In most animal species, reproductive success depends critically on precopulatory or solicitational behaviors that occur prior to mating. The specific sensory systems and behavioral strategies employed in precopulatory behaviors vary across species; in all cases, however, animals must be able to identify potential mating partners and solicit sexual interest. Female Syrian hamsters (Mesocricetus auratus) engage in multiple forms of precopulatory behaviors that are preferentially expressed to males or their odors, including vaginal scent marking and sexual odor preference. Conspecific odors relevant for precopulatory behaviors are processed by a network of forebrain areas that includes the bed nucleus of the stria terminalis (BNST) and the medial preoptic area (MPOA). The precise functional and neurochemical mechanisms whereby these areas regulate the expression of precopulatory behaviors, however, are unknown. Therefore, the aim of this dissertation is to address the following research questions: (1) Is the neuropeptide oxytocin (OT), acting within BNST or MPOA, necessary for the normal expression of odor-­guided precopulatory behaviors? (2) Is BNST or (3) MPOA required for the preferential expression of vaginal marking or investigation towards male odors?, and (4) Does OT interact with social odor processing to regulate vaginal marking? We found that blockade of OT receptors (OTRs) in MPOA and BNST decreased vaginal marking to male odors. There was no effect of OTR blockade on sexual odor preference. Selective lesions of BNST also disrupted preferential vaginal marking responses to male odors, without affecting sexual odor preference. In contrast, lesions of MPOA disrupted odor preference without affecting vaginal marking responses. Finally, central blockade of OTRs eliminated the normal pattern of increased activation of neurons to male vs. female odors in BNST, but not MPOA. Considered together, these results suggest that OT normally acts within BNST to drive preferential vaginal marking responses to male odors via selective facilitation of neural responses to these odors, and further, that there are separate and distinct neural circuits that regulate different forms of odor-guided precopulatory behaviors in females.

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No description available.

Male rat mating is a suite of individual behaviors mediated by the actions of two metabolites of testosterone (T), dihydrotestosterone (DHT) and estradiol (E2), on the brain. Individually, neither metabolite fully maintains or restores mating in castrated males, but both combined are as effective as T. Two hormone-responsive areas of the brain, the medial preoptic area (MPO) and the medial amygdala (MEA), are crucial for mating. These studies ask: by what mechanism(s) does E2 act in the MPO and MEA? We blocked the conversion of T to E2 in the MEA of intact male rats and sexual behavior was not maintained. We then infused antisense oligodeoxynucleotides (ODNs) to estrogen receptor-alpha (ER-á) mRNA bilaterally to the MPO or the MEA of intact male rats to block ER-á expression. ODN infusion of the MPO attenuated mating but infusion of the MEA had no effect. These results suggest that ER-á is the behaviorally relevant estrogen receptor (ER) in the MPO but not in the MEA. ER was originally described in the cytoplasm and nucleus of cells. Recently plasma membrane associated ERs (mER) have been reported. We conjugated E2 to Bovine Serum Albumin (BSA-E2), a large protein that will not penetrate the plasma membrane, thus restricting the action of E2 to mER, and chronically delivered it to the MPO and MEA. BSA-E2 maintained mating if put in the MPO, but not in the MEA, suggesting a surface action of E2 is sufficient in the MPO. The MPO and MEA are reciprocally connected and probably constitute elements of a larger, steroid-responsive neural network that mediates male mating behavior. To begin to describe this purported circuit, we injected Pseudorabies virus (PRV) into the prostate gland and dually labeled PRV-immunoreactive cells for ER or androgen receptors. We found dual labeling in a forebrain diencephalic circuit that includes the MPO, the medial preoptic nucleus, bed nucleus of stria terminalis, the zona incerta, the periaqueductal gray and other areas that presumably mediate both autonomic and motor aspects of male mating. Together, the results of these studies begin to elucidate locations and mechanisms of E2 mediation of male sexual behavior.

Male rat copulation is mediated by estrogen-sensitive neurons in the medial preoptic area (MPO) and medial amygdala (MEA); however, the mechanisms through which estradiol (E2) acts are not fully understood. We hypothesized that E2 acts through estrogen receptor α (ERα) in the MPO and MEA to promote male mating behavior. Antisense oligodeoxyneucleotides (AS-ODN) complementary to ERα mRNA were bilaterally infused via minipumps into either brain area to block the synthesis of ERα, which we predicted would reduce mating. Western blot analysis and immunocytochemistry revealed a knockdown of ERα in each brain region; however, compared to saline controls, males receiving AS-ODN to the MPO showed significant reductions in all components of mating, whereas males receiving AS-ODN to the MEA continued to mate normally. These results suggest that E2 acts differently in these brain regions to express sexual behavior and that ERα in the MPO, but not in the MEA, promotes mating.

Male rat copulatory behavior is dependent on Testosterone (T) and its metabolites, estradiol (E2) and dihydrotestosterone (DHT). The estrogen receptor (ER) isoforms, ERα and ERβ, exist in the medial Amygdala (MEA) and either receptor might mediate mating behavior. Therefore, the effects of selective estrogenic MEA implants: propyl pyrazole triol (PPT, ERα agonist), diarylpropionitrile (DPN, ERβ agonist), and 1-methyl-4-phenyl pyridinium (MPP, ERα antagonist) were compared to E2 in maintaining sexual behavior. Four groups of male rats were castrated and administered DHT s.c. and bilateral MEA implants containing either cholesterol, E2, PPT or DPN. An additional group of gonadally intact male rats received bilateral MPP-MEA implants. The post-surgical trials showed a significant decrease in the mating behavior of groups that received cholesterol, PPT, or DPN-MEA implants. However, sexual behavior was maintained in male rats that received the E2 or MPP-MEA implants. These results suggest a differential response of the MEA to E2.

Adaptive reproductive behavior requires the ability to recognize and approach possible mating partners in the environment. Syrian hamsters (Mesocricetus auratus) provide a useful animal model by which to study the neural processing of sexual signals, as mate recognition in this species relies almost exclusively on the perception of social odors. In the laboratory, male hamsters prefer to investigate female odors compared to male odors, and this opposite-sex odor preference provides a sensitive measure of the underlying neural processing of sexual stimuli. In addition to chemosensory cues, reproductive behavior in hamsters also requires sufficient levels of circulating gonadal steroid hormones, which reflect the reproductive state of the animal. These chemosensory and hormone signals are processed within an interconnected network of ventral forebrain nuclei, and within this network, the posteromedial cortical amygdala (PMCo) and medial amygdala (MA) are the only nuclei that both receive substantial chemosensory input and are also highly sensitive to steroid hormones. Although a large body of evidence suggests that the MA is critical for generating attraction to sexual odors, the specific role of the PMCo in regulating odor-guided aspects of male reproductive behavior has never been directly tested. Furthermore, detailed analyses of the MA suggest that separate, but interconnected sub-regions within this nucleus process odors differently. Specifically, the anterior MA (MeA) receives the majority of chemosensory input and responds to a variety of social odors, whereas the posterodorsal MA (MePD) receives less chemosensory input but contains the vast majority of steroid receptors. In order to further elucidate how the PMCo and/or MA process sexual odors, this dissertation addressed the following research questions: (1) Is the PMCo required for the expression of either opposite-sex odor preferences or male copulatory behavior? (2) Are functional interactions between MeA and MePD required for the expression of opposite-sex odor preferences? (3) How do MeA and MePD regulate odor responses within the MePD and MeA, respectively? (4) Are odor and/or hormone cues conveyed directly between MeA and MePD? Together, these experiments provide a comprehensive analysis of the functional and neuroanatomical substrates by which the brain processes sexual odors and generates appropriate behavioral responses to these stimuli.

In many species, including Syrian hamsters, male reproductive behavior depends on the perception of odor cues from conspecifics in the environment. Volatile odor cues are processed primarily by the main olfactory system, whereas non-volatile cues are processed primarily by the accessory olfactory system. Together, these two chemosensory systems mediate appetitive reproductive behaviors, such as attraction to female odors, and consummatory reproductive behaviors, such as copulation, in male Syrian hamsters. Main and accessory olfactory information are first integrated in the medial amygdala (MA), a limbic nucleus that is critical for the expression of reproductive behaviors. MA is densely interconnected with other ventral forebrain nuclei that receive chemosensory information and are sensitive to steroid hormones. Specifically, several lines of evidence suggest that MA may generate behavioral responses to socio-sexual odors via functional connections with the posterior bed nucleus of the stria terminalis (BNST) and medial preoptic area (MPOA). It is unknown, however, how these three nuclei act as functional circuit to adaptively regulate appetitive and consummatory reproductive behaviors. Therefore, the overarching goal of this dissertation was to determine how BNST and MPOA function, both uniquely and as a circuit with MA, to generate attraction to female odors and copulatory behaviors in male Syrian hamsters. We found that BNST is required for attraction to female odors, but not for copulation, in sexually-naïve males. In contrast, MPOA is required for both attraction to female odors and for copulation in sexually-naïve males. Surprisingly, prior sexual experience mitigated the requirement of BNST and MPOA for these behaviors. Next, we found that MA preferentially transmits female odor information to BNST and to MPOA, whereas BNST relays female and male odor information equivalently to MPOA. Finally, we found that the functional connections between MA and BNST are required for attraction to female odors but not for copulation, whereas the functional connections between MA and MPOA are required for copulation but not for attraction to female odors. Ultimately, these data may uncover a fundamental mechanism by which this ventral forebrain circuit regulates appetitive and consummatory reproductive behaviors across many species and modalities.

Submitted by Ana Maria Fiscina Sampaio (fiscina@bahia.fiocruz.br) on 2012-10-23T17:18:48Z No. of bitstreams: 1 Daniela Almeida Participaçao dos receptores....pdf: 1297031 bytes, checksum: 754812103a71d951c868e902bd43f90c (MD5) / Made available in DSpace on 2012-10-23T17:18:48Z (GMT). No. of bitstreams: 1 Daniela Almeida Participaçao dos receptores....pdf: 1297031 bytes, checksum: 754812103a71d951c868e902bd43f90c (MD5) Previous issue date: 2012 / Fundação Oswaldo Cruz. Centro de Pesquisas Gonçalo Moniz. Salvador, Bahia, Brasil / Situações de estresse repetido ou prolongado podem resultar em vários estados patológicos, como hipertensão arterial, arritmias cardíacas, infarto do miocárdio e até mesmo morte súbita. Embora se tenha muita informação sobre o controle cerebral da pressão arterial, as respostas cardiovasculares ao estresse não são totalmente compreendidas. Dados da literatura mostram a importância do núcleo medial da amigdala (MeA) e da neurotrasmissão histaminérgica no controle autonômico das funções cardiovasculares, no entanto, não há estudos evidanciando o papel das vias histaminérgicas no MeA nas adaptações cardiovasculares evocada pelo estresse emocional. Desta forma, o objetivo desta pesquisa foi estudar a participação dos receptores H1 e H2 no MeA sobre as respostas cardiovasculares em ratos estressados e não-estressados. Ratos Wistar (280-320g) foram submetidos à cirurgia estereotáxica para canulação bilateral do MeA. Passado cinco dias da estereotaxia, os animais foram submetidos a cateterização da artéria carótida esquerda. Vinte e quatro horas após a inserção do cateter, foram iniciados os experimentos com a gravação do registro da pressão arterial pulsátil (PAP) dos animais em condições basais e em livre movimento em suas respectivas caixas de forma continuada. As drogas utilizadas para a microinjeção central foram a mepiramina (antagonista dos receptores H1) nos grupos experimentais I e III e a cimetidina (antagonista dos receptores H2) nos grupos experimentais II e IV. Nos grupos experimentais I e II, 15 min após microinjeção central bilateral de mepiramina ou cimetidina respectivamente, em diferentes doses, os animais foram submetidos a estresse de restrição de movimentos em tubos de polietileno, e a PAP foi registrada continuamente durante 45 min. Após o período de estresse, os animais foram realocados em suas caixas e a PAP foi registrada por mais 30 min. Nos grupos experimentais III e IV, após as microinjeções centrais bilaterais no MeA, a PAP continuou sendo registrada por 75 min em animais sob condições basais e em livre movimento (não estressados). Os animais controles de todos os grupos experimentais receberam microinjeções de salina 0,9%. Os experimentos foram realizados entre 7h00min às 13h00min e os animais não tiveram acesso à água ou ração durante o experimento. Os dados estão expressos como média±E.P.M das variações da PAM e FC. Microinjeções de mepiramina nas doses de 50, 100 e 200 nmol promoveu bloqueio dose-dependente da resposta hipertensiva evocada pelo estresse de restrição. A cimetidina (100 e 200 nmol) atenuou a resposta hipertensiva ao estresse apenas na maior dose utilizada. A resposta anti-hipertensiva ao estresse foi maior nos animais que receberam microinjeções de mepiramina do que de cimetidina nas mesmas doses. Nenhuma das drogas alterou a resposta taquicárdica típica do estresse. Mepiramina ou cimetidina foram incapazes de alterar a PAM ou a FC de animais não estressados. Os dados sugerem que as vias histaminérgicas presentes no MeA medeiam a resposta pressora sem alterar a taquicardia evocadas pelo estresse de restrição, ativando preferencialmente os receptores do tipo H1. Além disto, os dados confirmam a hipótese de que a via histaminérgica no MeA não exerce modulação tônica do sistema cardiovascular. A obtenção de dados adicionais relativos ao papel fisiológico dos receptores histaminérgicos centrais no controle das funções cardiovasculares se reveste de grande importância para as ciências biológicas e para a clínica médica, principalmente quando vinculada à variável estresse. Os resultados deste trabalho contribuem para o esclarecimento da participação destes receptores no controle das funções cardiovasculares. / Repeated long lasting experiences of stress situations may result in various pathologic states such as arterial hypertension, cardiac dysrhythmias, myocardial stroke and even sudden death. Although there is a lot information about the neural control of the arterial blood pressure, especially by the brain stem and some other prosencephalic areas, stress-evoked cardiovascular responses are not totally understood. Previews studies shows the importance of the medial amygdala nucleus (MeA) and of the histaminergic neurotransmission on the autonomic control of cardiovascular functions, however there aren’t studies that evidence the role of the histaminergic pathways in MeA on emotional stress-evoked cardiovascular adaptations. Therefore, the aim of this study was investigate the participation of the histaminergic receptors H1 and H2 in MeA on the cardiovascular responses in stressed and non-stressed rats. Wistar rats (280-320g) were submitted to stereotaxic surgery for bilateral cannulation of MeA. Five days after surgery, animals were submitted to catheterization of the left carotid artery. Twenty four hours after catheter insertion, experiments were started and the pulsatile arterial pressure (PAP) of freely moving rats on basal conditions was recorded. Drugs used for central administration were mepyramine (H1 receptors antagonist) on experimental groups I and III and cimetidine (H2 receptors antagonist) on experimental groups II and IV. At experimental groups I and II, 15 min after central microinjections of mepyramine or cimetidine respectively, in different doses, the rats was submitted to restraint stress in a polyvinyl apparatus, and PAP were continuously recorded for 45 min. After stress period, rats were replaced in their own cages and an additional 30 min were recorded for PAP reestablishment. At experimental groups III and IV, after 30 min of basal recording, rats received bilateral central microinjections of mepyramine or cimetidine in a dose of 200 nmol, respectively, and an additional period of 75 min was recorded in freely moving rats on basal conditions. Saline 0,9% was administered as vehicle in control animals of all experimental groups. Mean arterial pressure (MAP) and heart rate (HR) were, then, calculated from the PAP signal. Experiments occurred between 7:00 and 13:00 and rat did not have access to water and food during the sessions. Data were expressed as mean±S.E.M. of MAP and HR variation. Mepyramine microinjections at doses of 50, 100 and 200 nmol promoted dose-dependent blockade of the restraint stress-evoked hypertensive response. Cimetidine (100 and 200 nmol) attenuated the hypertensive response to stress only at the highest dose administered. The anti-hypertensive response was bigger on animals which received mepyramine than cimetidine. Neither drugs altered the typical stress-evoked tachycardiac responses. Indeed, mepyramine or cimetidina were unable to modify the MAP or HR of freely moving rats on basal conditionals (non-stressed rats). These data suggest that histaminergic pathways in MeA mediates pressor responses without modifying the tachycardia promoted by restraint stress, activating preferentially H1 receptors. Besides, data corroborate to the hypothesis that histaminergic pathways in MeA do not plays tonic modulation of the cardiovascular system. Additional information acquired about physiologic role of central histaminergic receptor on the cardiovascular functions is important to biological science and to medical practice, especially when linked to the stress factor. These data contribute to clarify the role of these receptors on cardiovascular functions

The medial amygdala (MeA) is a central node in the interwoven circuits that regulate social behavior based on pheromones. Aromatase-expressing (arom+) neurons in the MeA are key for the establishment and maintenance of sex differences. Here, we characterized the intrinsic electrophysiological properties of arom+ neurons and non-aromatase (arom-) neurons in the MeA of male and female mice. Most electrophysiological properties were similar for arom+ neurons in the MeA between sexes, but the relative refractory period was twice as large in female mice. We also show that the firing pattern and firing frequency is markedly different between arom+ and arom- neurons. The activity of MeA neurons could be modulated by estradiol, which reduced activity in arom+ neurons in males. The differences between arom+ and arom- neurons were observed in both sexes suggesting that aromatase expression delineates a neural population in the MeA with similar and unique electrophysiological properties.