Sexual and olfactory preference in noncopulating male rats

In some species including rats, mice, gerbils, and rams, apparently normal males fail to copulate when repeatedly tested with receptive females. These animals are called "noncopulators (NC)," and the cause of this behavioral deficit is unknown. It has been shown that NC rats do not have hormonal alterations or deficits in the mechanisms that control penile function. The present study was designed to examine (Experiment 1) whether NC male rats prefer receptive females to sexually active males. In addition, the olfactory preference for bedding soiled from estrous or for anestrous bedding was investigated. These tests were performed in NC and copulating (C) male rats when the subjects were intact, gonadectomized (GDX), or GDX and treated with high doses of testosterone propionate (TP). Our results demonstrate that NC rats do not display sexual behavior even after high TP treatment. While C male rats have a clear preference for receptive females as opposed to a sexually active male, NC rats do not. In all hormonal conditions, the preference shown by NC rats for estrous bedding was significantly reduced in comparison to that seen in C rats. TP treatment in NC rats did not modify either partner or odor preference. In Experiment 2, we evaluated if NC rats are feminized and if it could be easier to induce feminine-like behavior by hormone treatment with estradiol benzoate (EB) or with EB plus progesterone (P) (EB+P). Odor preference for estrous or male bedding under these hormonal conditions was also compared. No differences between NC and C rats were found in feminine sexual behavior. In the olfactory test, we found that NC rats prefer odors from receptive females as opposed to male odors, but this preference is reduced compared to that of C rats. Males treated with EB or EB+P show no preference for female odors. These results demonstrate that treatment with EB or EB+P does not increase feminine sexual behavior in NC rats.     

Effect of sex steroids and coital experience on ferrets' preference for the smell,sight and sound of conspecifics

Previous research showed that ferrets of both sexes recognize potential opposite-sex mates on the basis of volatile body odors. We compared the ability of estrogen and androgen treatments to activate a preference in gonadectomized male and female ferrets for distal cues (volatile body odors alone or volatile odors+sight+sounds) from male versus female stimulus ferrets using an airtight Y-maze and a "stimulus proximity" or a "discrete trials" testing paradigm. Sexually naive, gonadectomized male and female ferrets that received either testosterone propionate (TP) or estradiol benzoate (EB) spent equal time in proximity to goal boxes that provided either volatile odors alone or odors+sight+sounds of male and female stimulus animals. After they received coital experience, male and female subjects (treated with either EB or TP) showed a significant preference for both types of opposite-sex stimuli. When discrete trials tests were given to these ferrets prior to receiving coital experience, EB-treated females preferred to approach odor only cues, as well as odors+sight+sounds of stimulus males, and this preference was further strengthened after coital experience. Sexually naive, TP-treated males preferred to approach volatile odor cues from stimulus females; however, these animals showed an equal preference for odors+sight+sounds of stimulus females and males. Again, after coital experience, males' preference for both sets of cues from stimulus females was significantly enhanced. Thus, in sexually naive ferrets, discrete trials, but not stimulus proximity tests, revealed a preference for distal cues (body odors with or without concurrent sight and sounds) from opposite-sex conspecifics in subjects of both sexes. Coital experience significantly enhanced these preferences for heterosexual distal cues under both testing paradigms.     

Responses of male mice to odors of females: Effects of T- and H-2-locus genotype

Male wild house mice (Mus musculus) were given a choice of odors of females whose T-locus genotype was +/ + or +/t. Males showed strong preferences for the odors of +/ + females. However, when males were tested with odors of recombinant females whose genotype differed at the T locus but which carried similar haplotypes at the H-2 locus, the preference for odors of +/+ females was not manifested. Consequently, differences in female odor production that are responsible for male odor preference are not due specifically to the female genotype at the T locus.This work was partially supported by NIH Grant HD R01 1997 and a BRSG Grant from Rutgers University.     

Attraction of beagles to conspecific urine, vaginal and anal sac secretion odors

Attraction of male and female Beagles to conspecific urine, vaginal and anal sac secretion odors was examined in four experiments. Males spent more relative time investigating female urine odors than odors of vaginal or anal sac secretions. Sexually experienced males, but not sexually inexperienced ones, spent more time investigating estrous than diestrous female urine and vaginal odors. Anal sac secretions from estrous bitches were not more attractive to males than those from diestrous bitches. Estrous females spent no more time than diestrous ones in the investigation of male anal sac secretion and urine odors. Male urine and anal sac secretions elicited little investigation from male conspecifics. Females spent more time investigating female urine odors than female anal sac or vaginal secretion odors, and exhibited a slight general preference for diestrous over estrous stimuli. A positive correlation between the odor investigation times of this study and investigation times of comparable animals to conspecifics in a social situation suggests odor preferences are relatively good indicators of social preferences, and vice versa, in this breed.     

Preferences of pre- and post-weanling long-evans rats for nest odors

Three experiments were conducted on the preferences of infant rats for nest odors. Pre-weanling rats preferred their own nest odor to nests of strange litters and showed low preferences for the nest odors of virgin females. Nest odors of unmated males were investigated less than the pup's own nest odor but more than the odors of virgin females. Male and female rats did not differ in odor preferences. Rats reared with both their dam and sire did not differ in their odor preferences from rats reared with their dam alone when tested at 16–20 days of age, but when tested at 33–38 days of age they showed a greater preference for male odors. Post-weanling rats did not show a preference for the nest odors of lactating females and began to show sex differences in their preferences.     

Ovarian hormones influence odor stimulated flank marking behavior in the hamster (Mesocricetus auratus)

The effect of ovarian hormones on the frequency of female flank marking in response to the odors of male hamsters was examined. The amount of flank marking differed significantly (p less than 0.01) among the days of the estrous cycle, with the lowest levels of flank marking occurring on estrus. Ovariectomy (OVX) eliminated the 4 day rhythm, and significantly (p less than 0.01) reduced the total amount of flank marking. Implantation of Silastic capsules containing estradiol benzoate (EB) in OVX hamsters increased flank marking by approximately two-fold when compared to hamsters implanted with blank Silastic capsules. In a second series of experiments, OVX hamsters given EB flank marked significantly (p less than 0.01) more than those with blank capsules. Subcutaneous injection of progesterone (P) into OVX-EB treated hamsters 6 hr prior to testing significantly reduced flank marking when compared to OVX-EB treated hamsters given oil. No differences were observed in flank marking between groups of OVX hamsters with blank capsules and injected with P or oil. In summary, these data indicate that the four day rhythm of odor stimulated flank marking in intact female hamsters may be the result of the 4 day fluctuations of estrogen and progesterone that occur during the estrous cycle.     

The role of the hippocampal system in social odor discrimination and scent-marking in female golden hamsters (Mesocricetus auratus)

Female golden hamsters (Mesocricetus auratus) received aspiration lesions of the parahippocampal region (PARA) or electrolytic lesions of the fimbria-fornix (FNX) and were tested for their (a) discrimination between odors of individual males in a habituation-discrimination task, (b) preference for male over female odors, and (c) scent-marking in response to conspecific odors. Both lesion groups habituated to repeated presentations of a male's odor. However, only FNX females discriminated between scents of individual males, whereas PARA females did not. Neither lesion eliminated female preferences for male odors. Females with FNX lesions showed decreased levels of scent marking, but those with PARA lesions had more subtle deficits. Thus, the PARA, but not the subcortical connections of the hippocampus, is critical for discrimination of the odors of individuals.     

Lesions centered on the medial amygdala impair scent-marking and sex-odor recognition but spare discrimination of individual odors in female golden hamsters.

The medial amygdala (Me) has been implicated in various social behaviors that depend on chemosensory cues, but its precise role in discriminating and learning social odors is not known. Female golden hamsters (Mesocricetus auratus) received electrolytic lesions of the Me or sham surgery and were tested for their ability to (a) discriminate between odors of individual males in a habituation-discrimination task, (b) show preferences for male over female odors in a Y maze, and (c) scent-mark in response to male and female odors. All females discriminated between scents of individual males. In contrast, Me lesions eliminated female preferences for male odors in a Y maze. Females with Me lesions also showed a substantial reduction in vaginal marking and virtually no flank marking in response to odors. Thus, the Me in female hamsters is critical for differential investigation of opposite-sex odors and for scent-marking behavior but is not involved in discrimination between odors of individuals.     

Sex differences in the response to neonatal steroid treatment in the Mongolian gerbil

The present study examines whether genetic male and female gerbils differ in response to neonatal steroid treatment as evidenced by the display of sexually dimorphic territorial marking behavior in adulthood. The advantage of this behavioral endpoint is that male and female marking differs only in frequency, permitting direct between-sex comparison of neonatal treatment effects. Females and day-2 castrated males received a single SC injection of 10 to 100 μg testosterone propionate (TP) or estradiol benzoate (EB) between birth and day-14 postpartum. Between 7 and 9 months of age, the territorial marking behavior response to exogenous TP was determined in all animals. Based on marking frequency, the time of greatest sensitivity to TP administered during the critical neonatal period occurred earlier in the male (days 3–5) than in the female (days 5–7). In addition females consistently showed greater sensitivity to given dose of either TP or EB than did males. EB was more effective than TP at the 10 μg dose in both sexes. These results indicate that sex differences exist in timing and steroid sensitivity in the sexual differentiation of the neural substrate regulating territorial marking behavior.     

Blocking oxytocin receptors inhibits vaginal marking to male odors in female Syrian hamsters

In Syrian hamsters (Mesocricetus auratus), precopulatory behaviors such as vaginal scent marking are essential for attracting a suitable mate. Vaginal marking is dependent on forebrain areas implicated in the neural regulation of reproductive behaviors in rodents, including the medial preoptic/anterior hypothalamus (MPOA-AH). Within MPOA-AH, the neuropeptide oxytocin (OT) acts to facilitate copulation (lordosis), as well as ultrasonic vocalizations towards males. It is not known, however, if OT in this area also facilitates vaginal marking. In the present study, a specific oxytocin receptor antagonist (OTA) was injected into MPOA-AH of intact female Syrian hamsters to determine if oxytocin receptor-dependent signaling is critical for the normal expression of vaginal marking elicited by male, female, and clean odors. OTA injections significantly inhibited vaginal marking in response to male odors compared with vehicle injections. There was no effect of OTA on marking in response to either female or clean odors. When injected into the bed nucleus of the stria terminalis (BNST), a nearby region to MPOA-AH, OTA was equally effective in decreasing marking. Finally, the effects of OTA appear to be specific to vaginal marking, as OTA injections in MPOA-AH or BNST did not alter general locomotor activity, flank marking, or social odor investigation. Considered together, these results suggest that OT in MPOA-AH and/or BNST normally facilitates male odor-induced vaginal marking, providing further evidence that OT generally supports prosocial interactions among conspecifics.     

Effects of vomeronasal organ removal on individual odor discrimination, sex-odor preference, and scent marking by female hamsters

Removal of the vomeronasal organ (VNX) did not eliminate the ability of female hamsters to discriminate between individual male's flank gland or urine odors in a habituation/discrimination task nor did it impair preference for male odors over female odors from a distance. Vomeronasal organ removal did reduce overall levels of investigation of flank gland odor in the habituation/discrimination task. Although VNX females did not show severe impairments in the frequency of either flank or vaginal marking in response to odors, they did show an abnormal pattern of marking. VNX females, unlike shams, did not flank mark more to female odors than to male odors, nor did they vaginal mark more to male odors than to female odors. Thus, the vomeronasal organ in female hamsters appears to be important for differences in scent marking toward male and female odors, but is not essential for discrimination of individual odors or for preferences for male over female odors.     

Photoperiod and gonadal hormones influence odor preferences of the male meadow vole, Microtus pennsylvanicus.

Male meadow voles housed in a long photoperiod (14 h light/day, LP) preferred female to male odors, whereas males maintained in a short photoperiod (10 h light/day, SP) did not display preferences for odors of either sex. These odor-preference patterns matched those of free-living males during spring and autumn, respectively. The preference of LP male voles for female over male odors was eliminated by gonadectomy and reinstated by treatment with testosterone. In SP males, although gonadectomy did not affect odor choices, a preference for female odors was induced by testosterone treatment. Treatment with estradiol did not alter odor preferences of LP or SP males. In conjunction with previous result, the present findings suggest that hormonal responsiveness of neural substrates that control odor preferences are sexually dimorphic and may reflect sex differences in reproductive strategies.     

Olfactory experience and the development of odor preference and vaginal marking in female Syrian hamsters

Rodent reproductive behavior relies heavily on odor processing, and evidence suggests that many odor-guided sexual behaviors are shaped by prior experience. We sought to determine if exposure to male odors during development is required for the adult expression of proceptive sexual behavior toward male odors in female Syrian hamsters. Exposure to male odors was restricted in na?ve subjects by removing all male siblings from the litter at three to five days of age. Control litters were also culled, but included equal numbers of male and female pups. As adults, na?ve females displayed investigatory preferences toward male odors in a Y-maze that were comparable to control females; this preference was observed whether contact with the odor stimuli was prevented of allowed. In contrast, na?ve females vaginal scent-marked equally toward male and female volatile odors, suggesting an inability to target behavior toward sexually relevant odors. However, na?ve females marked preferentially toward male odors when allowed to contact the odor stimuli. These results provide evidence for the experience-dependent development of vaginal marking behavior toward volatile components of sexual odors. Furthermore, they suggest that distinct mechanisms regulate the development of odor preferences and vaginal marking behavior in this species.     

Effect of prenatal androgen receptor antagonist or aromatase inhibitor on sexual behavior, partner preference and neuronal Fos responses to estrous female odors in the rat accessory olfactory system

Many socially relevant odors are detected in rodent species by the vomeronasal organ and subsequently processed by the accessory olfactory system (AOS). We previously found that gonadectomized male and female rats treated in adulthood with testosterone propionate (TP) showed equivalent Fos responses in the AOS to odors derived from estrous females. Likewise, in contrast with numerous other mammalian species, gonadectomized female rats show surprisingly high levels of male-typical mounting behavior in response to adult TP. We tested the hypothesis that prenatal testosterone (T) exposure, acting via androgen receptors (ARs) or via estrogen receptors, masculinizes the AOS in rats of both sexes. Pregnant dams were treated with either the AR blocker, Flutamide, the aromatase inhibitor, 1,4,6-androstatriene-3,17-dione (ATD), or nothing (control) to assess the role of prenatal androgen and estradiol receptor activation, respectively, in this masculinization. Beginning at birth, male and female offspring were injected subcutaneously (sc) every other day with either ATD (pre- and neonatal ATD group) or oil vehicle (Flutamide and control groups) until postnatal Day 12. Subjects were gonadectomized as adults, hormonally treated and tested for different behaviors before having their AOS Fos responses to estrous female odors assessed. Prenatal treatment with Flutamide (but not ATD) significantly decreased anogenital distance and severely impaired intromissive and ejaculatory behaviors in males tested after TP replacement without disrupting mounting capacity in either sex. Pre- and neonatal treatment with ATD (but not Flutamide) enhanced lordosis responsiveness in males tested after sc injections of estradiol and progesterone, whereas these perinatal treatments had no effect on any aspect of masculine coital performance in either sex. After TP treatment, male and female control subjects preferred to approach a tethered stimulus female as opposed to a male, and prenatal Flutamide or perinatal ATD treatments did not modify this pattern of partner preference. Neuronal Fos responses to estrous odors were (as in previous studies) identical in the AOS of gonadectomized TP-treated control males and females. Prenatal Flutamide or perinatal ATD treatments failed to disrupt consistently this profile of Fos responses to estrous odors in the AOS of rats of either sex. These behavioral and neuroanatomical findings raise the possibility that the similar level of male-typical responsiveness to social odors that occurs in male and female rats after adult TP treatment results from nonsteroid-hormone-dependent, species-specific factors that act perinatally in the brains of rats of both sexes.     

Suppression of adult copulatory behaviors following LiCl-induced aversive contingencies in juvenile male rats

Sexually immature juvenile male rats received an injection of .3 M (20 ml/kg) lithium chloride (LiCl) during each of 8-10 pairings (spaced at 2-3-day intervals) with an estrous female. Approximately seven weeks later, these males displayed fewer copulatory behaviors in pairings with estrous females than saline control rats. Noncontingent .3 M LiCl injections administered to juvenile males did not affect adult copulatory behaviors. Males that were either group or individually housed during the retention interval displayed comparable suppression, which suggests that copulatory suppression does not represent a generalized aversion to social interactions with another animal. Males housed during the retention interval in an environment that contained the odors of estrous females also displayed suppression comparable to that of males housed in the absence of such odors. This suggests that the associations that mediate copulatory suppression are not elicited solely by the odor cues of estrous females. These data demonstrate that LiCl-induced aversive contingencies in juvenile male rats subsequently suppress adult copulatory behaviors.     

Effect of flutamide (an antiandrogen) and diethylstilbestrol on the reproductive behavior of Japanese quail

Three experiments were conducted in order to investigate the role of brain androgen and estrogen receptors in sex hormone activated male reproductive behavior in Japanese quail. In Experiment 1, castrated male quail were injected with oil, testosterone propionate (TP), flutamide (FLUT), an androgen antagonist, or TP+FLUT. Males given TP+FLUT, compared with birds receiving TP alone, strutted much less and had smaller proctodeal (foam) glands. Copulation was reduced by FLUT only on the last test day and only on one measure (number of head grabs + mounts). These results suggest that binding of testosterone or one of its metabolites to an androgen receptor is part of the mechanism of TP activated strutting, and therefore that central nervous system androgen receptors are involved in a male reproductive behavior pattern. In Experiment 2, castrated male quail were injected with oil, with 50 micrograms estradiol benzoate (EB), or with 25, 50 or 100 micrograms diethylstilbestrol (DES), a synthetic estrogen that does not bind to androgen receptors. EB but not DES activated copulation to a significant extent. In Experiment 3 male and female quail with photically regressed gonads were given intraperitoneal Silastic implants of DES, estradiol (E) or cholesterol. DES was highly effective at activating male-typical copulation in males and receptivity in both sexes. Thus hormonal interaction with estrogen receptors alone is sufficient for the activation of male-typical as well as female-typical copulatory behavior in this species.     

Ejaculatory and postejaculatory behavior of male and female rats: effects of sex hormones and electric shock

Ejaculatory patterns were observed in normally reared, postpuberally castrated male and female rats treated with sex hormones and electrical shock in adulthood. In a preliminary experiment, 3 females treated with 50 microgram estradiol benzoate (EB) and 500 microgram progesterone (P) showed intromission and ejaculatory behavior when subjected to shock. Refractory periods were abnormally short and there was almost no postejaculatory vocalization. In Experiment 1, males and females were tested with electrical shock following daily treatment with 32 microgram EB with and without 500 microgram P on the test day. There was no difference between males and females in preejaculatory behavior, but females displayed abbreviated refractory periods and no postejaculatory vocalization. Progesterone had no observed effect. In Experiment 2 castrated males and females were subjected to shock after treatment with 8 microgram EB per day for 3 weeks. No P was given. Again females showed drastically reduced refractory periods and little vocalization. In Experiment 3, males and females treated with TP and shock displayed ejaculatory patterns, normal refractory periods and vocalization. Results show that female rats are capable of exhibiting the ejaculatory response without sex hormone treatment in perinatal life or androgen treatment in adulthood. It was also demonstrated that there is a sex difference in the postejaculatory behavior shown by estrogen-treated male and female rats.     

The role of estrogen and progesterone in the control of preputial gland sex attractant odors in the female rat

Sexually experienced male rats were used to test the attractiveness of body odors of female rats. The attractiveness of these odors varied with the estrous cycle. Odors from female rats in proestrus were the most attractive to male rats and those from female rats during the darkness hours of diestrus the least attractive. The preputial glands appeared to be the source of these odors for the male rats showed no preference for the odors of proestrous female rats that had been preputialectomized. Administration of 1 μg estrdiol benzoate (EB) for 5 days increased the attractiveness of body odors of ovariectomized rats. A higher dose of EB (5 μg) had the same effect when administered for 1 or 5 days although the increase that occurred after 3 days was not significant. A single dose of progesterone (P) (500 μg) on the other hand, decreased the attractiveness of ovariectomized female odors although no change was seen after 3 days of treatment. A single injection of P also decreased the attractiveness of odors of ovariectomized females that had received EB for 3 days. However, P failed to decrease the attractiveness of odors in ovariectomized females after preputialectomy. We conclude that the preputial glands are an important source of sex attractant odors in the female rat and that the changes in the release of these odors that occur throughout the estrous cycle and pregnancy are controlled by ovarian steroids. While estrogen acts to stimulate the production and release of these odors P appears to inhibit their release.     

Female rats prefer an area vacated by a high testosterone male

In each of 4 experiments, female rats were observed as they moved freely about an apparatus that had recently housed 2 males. The androgen levels of the males were manipulated with castrations and exogenous injections of testosterone propionate (TP). For the most part, female exploration times in the areas vacated by the 2 males did not differ. The urinary markings over the urine from the males, however, did differ. The female exhibited a decided marking preference for the area that had housed the male with higher titers of androgens. The preferences were for intact vis-a-vis castrated males, though injections of TP restored the castrates' attractiveness to the females. When both males possessed reproductive levels of TP, the females preferred to mark over the urine of the male with the higher levels of the androgen. The reproductive advantage for the female may be the choice of a mate with more viable sperm, because androgens are intimately involved in spermatogenesis.     

Odor modulation of ventral scent gland marking in the Mongolian gerbil

Gerbil odors were allowed to accumulate in individual home cages for several months in order to provide stable olfactory stimuli that might modulate the number of ventral scent gland marks produced by both male and female Mongolian gerbils. Male marking levels for most test conditions were 2-3 times larger than those in females. In general, the relative patterns of responsiveness to particular odors were similar in both sexes. The number of scent marks in home cages was significantly smaller than the number obtained in clean cages. This suppression was not strongly dependent on the odors emanating from an individual's own ventral gland sebum. Marking levels in the home cages of other gerbils were substantially higher than those seen in the test animal's own cage and were maximal in the home cages of other males. Behaviorally receptive females marked at significantly higher levels in home, clean, and conspecific cages than did nonreceptive females. Conspecific urine odors, especially those from females, increased marking levels significantly over those obtained in home cages. In males, home cage marking was significantly enhanced by the presence of a female. Females, on the other hand, marked at similar levels in home cages whether a potential mating partner was present or not. Both sexes showed significant reductions in marking levels in another's cage if the resident animal was present. These results suggest that when sufficient gerbil-related odors are available to a test animal, its rate of scent-marking is altered, depending on the particular odor environment. Some gerbil-related odors increase marking levels, whereas others decrease marking. Scent-marking then, like other olfactory guided behaviors and odor preferences, is context specific.