Two experiments examined probiotic pretreatment (Lactobacillus rhamnosus GG) on obsessive–compulsive disorder (OCD)-like behavior induction by RU 24969 in BALB/cJ house mice. In the first experiment, two groups were defined by their daily pretreatment by oral gavage of either (a) L. rhamnosus (1×109 CFU/day) or (b) the saline vehicle. Both a 2- and 4-week probiotic pretreatment attenuated OCD-like behavior induction (increased perseverative open-field locomotion, stereotypic turning, and marble burying) relative to saline pretreatment. Experiment 2 re-examined the 2-week probiotic pretreatment while also comparing it to a 4-week fluoxetine pretreatment. Again, groups were defined by daily pretreatment of either (a) L. rhamnosus for 2 weeks, (b) the saline vehicle for 2 weeks, or (c) fluoxetine (10 mg/kg) for 4 weeks. Pretreatment by either L. rhamnosus or fluoxetine blocked the induction of OCD-like behavior compared with saline pretreatment. Thus the 2-week probiotic pretreatment was again effective. Although side effects of fluoxetine or L. rhamnosus on androgen-dependent behaviors could not be demonstrated, L. rhamnosus treatment appeared comparable to fluoxetine treatment in attenuating mouse OCD-like behaviors.

Parental investment theory states that parents should contribute more to older offspring. Differences between the sexes also influence how each parent contributes to offspring in biparental species. Here, we examined a naturally occurring population of biparental convict cichlids in Costa Rica to determine how each parent cared for offspring during two distinct offspring development stages. Consistent with the predictions of the reproductive value hypothesis, we hypothesized that the levels of parental contribution would be relative to the value that each parent places on a brood. We predicted that female parents would contribute more than male parents because female convict cichlids have lower future reproductive success than males. Additionally, we predicted that both parents should contribute more to older offspring, either due to the young’s increased susceptibility to predation (i.e., the vulnerability hypothesis) or because of the longer period of time parents have been interacting with older offspring (i.e., feedback hypotheses). This increase in investment by males should coincide with a change in the coordination of care between parents. Detailed observations of parental pairs in their natural habitat supported these predictions. Females contributed more to broods than males and were relatively unaffected by offspring age while males spent significantly more time with older, free-swimming fry. Additionally, males tended to leave younger offspring more than females did, and were more likely to do so consecutively with younger offspring. This suggests that the coordination of duties between parents changes as parental investment changes. Overall, these data support both the reproductive value and the vulnerability hypotheses, but not necessarily the feedback hypothesis.

Ghrelin is a peptide hormone released by the stomach that stimulates hunger. Ghrelin also suppresses reproductive physiology by inhibiting the HPG axis. However, to our knowledge, our results are the first to demonstrate ghrelin's quick suppression of sex-hormone-regulated behaviors. In experiment 1, 2 orexigenic i.p. ghrelin injections (0.165 mg/kg and 0.33 mg/kg) suppressed male courtship behavior (ultrasonic calling to a female) and intermale aggression (latency to attack a stimulus male) 20 min following administration. Experiment 2 (examining only the 0.33 mg/kg dose ) replicated ghrelin's suppression of ultrasonic calling and intermale aggression; however, a third behavior, preference for volatile female odors (20 min following administration), was not significantly inhibited. In experiment 2, ghrelin treatment did not affect general locomotor activity (distance traveled 20 min following injection) or seminal vesicle weight (measured 5 days after completing ghrelin injections). We hypothesize that ghrelin's quick suppression of male aggression and ultrasonic mating calls was mediated through its effects on the brain (rather than indirectly through inhibition of the HPG axis).

An adult male house mouse typically begins calling at ultrasonic frequencies when presented with either a female mouse or her odors. Although direct experimental evidence has been difficult to obtain, ethological, genetic, endocrine, neurological, sound-spectrographic and perceptual findings all indirectly support male calling conveying important information to the female during sexual encounters. Since male calls correlate with the male's immediate sexual arousal and are also attractive to the female, these calls could help coordinate aspects of a mating bout. As might be expected of a reproductive behavior, male calling is hormonally and pheromonally regulated, similarly to other aspects of male reproductive physiology and behavior. Evidence is reviewed that androgen exerts a permissive activational effect during adulthood through its actions in the same brain area (the medial preoptic area) that regulates male-typical copulatory behavior. This activational effect also accounts for the sex difference in mouse calling. Evidence is also reviewed that male calling is pheromonally elicited by chemosignals in female urine.

Male mammals of many species exhibit reflexive testosterone release in mating situations. In house mice (Mus musculus), the dramatic robustness of such release, occurring primarily in response to a novel female, suggests some function. The resulting testosterone elevations typically peak during copulatory behavior and may serve to activate transitory motivational and physiological responses that facilitate reproduction. However, such a function requires that testosterone be working through either nongenomic, or very quick genomic, mechanisms. The first part of the review describes reflexive sex hormone release in house mice. The second part summarizes research implicating testosterone’s fast actions in affecting anxiety, reward, learning, analgesia, and penile reflexes in rodents, all of which could optimize male mating success. The review concludes with a speculative model of how spontaneous and reflexive hormone release might interact to regulate reproductive behavior and why mice appear to be an ideal species for examining testosterone’s quick effects.

In virtually every mammalian species examined, some males exhibit reflexive testosterone release upon encountering a novel female (or female-related stimulus). At the same time, not every individual male (or every published study) provides evidence for reflexive testosterone release. Four experiments using house mice (Mus musculus) examined the hypothesis that both the male's genotype and his degree of sexual arousal (as indexed by ultrasonic mating calls) are related to such variability. In Experiment 1, CF-1 males exhibited reflexive testosterone elevations 30 min after encountering female urine. CK males, on the other hand, did not exhibit testosterone elevations 20, 30, 50, 60, or 80 min after encountering female urine (Experiments 1 and 2) suggesting this strain incapable of reflexive release. In Experiment 3, we measured both mating calls and reflexive testosterone release in response to female urine in CF-1 and CK males. Most males of both strains called vigorously to female urine but not to water. But, only CF-1 males exhibited significant testosterone elevations to female urine. In Experiment 4, DBA/2J males called vigorously to females followed by testosterone elevations 30 min later. The first 3 experiments support the hypothesis that male genotype is an important variable underlying mammalian reflexive testosterone release. Statistically significant correlations between mating calls in the first minute after stimulus exposure and testosterone elevations 30 min later (Experiments 3 and 4) support the hypothesis that, in capable males, reflexive testosterone release is related to the male's initial sexual arousal.

Male mammals reflexively release an endogenous pulse of testosterone in response to either a female or her urinary pheromones. Two experiments examined the hypothesis that such pulses have quick-acting effects upon the expression of reproductive behavior in male house mice. In Experiment 1, 30 min after exposure to female urine, males exhibited significantly reduced latency to mount a receptive female (when they should have been expressing an endogenous testosterone pulse). In Experiment 2, gonadally intact males received a simulated testosterone pulse via a subcutaneous injection of 500 μg of testosterone propionate. At 60 min after injection, males mounted a receptive female significantly more quickly than if they had not received such an injection. These experiments provide evidence that elevations in testosterone titers above baseline can, under certain conditions, rapidly alter the expression of male-typical behaviors.

Eight experiments supported the hypotheses that reflexive testosterone release by male mice during sexual encounters reduces male anxiety (operationally defined in terms of behavior on an elevated plus-maze) and that this anxiolysis is mediated by the conversion of testosterone to neurosteroids that interact with GABAA receptors. In Experiment 1, a 10-min exposure to opposite-sex conspecifics significantly reduced both male and female anxiety 20 min later (as indexed by increased open-arm time on an elevated plus-maze) compared to control mice not receiving this exposure. In contrast, locomotor activity (as indexed by enclosed-arm entries on the elevated plus-maze) was not significantly affected. The remaining experiments examined only male behavior. In Experiment 2, exposure to female urine alone was anxiolytic while locomotor activity was not significantly affected. Thus, urinary pheromones of female mice likely initiated the events leading to the male anxiolysis. In phase 1 of Experiment 3, sc injections of 500 μg of testosterone significantly reduced anxiety 30 min later while locomotor activity was not significantly affected. Thus, testosterone elevations were associated with reduced male anxiety and the time course consistent with a nongenomic, or very rapid genomic, mechanism of testosterone action. In phase 2 of Experiment 3, the anxiolytic effect of testosterone was dose dependent with a 250 μg sc injection required. Thus, testosterone levels likely must be well above baseline levels (i.e., in the range induced by pulsatile release) in order to induce anxiolysis. In Experiment 4, a high dosage of 5α-dihydrotestosterone was more anxiolytic than a high dosage of estradiol benzoate, suggesting that testosterone action may require 5α-reduction. In Experiments 5 and 6, 3α,5α-reduced neurosteroid metabolites of testosterone (androsterone and 3α-androstandione) were both anxiolytic at a lower dosage (100 μg/sc injection) than testosterone, supporting the notion that testosterone is converted into neurosteroid metabolites for anxiolytic activity. Experiments 7 and 8 found that either picrotoxin or bicucculine, noncompetitive and competitive antagonists of the GABAA receptor, respectively, blocked the anxiolytic effects of testosterone. However, conclusions from these 2 experiments must be tempered by the reduction in locomotor activity that was also seen. The possible brain locations of testosterone action as well as the possible adaptive significance of this anxiolytic response are discussed.

This experiment examined whether testosterone proprionate (T) action in the medial preoptic area (MPO) would synergize with T action in the medial nucleus of the amygdala (AME) for the expression of androgen‐dependent behaviors in house mice. Cannulae containing T were bilaterally implanted into the MPO, the AME, or both areas concurrently (MPO/AME) of castrated males. In addition, other castrates were implanted subcutaneously with empty Silastic capsules (BSIL) or Silastic capsules containing T (TSIL). All subjects were examined for the following androgen‐dependent, male‐typical behaviors: mounting, urinary scent marking, preference for female urine over male urine, preference for female over male conspecifics and ultrasonic mating vocalizations. MPO implants restored ultrasonic vocalizations and preference for females, but had little or no effect upon urine marking, mounting or preference for female urine. In contrast, AME implants were ineffective at restoring any of these male‐typical behaviors. The combined MPO/AME implants were not more effective in restoring male‐typical behaviors than MPO implants alone, thus providing no evidence for synergy in hormone action between these two brain areas. In general, castration (BSIL) resulted in low levels of all behaviors whereas systemic T replacement (TSIL) resulted in high levels of behavior, verifying the androgen‐dependence of these behaviors. Group differences in male‐typical behavior could not be accounted for by differences in general activity levels. Moreover, none of the brain‐implanted groups had larger seminal vesicles than those of the BSIL. Thus, when the brain implants affected behavior, they most probably did so through their effects within the brain. Although the AME is a target for steroid hormones and is an important area for the expression of male‐typical behaviors, intracranial T implants into the AME did not demonstrate a role for androgen in the AME in restoring male‐typical behaviors in castrated mice.

White, N. R., M. Prasad, R. J. Barfield and J. G. Nyby. 40- and 70-kHz vocalizations of mice (Mus musculus) during copulation. Physiol Behav 63(4) 467–473, 1998. Ultrasonic vocalizations were tape recorded from five pairs of copulating mice and subjected to spectrographic analysis. As expected, the mice emitted numerous 70-kHz vocalizations. At the beginning of the test, before copulation began, 70-kHz calls were emitted almost continuously, while calls with lower spectrographic frequencies were not observed. Subsequently, bursts of 70-kHz calling generally began shortly before mounts and intromissions and persisted until dismount. Intermixed with these 70-kHz calls were additional vocalizations of about 40 kHz. Calling rates were highest just prior to intromission. Once intromissions began, 70-kHz calls continued at a lower rate until dismount; however, 40-kHz calls occurred infrequently. In a second experiment, the male was found to emit the majority of the 70-kHz calls and all of the 40-kHz calls. When the male was devocalized, few calls were detected, regardless of whether the female was able to call. If the male was not devocalized, high rates of calling were detected, even if the female was devocalized.

Cannulae containing testosterone proprionate (T) were bilaterally implanted into the medial preoptic area (MPO), the ventral tegmental area (VTA), or both areas concurrently (MPO/VTA) of castrated male house mice. A fourth group of castrates in which the intracranial implants missed their targets (MIMP) served as controls. In addition, other castrates were implanted subcutaneously with empty silastic capsules (BSIL) or silastic capsules containing T (TSIL). All subjects were examined for the following male-typical behaviors: mounting, attraction to female urine, ultrasonic mating vocalizations, and urinary scent marking. In addition, the males were tested for activity levels to insure that they were not motorically impaired. In general, TSIL implants restored all male-typical behaviors to normal levels, whereas BSIL and MIMP implants were generally ineffective. Similar to previous findings, MPO implants alone completely restored ultrasonic vocalizations, partially restored urine marking, and had little or no effect upon mounting or urine preference. In contrast, VTA implants alone were ineffective at restoring any of these male-typical behaviors. However, the combined MPO/VTA implants were the most effective in restoring male-typical behaviors. In fact, a synergism between concurrent hormone action in the MPO and VTA was seen for mounting and urine preference. We interpret these data to indicate that androgen may act simultaneously in the MPO and VTA for more complete expression of some male-typical reproductive behaviors.

Two experiments examined the chemosensory modalities by which males detect an ephemeral sex pheromone in the freshly voided urine of female mice. Experiment 1 examined the interaction of deafferenting the accessory olfactory system (vomeronasal organ removal) and subsequent sexual experience upon ultrasonic vocalizations by male mice to freshly voided female urine. In general, sexually experienced males vocalized substantially more than sexually naive males. In addition, males possessing a vomeronasal organ vocalized slightly more than those without. Nonetheless, a functioning vomeronasal organ clearly was not essential for vocalizing to fresh female urine. Experiment 2 examined the effects of deafferenting the main olfactory system (ZnSO4 nasal irrigation) and/or the accessory olfactory system (vomeronasal removal) in sexually experienced males. Males with both olfactory systems functioning vocalized at high levels to fresh urine, while males with only one functioning system vocalized at intermediate levels. Males with neither system functioning did not vocalize at all to fresh urine. In contrast, when female mice themselves served as stimuli, all groups of males vocalized at high levels. We conclude that adult male mice can detect the ephemeral pheromone via either the main olfactory system or the accessory olfactory system. However, vocalizations to the female herself can be mediated by other sensory systems as well.

At least two pheromones exist in female house mouse urine that elicit 70-kHz ultrasonic mating vocalizations from males: (1) a potent albeit ephemeral pheromone found in freshly voided urine which disappears within 15-18h and (2) a more persistent, but less potent pheromone that remains for at least 30 days. Three experiments examined whether male responsiveness to the ephemeral pheromone and its loss in aged urine were due to volatility or oxidation. Experiment 1 demonstrated that both sexually experienced and inexperienced males must physically contact fresh urine before vocalizing. Experiment 2 indicated that storing freshly voided urine in an air-tight syringe does not attenuate the loss of the ephemeral pheromone. Experiment 3 demonstrated that the loss of activity of the ephemeral pheromone can be prevented with either β-mercaptoethanol or dithiotreitol. Thus, the ephemeral pheromone is non-volatile and appears to be degraded by oxidation.

Castrated male mice were bilaterally implanted with 27 ga cannulae containing testosterone into either the septum, medial preoptic area (MPO), or corticomedial amygdala. One additional group of castrates received no hormone and another received only systemic testosterone via subcutaneous silastic capsules. All males were subsequently tested for ultrasonic mating vocalizations, urine marking, mounting behavior, aggression and gender preference, all of which are androgen-dependent, male-typical behaviors. In general castrates receiving no hormone performed these behaviors at low levels and animals receiving systemic testosterone performed the behaviors at normal male-typical levels. Ultrasonic vocalizations in response to female urine were activated by MPO implants. Urine marking in response to female urine appeared to be partially activated only with MPO implants. Very little mounting or fighting were seen in the brain implanted groups. Gender preference (for females over males) was restored with MPO implants and appeared to be partially activated with septal implants. The seminal vesicles of the castrates receiving brain implants were not significantly different from those receiving no hormone indicating that little or no implanted hormone was exiting the brain into general circulation. The implications of these findings for the neuroanatomy of sexual motivation and performance are discussed.

Male mice will emit ultrasounds to at least two chemosignals present in female mouse urine: a) an ephemeral unconditioned stimulus (UCS) present in freshly voided urine and b) a conditioned stimulus (CS) that remains after the UCS has disappeared. In this study, four phases examined the time frame over which the UCS disappeared. Urine that had been aged for either 12, 15, 18, or 24 h was compared for vocalization elicitation with fresh urine (which contains the UCS) and metabolic cage-collected urine ( which does not). Under the conditions utilized, significant pheromone fade-out occurred between 15 and 18 h.

Appetitive and aversive experiences influence whether odors elicit precopulatory behavior from male rodents. A role for aversive experience in odor-elicited reproductive behaviors had been demonstrated for hamsters and rats, but similar work on house mice had not been performed. Four experiments examined whether lithium chloride (LiCl) aversive conditioning would alter two precopulatory behaviors (ultrasonic vocalizations and olfactory preference) that male house mice normally exhibit to female urine. Lithium chloride was used to aversively condition male house mice to either female urine odor, female urine in drinking water, the female herself, or a novel odor. Independent tests of taste aversion establishment were also conducted. In these experiments, LiCl aversive conditioning produced robust taste aversions to water adulterated with either female urine or a novel odorant/tastant (isoamylacetate), but only transitory decrements in odor-elicited, male-typical precopulatory behaviors. We conclude that aversive conditioning is unlikely to be a significant factor affecting male mouse precopulatory behavior.