-releasing hormone neuronal activation in anosmic female ferrets.

In females of both spontaneously and induced ovulating species, pheromones from male conspecifics can directly stimulate GnRH neuronal activity, thereby inducing pituitary LH secretion and stimulating the onset of estrus. However, whether pheromones contribute to the steroid- or mating-induced preovulatory activation of GnRH neurons is less clear. Previous studies in the ferret, an induced ovulator, raised the possibility that olfactory cues contribute to the ability of genital-somatosensory stimulation to activate GnRH neurons in the mediobasal hypothalamus (MBH). In the present study the percentage of GnRH neurons colabeled with Fos-immunoreactivity (IR), used as a marker for neuronal activation, was investigated in the MBH of mated gonadectomized, estradiol-treated female ferrets in which both nares were occluded. In addition, the percentage of GnRH neurons colabeled with Fos-IR was examined in the MBH of gonadectomized, estradiol-treated female ferrets exposed to male bedding. Bilateral nares occlusion successfully blocked mating or odor-induced increments in Fos-IR in central olfactory regions, including the cortical and medial amygdala. By contrast, the percentage of GnRH neurons expressing Fos-IR did not differ between mated nares- and sham-occluded females. Exposure to male bedding alone failed to induce Fos-IR in MBH GnRH neurons. Thus, the mating-induced preovulatory activation of GnRH neurons in the female ferret's MBH appears to rely solely on genital-somatosensory as opposed to olfactory inputs.

Central forebrain Fos responses to familiar male odours are attenuated in recently mated female mice.

Exposure of recently mated female mice to the urinary odours of an unfamiliar male blocks pregnancy (the Bruce effect). The absence of a pregnancy block in response to the stud male's familiar odours depends on an olfactory memory that is formed in the accessory olfactory bulb (AOB) in response to vomeronasal organ (VNO) inputs during mating. Sexually naive Balb/c female mice in pro-oestrus/oestrus were either placed onto soiled bedding ('bedding-only' females) from, or allowed to mate with, a Balb/c male ('recently mated' females). After 42 h, females were placed for 90 min onto clean bedding (controls) or onto soiled bedding from either a C57BL/6 male (unfamiliar bedding) or a Balb/c male (familiar bedding). Significant increases in Fos-immunoreactivity (Fos-IR, a marker of neuronal activation) occurred in the medial amygdala and the medial preoptic area (MPA) of 'bedding only' females exposed to either unfamiliar or familiar bedding and in 'recently mated' females exposed to unfamiliar bedding but not to familiar bedding. This suggests that a mating-induced memory prevents the later activation by the familiar stud male's odours of neurons in forebrain regions that receive inputs from the VNO--AOB. 'Bedding-only' females later exposed to either familiar or unfamiliar bedding had increased Fos-IR in the G alpha(o) protein-expressing basal zone of the VNO whereas no such effect occurred in 'recently mated' females. Familiar, as well as unfamiliar, male odours augmented Fos-IR in significantly more rostral than caudal AOB granule cells in all groups, with the effect being strongest in 'recently mated' females exposed to familiar male bedding. This outcome is consistent with the absence of odour-induced Fos-IR in forebrain regions of these females and, presumably, the absence of a pregnancy block.