The anterior medial amygdala transmits sexual odor information to the posterior medial amygdala and related forebrain nuclei

In Syrian hamsters, reproductive behavior relies on the perception of chemical signals released from conspecifics. The medial amygdala (MEA) processes sexual odors through functionally distinct, but interconnected, sub‐regions; the anterior MEA (MEAa) appears to function as a chemosensory filter to distinguish between opposite‐sex and same‐sex odors, whereas the posterodorsal MEA (MEApd) is critical for generating attraction specifically to opposite‐sex odors. To identify how these sub‐regions interact during odor processing, we measured odor‐induced Fos expression, an indirect marker of neuronal activation, in the absence of either MEAa or MEApd processing. In Experiment 1, electrolytic lesions of the MEAa decreased Fos expression throughout the posterior MEA in male hamsters exposed to either female or male odors, whereas MEApd lesions had no effect on Fos expression within the MEAa. These results indicate that the MEAa normally enhances processing of sexual odors within the MEApd and that this interaction is primarily unidirectional. Furthermore, lesions of the MEAa, but not the MEApd, decreased Fos expression within several connected forebrain nuclei, suggesting that the MEAa provides the primary excitatory output of the MEA during sexual odor processing. In Experiment 2, we observed a similar pattern of decreased Fos expression, using fiber‐sparing, NMDA lesions of the MEAa, suggesting that the decreases in Fos expression were not attributable exclusively to damage to passing fibers. Taken together, these results provide the first direct test of how the different sub‐regions within the MEA interact during odor processing, and highlight the role of the MEAa in transmitting sexual odor information to the posterior MEA, as well as to related forebrain nuclei.     

Chemosensory and steroid-responsive regions of the medial amygdala regulate distinct aspects of opposite-sex odor preference in male Syrian hamsters

In rodent species, such as the Syrian hamster, the expression of sexual preference requires neural integration of social chemosensory signals and steroid hormone cues. Although anatomical data suggest that separate pathways within the nervous system process these two signals, the functional significance of this separation is not well understood. Specifically, within the medial amygdala, the anterior region (MEa) receives input from the olfactory bulbs and other chemosensory areas, whereas the posterodorsal region (MEpd) contains a dense population of steroid receptors and receives less substantial chemosensory input. Consequently, the MEa may subserve a primarily discriminative function, whereas the MEpd may mediate the permissive effects of sex steroids on sexual preference. To test these hypotheses, we measured preference and attraction to female and male odors in males with lesions of either the MEa or MEpd. In Experiment 1, lesions of either region eliminated opposite-sex odor preferences. Importantly, MEpd-lesioned males displayed decreased attraction toward female odors, suggesting decreased sexual motivation. In contrast, MEa-lesioned males displayed high levels of investigation of both male and female odors, suggesting an inability to categorize the relevance of the odor stimuli. In Experiment 2, we verified that both MEa- and MEpd-lesioned males could discriminate between female and male odors, thereby eliminating the possibility that the observed lack of preference reflected a sensory deficit. Taken together, these results suggest that both the MEa and MEpd are critical for the expression of opposite-sex odor preference, although they appear to mediate distinct aspects of this behavior.     

Excitotoxic lesions of the tegmental pedunculopontine nucleus impair copulation in naive male rats and block the rewarding effects of copulation in experienced male rats

The tegmental pedunculopontine nucleus (TPP) of the brainstem mediates food reward in food-sated animals and opiate reward in drug-naive animals. In the present study, we examine the effect of excitotoxic lesions of the TPP on sexual behaviour in naive and experienced male rats. Male, Long-Evans rats received either 0.25 micro L injections of NMDA (4.2 micro g/side) or vehicle (shams) into the TPP. In sexually naive males, complete bilateral TPP lesions decreased all measure of copulation (i.e. mounts, intromissions and ejaculations), prevented acquisition of conditioned sexual excitement, decreased approach preference for a receptive female over a non-receptive one, and decreased non-contact erections; unilateral or bilateral posterior-sparing TPP lesions did not affect any of these measures. Conversely, in sexually experienced males, lesions not only failed to disrupt copulation, but also increased conditioned sexual excitement, decreased post-ejaculatory interval and blocked the effect of prolonged copulation on conditioned sexual excitement. Following differential pairing of distinctive environments with and without copulation, sham males with sexual experience displayed a significant preference for the environment paired with copulation, whereas the lesion males with sexual experience displayed a significant aversion for the environment paired with copulation. These findings indicate that the TPP is critical for the acquisition of copulation in naive males and mediates the rewarding consequences of copulation in experienced males. Together these findings demonstrate that the TPP mediates sexual reward, but that sexual experience is not sufficient to produce a deprivation state.     

Remodeling of the number and structure of dendritic spines in the medial amygdala: From prepubertal sexual dimorphism to puberty and effect of sexual experience in male rats

The posterodorsal medial amygdala (MePD) is a sexually dimorphic area and plays a central role in the social behavior network of rats. Dendritic spines modulate synaptic processing and plasticity. Here, we compared the number and structure of dendritic spines in the MePD of prepubertal males and females and postpubertal males with and without sexual experience. Spines were classified and measured after three‐dimensional image reconstruction using DiI fluorescent labeling and confocal microscopy. Significantly differences are as follows: (a) Prepubertal males have more proximal spines, stubby/wide spines with long length and large head diameter and thin and mushroom spines with wide neck and head diameters than prepubertal females, whereas (b) prepubertal females have more mushroom spines with long neck length than age‐matched males. (c) In males, the number of thin spines reduces after puberty and, compared to sexually experienced counterparts, (d) naive males have short stubby/wide spines as well as mushroom spines with reduced neck diameter. In addition, (e) sexually experienced males have an increase in the number of mushroom spines, the length of stubby/wide spines, the head diameter of thin and stubby/wide spines and the neck diameter of thin and mushroom spines. These data indicate that a sexual dimorphism in the MePD dendritic spines is evident before adulthood and a spine‐specific remodeling of number and shape can be brought about by both puberty and sexual experience. These fine‐tuned ontogenetic, hormonally and experience‐dependent changes in the MePD are relevant for plastic synaptic processing and the reproductive behavior of adult rats.     

Lesions of the SDN-POA inhibit sexual behavior of male Wistar rats

Discrete bilateral lesions in the SDN-POA of sexually naive adult male rats were found to decrease the number of animals ejaculating and/or to increase latencies to the first mount, intromission and ejaculation. The deleterious effects of the lesions disappeared after 4 tests for sexual behavior but were reinstated when the males were tested under suboptimal conditions, i.e., when they were tested with a marginally receptive female or when they had only limited access to the stimulus female. It was subsequently shown that males with a bilaterally lesioned SDN-POA still showed an increase in plasma testosterone. LH and prolactin levels in response to sexual stimulation. Effects of the lesions on scent marking were not found. Together with previous data indicating that SDN-POA-lesions disrupt masculine sexual behavior in females, these data are taken as evidence that the SDN-POA plays a role in the regulation of masculine sexual behavior. The data further suggest that previously reported negative results of SDN-POA-lesions on masculine sexual behavior in male rats might be attributed to the use of sexually experienced instead of sexually inexperienced animals.     

Lesions of the nucleus paragigantocellularis: effects on mating behavior in male rats

We evaluated the effects of bilateral radio-frequency lesions of the paragigantocellular (PGi) reticular nucleus in the ventral medulla on male rat copulatory behavior. In Experiment 1, sexually naive male rats with such lesions were more likely than sham-operated controls to copulate to ejaculation during their first exposure to an estrous female. Additionally, among the rats that copulated to ejaculation, those with lesions demonstrated a reduction in mount frequency (MF), intromission frequency (IF), and ejaculation latency (EL), and an increase in copulatory efficiency (CE). In Expt. 2, sexually experienced male rats were allowed to mate to sexual exhaustion. Males with PGi lesions showed an increased latency to sexual exhaustion and an increased number of ejaculations prior to exhaustion. Additionally, rats with PGi lesions displayed reductions in IF, EL, and post-ejaculatory interval (PEI) as they approached sexual exhaustion. Our results provide further evidence that the PGi is a supraspinal locus of descending inhibitory influence on spinal nuclei mediating ejaculatory reflexes in the male rat.     

Conspecific odor preferences of male albino rats are reversed by intracerebral 6-hydroxydopamine

The odor of pine shavings from the nest of a female and her litter attracted sexually experienced male Sprague-Dawley rats tested in a two-choice situation. The preference persisted after surgery in controls treated with bilateral intracerebral injections of saline-ascorbic into the vicinity of the ascending noradrenergic bundle. In contrast, 7 out of 8 animals receiving bilateral injections of the neurotoxin 6-hydroxydopamine (6-OHDA), preferred the odor of clean pine to pine nest odor after surgery, indicating preference reversal. 6-hydroxydopamine also reduced olfactory cortex norepinephrine (NE) concentrations by 85%. Pretreatment with intracerebral injection of amphetamine, a catecholamine uptake inhibitor and releaser, prevented 6-OHDA-induced preference reversal in 7 out 8 animals and limited NE reduction to 38% of concentrations measured in amphetamine-pretreated vehicle-injected controls. The data implicate central NE in the modulation of responses to conspecific odors.     

Oestrogen and androgen receptor activation contribute to the masculinisation of oxytocin receptors in the bed nucleus of the stria terminalis of rats

Oxytocin (OT) often regulates social behaviours in sex‐specific ways, and this may be a result of sex differences in the brain OT system. Adult male rats show higher OT receptor (OTR) binding in the posterior bed nucleus of the stria terminalis (pBNST) than adult female rats. In the present study, we investigated the mechanisms that lead to this sex difference. First, we found that male rats have higher OTR mRNA expression in the pBNST than females at postnatal day (P) 35 and P60, which demonstrates the presence of the sex difference in OTR binding density at message level. Second, the sex difference in OTR binding density in the pBNST was absent at P0 and P3, but was present by P5. Third, systemic administration of the oestrogen receptor (ER) antagonist fulvestrant at P0 and P1 dose‐dependently reduced OTR binding density in the pBNST of 5‐week‐old male rats, but did not eliminate the sex difference in OTR binding density. Fourth, pBNST‐OTR binding density was lower in androgen receptor (AR) deficient genetic male rats compared to wild‐type males, but higher compared to wild‐type females. Finally, systemic administration of the histone deacetylase inhibitor valproic acid at P0 and P1 did not alter pBNST‐OTR binding density in 5‐week‐old male and female rats. Interestingly, neonatal ER antagonism, AR deficiency, and neonatal valproic acid treatment each eliminated the sex difference in pBNST size. Overall, we demonstrate a role for neonatal ER and AR activation in setting up the sex difference in OTR binding density in the pBNST, which may underlie sexual differentiation of the pBNST and social behaviour.     

Influence of a natural stressor (predator odor) on locomotor activity in the meadow vole (Microtus pennsylvanicus): modulation by sex, reproductive condition and gonadal hormones

Sex differences in a variety of non-reproductive behaviors have been indicated to occur in seasonally breeding polygynous promiscuous rodents such as the meadow vole, Microtus pennsylvanicus. The present study was designed to assess the effects of reproductive and hormonal status on the locomotor responses of meadow voles following brief exposure to the odors of a natural predator, the Red fox (Vulpes vulpes). Adult male and female meadow voles, which are seasonal photoperiodically-induced breeders, were housed in either mixed sex pairs under a long, reproductively stimulatory photoperiod (simulating breeding: long light cycle, paired: LLC + P) or in same-sex pairs under a short, reproductively inhibitory photoperiod (simulated non-breeding: short light cycle, non-paired: SLC-NP). On 2 consecutive days following 1 day of baseline activity monitoring, voles were exposed individually for 3 min to fox odor and a novel pungent control odor (extract of almond). The levels of various measures of activity that were displayed by the voles were assessed by an automated Digiscan activity monitoring system. LLC + P (simulated breeding) voles displayed higher basal levels of activity relative to SLC + NP (simulated non-breeding) voles, with males displaying greater activity than females. LLC + P (simulated breeding) males displayed a significant reduction in activity levels following exposure to fox odor relative to control odor. The reductions in activity following fox odor exposure were related to plasma testosterone levels such that a larger behavioral response (i.e. greater reduction) was associated with higher levels of testosterone. Furthermore, dividing males into high and low testosterone groups based on the median levels of testosterone revealed that high but not low testosterone males displayed reductions in activity following exposure to fox odor relative to control odor. No changes in activity levels following exposure to fox odor were noted in SLC-NP males, and either SLC-NP or LLC + P females. These results show that this sexually dimorphic non-reproductive behavior is significantly influenced by reproductive condition and gonadal hormone levels.     

Corticosterone rapidly reduces male odor preferences in female mice

There is accumulating evidence for rapid, non-genomic behavioral effects of various steroids including that of the glucocorticoid, corticosterone. Using an odor preference test, the responses of which are indicative of mate preferences and sexual interest, we examined the effects of acute corticosterone on the responses of oestrous female mice to male odors. Control female mice displayed an overwhelming preference for the odors of male mice. Peripheral administration of corticosterone elicited a significant dose-related (1.0-5.0 mg/kg) decrease in female preference for male odors at 10 min, but not at 60 min, after administration. These inhibitory effects of corticosterone on odor preferences were significantly reduced by the competitive NMDA antagonist, NPC 12626, and enhanced by the GABA antagonist bicuculline. This indicates that corticosterone has rapid inhibitory effects on olfactory mediated female mate preferences and responses to male odor that in part involve interactions with NMDA and GABA receptor mechanisms.     

Neonatal 6-hydroxydopa alters conspecific odor investigation by male rats

Odor-guided behavior was examind in male rats injected at birth and 48 h later with either the catecholaminergic neurotoxin 6-hydroxydopamine (60 μg/g, i.p.) or vehicle. In odor preference tests administered 8 or 74 days postnatally, drug-treated animals avoided novel odors which were neutral for controls and showed reduced preference for conspecific nest odors. In emergence tests administered 73–75 days postnatally, odors from an anesthetized female reduced approach latency and increased investigation of familiar conspecific odors for control but not drug-treated males. Neonatal drug treatment decreased adult olfactory cortex norepinephrine (NE) levels by 50%, but did not significantly influence either cardiac NE or olfactory cortex dopamine. The results imply that brain NE facilitates attraction to familiar conspecific odors.     

Effects of discrete lesions of the sexually dimorphic nucleus of the preoptic area or other medial preoptic regions on the sexual behavior of male rats

The sexually dimorphic nucleus of the rat medial preoptic area (SDN-POA) has a volume five times larger in the adult male compared with that of the adult female. In the present study, the effects of discrete electrolytic destruction of the SDN-POA or other specific medial preoptic (MPOA) regions on masculine sexual behavior were determined in adult, sexually experienced male rats. Small lesions encompassing the SDN-POA had no effect on the maintenance of copulatory behavior. Lesions of similar size placed within the ventral or anterio-dorsal MPOA also did not consistently affect the display of masculine sexual behavior. However, animals that received small lesions within their dorsal MPOA showed a substantial, long-term decrease in number of mounts, intromissions, and ejaculations compared to these parameters in sham-lesioned control rats, thus indicating a lesion-induced disruption of those neural mechanisms mediating these behaviors. Collectively these data suggest that the SDN-POA is not critical for a full expression of male sexual behavior and that the dorsal MPOA may be more important than other MPOA regions for copulatory behavior.     

Male aromatase knockout mice acquire a conditioned place preference for cocaine but not for contact with an estrous female

We have previously shown that male mice carrying a targeted mutation in the Cyp19 gene which encodes the aromatase enzyme (aromatase knockout or ArKO), showed a reduced interest to investigate volatile odors from conspecifics in a Y-maze. We asked here whether the incentive value of reproductively relevant odors is reduced in ArKO males by comparing the ability of male wild-type (WT) and ArKO mice to learn a conditioned place preference using exposure to reproductively relevant odors as incentive stimuli. When the presence of an anesthetized estrous female or soiled bedding from estrous females was used as incentive stimuli, only WT and not male ArKO mice showed conditioned place preference suggesting that the reward value of these stimuli is reduced in ArKO males. However, ArKO males showed conditioned place preference when cocaine was used as incentive stimulus, indicating that ArKO males are able to learn the conditioned place preference procedure. These results thus further confirm the important role of estradiol in sexually related behavioral responses in male mice.     

Relationship between sexual behavior and sexually dimorphic structures in the anterior hypothalamus in control and prenatally stressed male rats

The present study was designed to examine the effects of prenatal stress on the morphological development of sexually dimorphic structures in the anterior hypothalamus in male rats and to determine if there is a relationship between morphologic development of the brain and copulatory behavior in individual animals. Dams in the stress group were subjected to treatments of heat-light restraint during the third trimester of gestation (day 14 to parturition) three times daily for 45-min periods. At 90 days of age, prenatally stressed and control male offspring were tested during the dark cycle for spontaneous male sexual behavior. Volumes of the sexually dimorphic nucleus of the preoptic area (SDN-POA) and the anteroventral periventricular nucleus (AVPV) were measured. Comparisons were made between copulatory behavior and hypothalamic nuclear volumes. SDN-POA volumes were significantly reduced (feminized; males have a larger SDN-POA than females) in prenatally stressed males that did not copulate, whereas, SDN-POA volumes in prenatally stressed males that copulated were not altered. The few control males that did not copulate (sexually non-active) also had significantly reduced SDN-POA volumes compared to the control males that did copulate (sexually active). The volume of the AVPV was significantly increased (feminized; males have a smaller AVPV than females) in prenatally stressed males that were sexually non-active compared to AVPV volumes in sexually active males. The results obtained in this study provide a strong positive relationship between sexual behavior and the morphology of the two sexually dimorphic structures measured.     

Copulatory behavior is controlled by the sexually dimorphic nucleus of the quail POA

The medial preoptic nucleus (POM) of the quail preoptic area is sexually dimorphic and testosterone sensitive. Stereotaxic implantation of needles filled with crystalline testosterone demonstrated that the POM is a critical site of steroid action in the control of copulatory behavior. Only implants located in the POM reliably restored the behavior in castrated birds. Implants around the nucleus weakly activated the behavior; those which were distant by more than 200 microns were totally inactive. Electrolytic lesions confirmed the role of the POM in the control of copulatory behavior. The percentage of the POM which was lesioned was highly correlated to the behavioral deficit while the absolute size of the lesion was not. Electrolytic lesions in or around POM also significantly decreased the volume of the nucleus suggesting that the afferents and efferents of the nucleus are required for its full development. The total volume of the POM was correlated with the sexual behavior of the birds. The morphological changes in POM observed following exposure to testosterone probably represent the signature of the behavioral effects of the steroid. The sexually dimorphic testosterone-sensitive POM is therefore an excellent animal model to study the brain-steroid interactions which mediate the activation of male reproductive behavior.     

Effects of alpha-methyl-para-tyrosine on monoamine levels in the Japanese quail: sex differences and testosterone effects.

Experiments were performed to obtain more information on the regulation by steroids of catecholaminergic systems in the brain of Japanese quail. Dose-response and time-response experiments were first performed to determine optimal conditions for measuring turnover in the quail brain. The norepinephrine and dopamine turnover were then estimated in microdissected brain nuclei of birds that were either sexually mature or gonadectomized or gonadectomized and treated with testosterone. Two major facts that bear direct relationship with the control of masculine reproductive behavior were demonstrated. On one hand, the dopamine turnover in the medial preoptic nucleus, a sexually dimorphic brain structure which is critically implicated in the control of copulatory behavior was much higher in male than in female quail irrespective of the hormonal condition of the birds. On the other hand, norepinephrine concentrations appeared to be higher in several nuclei of the female brain by comparison with males. These sex differences might represent part of the causal factors that underlie the sex dimorphism in reproductive behavior in quail.     

Effects of α-methyl-para-tyrosine on monoamine levels in the japanese quail: Sex differences and testosterone effects

Experiments were performed to obtain more information on the regulation by steroids of catecholaminergic systems in the brain of Japanese quail. Dose-response and time-response experiments were first performed to determine optimal conditions for measuring turnover in the quail brain. The norepinephrine and dopamine turnover were then estimated in microdissected brain nuclei of birds that were either sexually mature or gonadectomized or gonadectomized and treated with testosterone. Two major facts that bear direct relationship with the control of masculine reproductive behavior were demonstrated. On one hand, the dopamine turnover in the medial preoptic nucleus, a sexually dimorphic brain structure which is critically implicated in the control of copulatory behavior was much higher in male than in female quail irrespective of the hormonal condition of the birds. On the other hand, norepinephrine concentrations appeared to be higher in several nuclei of the female brain by comparison with males. These sex differences might represent part of the causal factors that underlie the sex dimorphism in reproductive behavior in quail.     

The nature of the conditioned response mediating olfactory conditioned ejaculatory preference in the male rat

We have developed a model to study the influence of conditioning on sexual partner preference in the rat. In this model, pairing a neutral odor (almond) with copulation to ejaculation produces a subsequent preference to ejaculate with females bearing that odor. We refer to this phenomenon as a conditioned ejaculatory preference (CEP). The present study investigates the nature of the conditioned response that mediates CEP. Given equal mount and intromission distributions, but an unequal ejaculation distribution, we hypothesized that two mechanisms could account for CEP: facilitated ejaculation or selective ejaculation. To test these hypotheses, we examined the effect of omitting the olfactory conditioned stimulus (CS) or applying it to a nonreceptive female. Males trained with the CS paired with copulation (paired-trained males) failed to display evidence of delayed ejaculation when copulating with unscented females. Conversely, paired-trained males displayed CS-elicited copulatory behavior with CS-bearing nonreceptive females. In addition, we re-analyzed the data from the copulatory preference tests of previous experiments for CEP-displaying males. Again, we failed to find evidence of facilitated ejaculation with the scented female. However, the time between the last mount or intromission and ejaculation was increased if either occurred with the scented female. Furthermore, more mounts were directed towards the scented female near the end, but not at other points, of an ejaculatory series. These findings suggest that the paired-trained males attend to the scented female near the point of ejaculation, and are consistent with the hypothesis that CEP is mediated by selective ejaculation.     

Effects of a D1 antagonist and of sexual experience on copulation-induced Fos-like immunoreactivity in the medial preoptic nucleus.

The medial preoptic nucleus (MPN) of the medial preoptic area (MPOA) and the medial amygdala are two brain regions in which male rat sexual behavior increased Fos-like immunoreactivity (Fos-Li). Dopamine is released in the MPOA during male rat sexual behavior and facilitates copulation. Psychostimulants, which increase dopamine levels, induce Fos-Li in the striatum through D1 receptors. We examined whether copulation-induced Fos-Li in the MPN was also mediated through D1 receptors. In Experiment 1, sexually inexperienced male rats that received the D1 antagonist Schering 39166 prior to their first sexual experience had fewer Fos-Li cells in the MPN than did those that received vehicle. In Experiment 2, no significant effect of the D1 antagonist was observed on copulation-induced Fos-Li in male rats that had received repeated sexual experiences prior to the drug test day. Sexual experience increases copulatory efficiency; the mechanisms by which this improvement occurs are unclear. In Experiment 3, copulation by highly experienced male rats led to greater Fos-Li in the MPN than did copulation by sexually naive males. Although there were no differences between groups in amygdala Fos-Li in these studies, in several groups Fos-Li in the medial amygdala was positively correlated with the post-ejaculatory interval. These experiments indicate that (1) stimulation of D1 receptors may contribute to the transient copulation-induced increase in Fos-Li in the MPN, and (2) repeated sexual experiences enhanced copulation-induced Fos-Li in the MPN, which may represent a marker of altered responsiveness of neurons in the MPN to sexual or conditioned stimuli.     

MPOA and BNST lesions in male Syrian hamsters: differential effects on copulatory and chemoinvestigatory behaviors

Electrolytic lesions were made in the medial preoptic area (MPOA) and bed nucleus of the stria terminalis (BNST) to evaluate their participation in the neural regulation of copulatory and chemoinvestigatory behaviors in male hamsters. Damage to either the MPOA or the BNST caused severe deficits in copulatory performance in a subset of the animals in each group. In the MPOA group all males displaying severe deficits had lesions which included a small central region of the caudal MPOA. In the BNST group, animals with severe copulatory deficits all had large lesions which covered most of both the medial and lateral parts of the nucleus. In contrast, MPOA and BNST lesions differentially affected chemoinvestigatory behaviors. MPOA lesions did not affect any of the males' anogenital investigation rates or attraction to female odors, even though some of these hamsters had stopped mating completely. Males with BNST lesions, on the other hand, all displayed significant reductions in their chemoinvestigatory responding even though the majority of them continued to mate normally. We suggest that the MPOA and BNST may in part regulate male sexual behavior by differentially responding to 'attractant' and 'mounting' substances within female hamster vaginal secretion.