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Synopsis Like many scientific disciplines, the field of reproductive biology is subject to biases in terminology and research foci. For example, females are often described as coy and passive players in reproductive behaviors and are termed “promiscuous” if they engage in extra-pair copulations. Males on the other hand are viewed as actively holding territories and fighting with other males. Males are termed “multiply mating” if they mate with multiple females. Similarly, textbooks often illustrate meiosis as it occurs in males but not females. This edition of Integrative and Comparative Biology (ICB) includes a series of papers that focus on reproduction from the female perspective. These papers represent a subset of the work presented in our symposium and complementary sessions on female reproductive biology. In this round table discussion, we use a question and answer format to leverage the diverse perspectives and voices involved with the symposium in an exploration of theoretical, cultural, pedagogical, and scientific issues related to the study of female biology. We hope this dialog will provide a stepping-stone toward moving reproductive science and teaching to a more inclusive and objective framework.

Synopsis In the search for understanding female sexual decision-making, progress has been made in uncovering a variety of perceptual biases and most of these concern the animal’s sensory biology and cognitive processes. We are now poised to dig deeper into the female’s decision-making and ask if incentive salience, which plays a role in all types of appetitive behaviors, also influences a female’s “taste for the beautiful.” The incentive salience hypothesis suggests that dopamine assigns value or salience to objects or actions. After value is assigned to all potential actions, an action selection system then chooses among potential options to select the most valuable action. In this view, dopamine stimulates reward-seeking behavior by assigning incentive salience to specific behavioral actions, which in turn, increases pursuit and focus on objects or stimuli that represent the valuable action. Here, we apply this framework to understand why females are compelled to respond maximally to some male courtship signals over others and how this process may reveal a female’s hidden mate preferences. We examine studies of dopamine and the mesolimbic reward system because these may play a role in expanding the female’s perceptual landscape for novelty in male courtship signals and establishing novel hidden preferences. We review three avenues of research that may identify signatures of incentive salience in females during sexual decision-making. This review includes studies of dopamine agonist or antagonist administration in females during mate choice or partner preference tests, measures of neural activity in dopaminergic neural circuits during mate choice or partner preference tests, and social regulation of dopamine in females when entering reproductive contexts and/or exposure to mate signals. By applying the incentive salience hypothesis to female reproductive decision-making, it redefines how we see the female’s role in sexual encounters. Females cannot be considered passive during reproductive encounters; rather they are seeking sexual encounters, particularly with males that tap into their perceptual biases and initiate a reward-seeking response. Incentive salience applied to reproductive behavior requires considering females as viewing sexual stimuli as rewarding and initiating action to seek out this reward, all of which indicates females are driving sexual encounters.

Obligate brood-parasitic birds never build nests, incubate eggs or supply nestlings with food or protection. Instead, they leave their eggs in nests of other species and rely on host parents to raise their offspring, which allows the parasite to continue reproducing throughout the breeding season. Although this may be a clever fitness strategy, it is loaded with a set of dynamic challenges for brood parasites, including recognizing individuals from their own species while growing up constantly surrounded by unrelated individuals, remembering the location of potential host nests for successful reproduction and learning the song of their species while spending time being entirely surrounded by another species during a critical developmental period, a predicament that has been likened to being ‘raised by wolves’. Here, I will describe what we currently know about the neurobiology associated with the challenges of being a brood parasite and what is known about the proximate mechanisms of brood parasite evolution. The neuroethology of five behaviors (mostly social) in brood parasites is discussed, including: (1) parental care (or the lack thereof), (2) species recognition, (3) song learning, (4) spatial memory and (5) pair-bonding and mate choice. This Review highlights how studies of brood parasites can lend a unique perspective to enduring neuroethological questions and describes the ways in which studying brood-parasitic species enhances our understanding of ecologically relevant behaviors.

Parental care is critical for offspring survival in many species. However, parental behaviors have been lost in roughly 1% of avian species known as the obligate brood parasites. To shed light on molecular and neurobiological mechanisms mediating brood parasitic behavior, we compared brain gene expression patterns between two brood parasitic species and one closely related non-parasitic Icterid (blackbird) species. Our analyses focused on gene expression changes specifically in the preoptic area (POA), a brain region known to play a critical role in parental behavior across vertebrates. Using comparative transcriptomic approaches, we identified gene expression patterns associated with brood parasitism. We evaluated three non-mutually exclusive alternatives for the evolution of brood parasitism: (1) retention of juvenile-like (neotenic) gene expression, (2) reduced expression of maternal care-related genes in the POA, and/or (3) increased expression of genes inhibiting maternal care. We find evidence for neotenic expression patterns in both species of parasitic cowbirds as compared to maternal, non-parasites. In addition, we observed differential expression in a number of genes with previously established roles in mediating maternal care. Together, these results provide the first insight into transcriptomic and genetic mechanisms underlying the loss of maternal behavior in avian brood parasites.

Prolactin is often referred to as the "parental hormone" but there are examples in which prolactin and parental behavior are disconnected. One intriguing example is in avian obligate brood parasites; species exhibiting high circulating prolactin but no parental care. To understand this disconnect, we examined transcriptional and behavioral responses to prolactin in brown-headed (Molothrus ater) and bronzed (M aeneus) brood parasitic cowbirds. We first examine prolactin-dependent regulation of transcriptome wide gene expression in the preoptic area (POA), a brain region associated with parental care across vertebrates. We next examined prolactin-dependent abundance of seven parental care-related candidate genes in hypothalamic regions that are prolactin-responsive in other avian species. We found no evidence of prolactin sensitivity in cowbirds in either case. To understand this prolactin insensitivity, we compared prolactin receptor transcript abundance between parasitic and nonparasitic species and between prolactin treated and untreated cowbirds. We observed significantly lower prolactin receptor transcript abundance in brown-headed but not bronzed cowbird POA compared with a nonparasite and no prolactin-dependent changes in either parasitic species. Finally, estrogen-primed female brown-headed cowbirds with or without prolactin treatment exhibited significantly greater avoidance of nestling begging stimuli compared with untreated birds. Taken together, our results suggest that modified prolactin receptor distributions in the POA and surrounding hypothalamic regions disconnect prolactin from parental care in brood parasitic cowbirds.

Species recognition mediates the association of individuals with conspecifics. Learned cues often facilitate species recognition via early social experience with parents and siblings. Yet, in some songbirds, the production of species-typical vocalizations develops in the absence of early social experiences. Here, we investigate the auditory-evoked neural responses of juvenile red-winged blackbirds (Agelaius phoeniceus), a nonparasitic (parental) species within the Icterid family and contrast these results with a closely related Icterid parasitic species that lacks parental care, the brown-headed cowbird (Molothrus ater). We demonstrate that immediate early gene (IEG) activity in the caudomedial mesopallium (CMM) is selectively evoked in response to conspecific non-learned vocalizations in comparison to 2 types of heterospecific non-learned vocalizations, independent of the acoustic similarity patterns between the playback stimuli. This pattern, however, was not detected in the caudomedial nidopallium (NCM). Because the red-winged blackbird is a parental species, the conspecific non-learned vocalization is presumably a familiar sound to the juvenile red-winged blackbird, whereas the heterospecific non-learned vocalizations are novel. We contrast results reported here with our recent demonstration of selective IEG induction in response to non-learned conspecific vocalizations in juvenile parasitic brown-headed cowbirds, in which conspecific non-learned vocalizations are presumably novel. In this case, selective IEG induction from conspecific non-learned vocalization occurred within NCM but not within CMM. By comparing closely related species with stark differences in the early exposure to conspecifics, we demonstrate that CMM and NCM respond to familiar vs. novel non-learned vocalizations in a manner that parallel previously reported regional responses to learned vocalizations such as conspecific songs.

Brood parasitic songbirds are a natural system in which developing birds are isolated from species-typical song and therefore present a unique opportunity to compare neural plasticity in song learners raised with and without conspecific tutors. We compared perineuronal nets (PNN) and parvalbumin (PV) in song control nuclei in juveniles and adults of two closely related icterid species (i.e. blackbirds): brown-headed cowbirds (Molothrus ater; brood parasite) and red-winged blackbirds (Agelaius phoeniceus; non-parasite). The number of PV cells per nucleus was significantly higher in adults compared with juveniles in the nucleus HVC and the robust nucleus of the arcopallium (RA), whereas no significant species difference appeared in any region of interest. The number of PNN per nuclei was significantly higher in adults compared with juveniles in HVC, RA and Area X, but only RA exhibited a significant difference between species. PV cells surrounded by PNN (PV+PNN) also exhibited age-related differences in HVC, RA and Area X, but RA was the only region in which PV+PNN exhibited significant species differences. Furthermore, a significant interaction existed in RA between age and species with respect to PNN and PV+PNN, revealing RA as a region displaying differing plasticity patterns across age and species. Additional comparisons of PNN and PV between adult male and female cowbirds revealed that males have greater numbers of all three measures in RA compared with females. Species-, sex- and age-related differences in RA suggest that species differences in neural plasticity are related to differences in song production rather than sensitivity to song learning, despite a stark contrast in early exposure to conspecific male tutors.

Auditory communication in humans and other animals frequently takes place in noisy environments with many co-occurring signallers. Receivers are thus challenged to rapidly recognize salient auditory signals and filter out irrelevant sounds. Most bird species produce a variety of complex vocalizations that function to communicate with other members of their own species and behavioural evidence broadly supports preferences for conspecific over heterospecific sounds (auditory species recognition). However, it remains unclear whether such auditory signals are categorically recognized by the sensory and central nervous system. Here, we review 53 published studies that compare avian neural responses between conspecific versus heterospecific vocalizations. Irrespective of the techniques used to characterize neural activity, distinct nuclei of the auditory forebrain are consistently shown to be repeatedly conspecific selective across taxa, even in response to unfamiliar individuals with distinct acoustic properties. Yet, species-specific neural discrimination is not a stereotyped auditory response, but is modulated according to its salience depending, for example, on ontogenetic exposure to conspecific versus heterospecific stimuli. Neuromodulators, in particular norepinephrine, may mediate species recognition by regulating the accuracy of neuronal coding for salient conspecific stimuli. Our review lends strong support for neural structures that categorically recognize conspecific signals despite the highly variable physical properties of the stimulus. The available data are in support of a 'perceptual filter'-based mechanism to determine the saliency of the signal, in that species identity and social experience combine to influence the neural processing of species-specific auditory stimuli. Finally, we present hypotheses and their testable predictions, to propose next steps in species-recognition research into the emerging model of the neural conceptual construct in avian auditory recognition.

Social regulation of reproductive hormones is a means by which conspecific males and females orchestrate successful reproductive efforts. There is variation, however, in the range of social cues that will initiate a hormone response in the receiver of social signals. We investigate whether social cues modify activity within the hypothalamic-pituitary-gonadal (HPG) axis and the specificity of this response in a social parasite that is known to eavesdrop on the communication signals of other species: the brown-headed cowbird (Molothrus ater). Brown-headed cowbirds are obligate brood parasites that do not build nests or care for their own young. Instead, obligate brood parasites always leave their eggs in the nest of a host species thereby receiving the benefits of parental care toward offspring without paying any of the costs. Thus, social parasites must coordinate their breeding attempts with conspecifics as well as potential heterospecific hosts and therefore, social parasites such as cowbirds rely on the communication signals of host species to help locate nests to parasitize during the breeding season. Here, we explore whether the vocal signals of potential host species can also be used as a social cue that modifies the HPG axis of female brown-headed cowbirds. Results reveal that both conspecific and heterospecific song-exposed females exhibit significantly greater circulating estradiol concentrations as compared to silence-exposed females. While conspecific song induces the greatest elevation in circulating estradiol, there is no significant difference in circulating estradiol levels in females exposed to either conspecific or heterospecific songs. This pattern suggests both song types are effective at evoking a reproductive physiological response. On the other hand, circulating progesterone concentrations did not differ among the song- and silence-exposed groups nor did the size of the female’s ovarian follicles. These results indicate that heterospecific vocal communication signals can effectively be used as a social cue that simultaneously provides necessary information regarding breeding status of hosts and modifies breeding condition of the eavesdropper.HighlightsBrood parasites may coordinate reproductive attempts with potential hosts.Therefore, parasites eavesdrop on other species, which helps them find nests.Vocal signals from conspecific and heterospecifics elevate estrogen in parasites.Hearing potential host songs may enhance reproductive hormones in brood parasites.

Mate choice decision-making requires four components: sensory, cognitive, motivation, and salience. During the breeding season, the neural mechanisms underlying these components act in concert to radically transform the way a female perceives the social cues around her as well as the way in which cognitive and motivational processes influence her decision to respond to courting males. The role of each of these four components in mate choice responses will be discussed here as well as the brain regions involved in regulating each component. These components are not independent, modular systems. Instead, they are dependent on one another. This review will discuss the many ways in which these components interact and affect one another. The interaction of these components, however, ultimately leads back to a few key neuromodulators that thread motivation, sensory, salience, and cognitive components into a set of inter-dependent processes. These neuromodulators are estrogens and catecholamines. This review will highlight the need to understand estrogens in reproductive contexts not just as simply a 'sexual motivation modulator' or catecholamines as 'cognitive regulators' but as neuromodulators that work together to fully transform a non-breeding female into a completely reproductive female displaying: heightened sexual interest in courting males, greater arousal and selective attention toward courtship signals, improved signal detection and discrimination abilities, enhanced contextual signal memory, and increased motivation to respond to signals assigned incentive salience. The aim of this review is to build a foundation in which to understand the brain regions associated with cognitive, sensory, motivational, and signal salience not as independently acting systems but as a set of interacting processes that function together in a context-appropriate manner.

Obligate avian brood parasites are raised by heterospecific hosts and, therefore, lack crucial early exposure to relatives and other conspecifics. Yet, young brood parasites readily recognize and affiliate with others of their own species upon independence. One solution to this social recognition paradox is the ontogenetic 'password' mechanism used by obligate parasitic brown-headed cowbirds (Molothrus ater), whereby conspecific identification is initially mediated through the cowbird chatter: a non-learned vocal cue. We explored the neural basis of such password-based species recognition in juvenile and adult male cowbirds. We found that cowbird auditory forebrain regions express greater densities of the protein product of the immediate-early gene ZENK in response to the password chatter call relative to control sounds of mourning dove (Zenaida macroura) coos. The chatter-selective induction of ZENK expression occurs in both the caudal medial nidopallium (NCM) and the caudal medial mesopallium (CMM) in adults, but only within the NCM in juveniles. In contrast, we discovered that juvenile cowbirds exhibit neural selectivity to presentations of either conspecific or heterospecific songs, but only in CMM and only after recent experience. Juvenile cowbirds that did not have previous experience with the song type they were exposed to during the test period exhibited significantly lower activity-dependent gene expression. Thus, in juvenile male cowbirds, there is early onset of species-specific selective neural representation of non-learned calls in NCM and recently experienced song in CMM. These results suggest that NCM is evolutionarily co-opted in parasitic cowbirds to selectively recognize the password chatter, allowing juvenile cowbirds to identify adult conspecifics and avoid mis-imprinting upon unrelated host species. These ontogenetic comparisons reveal novel insights into the neural basis of species recognition in brood parasitic species.

It is well established that auditory forebrain regions of oscine birds are essential for the encoding of species-typical songs and are, therefore, vital for recognition of song during sociosexual interactions. Regions such as the caudal medial nidopallium (NCM) and the caudal medial mesopallium (CMM) are involved in perceptual processing of song and the formation of auditory memories. There is an additional telencephalic nucleus, however, that has also been implicated in species recognition. This nucleus is HVC, a prominent nucleus that sits at the apex of the song system, and is well known for its critical role in song learning and song production in male songbirds. Here, we explore the functional relationship between auditory forebrain regions (i.e., NCM and CMM) and HVC in female canaries (Serinus canaria). We lesion HVC and examine immediate early gene responses to conspecific song presentation within CMM and NCM to explore whether HVC can modulate auditory responses within these forebrain regions. Our results reveal robust deficits in ZENK-ir in CMM and NCM of HVC-lesioned females when compared with control- and sham-lesioned females, indicating that functional connections exists between HVC and NCM/CMM. Although these connected regions have been implicated in song learning and production in males, they likely serve distinct functions in female songbirds that face the task of song recognition rather than song production. Identifying functional connections between HVC and auditory regions involved in song perception is an essential step toward developing a comprehensive understanding of the neural basis of song recognition. © 2012 Wiley Periodicals, Inc. Develop Neurobiol, 2013

Rather than being a static, species specific trait, reproductive behavior in female amphibians is variable within an individual during the breeding season when females are capable of reproductive activity. Changes in receptivity coincide with changes in circulating estrogen. Estrogen is highest at the point when females are ready to choose a male and lay eggs. At this time female receptivity (her probability of responding to a male vocal signal) is highest and her selectivity among conspecific calls (measured by her probability of responding to a degraded or otherwise usually unattractive male signal) is lowest. These changes occur even though females retain the ability to discriminate different acoustic characteristics of various conspecific calls. After releasing her eggs, female amphibians quickly become less receptive and more choosy in terms of their responses to male sexual advertisement signals. Male vocal signals stimulate both behavior and estrogen changes in amphibian females making mating more probable. The changes in female reproductive behavior are the same as those generally accepted as indicative of a change in female sexual arousal leading to copulation. They are situationally triggered, gated by interactions with males, and decline with the consummation of sexual reproduction with a chosen male. The changes can be triggered by either internal physiological state or by the presence of stimuli presented by males, and the same stimuli change both behavior and physiological (endocrine) state in such a way as to make acceptance of a male more likely. Thus amphibian females demonstrate many of the same general characteristics of changing female sexual state that in mammals indicate sexual arousal.

Birdsong is a species-typical vocal signal that facilitates reproduction and deters competitors. Song production is regulated by a clearly defined and specialized neural circuitry in which high concentrations of catecholamines are present. The nuclei within the song control circuit receive projections from catecholaminergic cell populations involved in attention, arousal and motivation, including periaqueductal gray (PAG), ventral tegmental area (VTA), locus coeruleus (LoC) and sub coeruleus (SC). Here, we examine whether catecholamine-containing neurons in these regions exhibit the immediate early gene, ZENK, during spontaneous, undirected song production in male zebra finches (Taeniopygia guttata). Males were assigned to "singing" or "silent" groups based on the total duration of spontaneous, undirected song produced within a 30 min period. We quantified the number of cells expressing both ZENK-ir and tyrosine hydroxylase (TH)-ir within the VTA, PAG, LoC and SC. The number of cells expressing co-localized ZENK and TH-ir was significantly elevated within the PAG in males that were singing compared to silent males. The number of cells expressing ZENK-ir alone was also elevated in the VTA and SC in singing males compared to silent males. Although ZENK expression is elevated in singing birds it does not positively correlate with the amount of singing produced. It is therefore likely that catecholaminergic PAG neurons are involved in motivational or attentional components of vocal expression rather than vocal motor output. Overall, our study is consistent with the hypothesis that PAG catecholamine-containing neurons as well as VTA and SC neurons play a role in vocal communication of male songbirds.

During acoustic communication, animals must attend to sounds from a particular source while simultaneously rejecting intrusion from other sources. One possible candidate mechanism for this process is the noradrenergic system. Noradrenaline is a neuromodulator that tunes sensory processing systems and regulates attention. We examined whether pharmacological degradation of the noradrenergic system using N-(2-chloroethyl)-N-2-bromobenzyl-amine hydrochloride (DSP-4) modifies processing of species-typical auditory signals in female canaries (Serinus canaria). We measured auditory responses to conspecific and heterospecific songs using ZENK protein expression within the caudomedial nidopallium (NCM) and the mesopallium caudomedial (CMM). Song-induced ZENK expression in these auditory forebrain areas is typically higher in birds exposed to conspecific songs as opposed to heterospecific songs. Our results reveal that this differential ZENK induction is abolished specifically within dNCM and CMM in female canaries treated with DSP-4. Furthermore, in DSP-4-treated birds, conspecific song-induced ZENK expression is significantly reduced when compared to saline-treated birds. This suggests that the noradrenergic system modifies auditory processing by enhancing neuronal responses to signals relevant to survival and reproduction rather than inhibiting neuronal responses to signals that are less relevant. Overall, our results reveal that noradrenaline plays a significant neuromodulatory role during the reception of species-typical communication signals.

In many vertebrates, the production and reception of species-typical courtship signals occurs when gonadotropin and gonadal hormone levels are elevated. These hormones may modify sensory processing in the signal receiver in a way that enhances behavioral responses to the signal. We examined this possibility in female túngara frogs (Physalaemus pustulosus) by treating them with either gonadotropin (which elevated estradiol) or saline and exposing them to either mate choruses or silence. Expression of an activity-dependent gene, egr-1, was quantified within two sub-nuclei of the auditory midbrain to investigate whether gonadotropin plus chorus exposure induced greater egr-1 induction than either of these stimuli alone. The laminar nucleus (LN), a sub-nucleus of the torus semicircularis that contains steroid receptors, exhibited elevated egr-1 induction in response to chorus exposure and gonadotropin treatment. Further analysis revealed that neither chorus exposure nor gonadotropin treatment alone elevated egr-1 expression in comparison to baseline levels whereas gonadotropin + chorus exposure did. This suggests that mate signals and hormones together produce an additive effect so that together they induce more egr-1 expression than either alone. Our previously published studies of female túngara frogs reveal that (1) gonadotropin-induced estradiol elevations also increase behavioral responses to male signals, and (2) reception of male signals elevates estradiol levels in the female. Here, we report data that reveal a novel mechanism by which males exploit female sensory processing to increase behavioral responses to their courtship signals.

Amphibian behavioral endocrinology has focused on reproductive social behavior and communication in frogs and newts. Androgens and estrogens are critical for the expression of male and female behavior, respectively, and their effects are relatively clear. Corticosteroids have significant modulatory effects on the behavior of both sexes, as does the peptide neuromodulator arginine vasotocin in males, but their effects and interactions with gonadal steroids are often complex and difficult to understand. Recent work has shown that the gonadal hormones and social behavior are mutually reinforcing: engaging in social interactions increases hormone levels just as increasing hormone levels change behavior. The reciprocal interactions of hormones and behavior, as well as the complex interactions among gonadal steroids, adrenal steroids, and peptide hormones have implications for the maintenance and evolution of natural social behavior, and suggest that a deeper understanding of both endocrine mechanisms and social behavior would arise from field studies or other approaches that combine behavioral endocrinology with behavioral ecology.

Tropically breeding anurans that require heavy rainfall in order to reproduce are subject to favorable breeding conditions that are sporadic. Although there is an increased probability of rain during the rainy season, the probability of local rainfall is unpredictable and this may influence female anuran reproductive strategies. The female tungara frog, a neotropical frog that requires standing water to breed, maintains readiness to breed at any time via asynchronous oogenesis. Although females constantly produce and maintain oocytes during the breeding season, this study shows that they have cyclic fluctuations in gonadal hormone levels. Plasma levels of estrogen significantly change during three reproductive stages within a single reproductive cycle (P=0.03), as do plasma levels of progesterone and androgen (P

Quantitative geneticists traditionally attribute phenotypic variation to genetic or environmental sources. New findings with near-clonal mice raised in a single enriched environment demonstrated significant developmental and behavioural divergence. This suggests intriguing possibilities for how (epi)genomes and environments might interact to drive phenotypic individuality, thus shaping evolutionary pathways.

Genes that mediate mate preferences potentially play a key role in promoting and maintaining biological diversity. In this study, we compare mate preference behavior in two related poeciliid fishes with contrasting behavioral phenotypes and relate these behavioral differences to gene profiles in the brain. Results reveal that one poeciliid fish, the Northern swordtail, exhibits robust mate preference as compared to the Western mosquitofish, which utilizes a coercive mating system. Female swordtails display no significant difference in association time between male- and female-exposure trials, whereas female mosquitofish spend significantly less time associating with males relative to females. Furthermore, the preference strength for large males is significantly lower in female mosquitofish relative to swordtails. We then examine expression of three candidate genes previously shown to be associated with mate preference behavior in female swordtails and linked to neural plasticity in other vertebrates: neuroserpin (NS), neuroligin-3 (NLG-3) and N-methyl-d-aspartate receptor (NMDA-R). Whole brain gene expression patterns reveal that two genes (NS and NLG-3) are positively associated with mate preference behavior in female swordtails, a pattern opposing that of the mosquitofish. In mosquitofish females, these genes are downregulated when females express biases toward males yet are elevated in association with total motor activity patterns under asocial conditions, suggesting that the presence of males in mosquitofish species may inhibit expression of these genes. Both gene expression and female behavioral responses to males exhibit opposing patterns between these species, suggesting that this genetic pathway may potentially act as a substrate for the evolution of mate preference behavior.

Temperate zone animals exhibit seasonal variation in reproductive physiology. In most cases, seasonal changes in reproductive states are regulated by changes in GnRH1 secretion, rather than synthesis, from the preoptic area (POA)/anterior hypothalamus. An important exception occurs in some songbirds that become photorefractory to the stimulatory effects of long days and show profound decreases in brain GnRH1 protein content. Whether this decline reflects changes in gene expression is unknown because of past failures to measure GNRH1 mRNA levels, due in large part to the absence of available GNRH1 gene sequence in this taxon. Here, we report the first cloning of GNRH1 cDNAs in two songbirds: European starlings and zebra finches. Consistent with the size of the prepro-hormone in other avian and non-avian species, the open-reading frames predict proteins of 91 and 92 amino acids, respectively. Whereas the decapeptide in both species is perfectly conserved with chicken GnRH1, the amino acid identity in the signal peptide and GNRH associated peptide subdomains are significantly less well conserved. At the nucleotide level, the starling and zebra finch coding sequences are approximately 88% identical to each other but only approximately 70% identical to chicken GNRH1. In situ hybridization using radiolabeled cRNA probes demonstrated GNRH1 mRNA expression primarily in the POA, consistent with previous studies on the distribution of the GnRH1-immunoreactive cell bodies. Furthermore, we provide evidence for photoperiod-dependent regulation of GNRH1 mRNA in male starlings. Declines in GNRH1 mRNA levels occur in parallel with testicular involution. Thus, photorefractoriness is associated with decreases in GNRH1 gene expression in the medial POA.

Females alter their mate choices as they transition through different reproductive stages; however, the proximal mechanisms for such behavioral fluctuation are unclear. In many taxa, as females transition through different reproductive stages, there is an associated change in hormone levels; therefore, we examined whether fluctuation in hormone levels serves as a proximal mechanism for within-individual variation in mate choice in female túngara frogs (Physalaemus pustulosus). We manipulated hormone levels of females by administering 0, 10, 100, 500 or 1,000 IU of human chorionic gonadotropin (HCG), which is a ligand for luteinizing hormone (LH) receptors and will therefore cause increased gonadal hormone production. Phonotaxis assays were conducted to measure three aspects of mate choice behavior before and after HCG administration; receptivity (response to a conspecific mate signal), permissiveness (response to a signal that is less attractive than conspecific signals) and discrimination (ability to discern signal differences). The probability of response to a conspecific and an artificial hybrid signal significantly increased at the highest HCG doses. The difference in mean response time between pre- and post-HCG tests was significantly different for both the receptivity and permissiveness tests among the five doses. Increased permissiveness, however, was not due to decreased discrimination because females could discriminate between calls even at the highest HCG doses. These hormonal manipulations caused the same behavioral pattern we reported in females as they transitioned through different reproductive stages (Lynch, K.S., Rand, A.S., Ryan, M.J., Wilczynski, W., 2005. Plasticity in female mate choice associated with changing reproductive states. Anim. Behav. 69, 689-699), suggesting that changes in hormone levels can influence the female's mate choice behavior.