Efeito do tratamento cr?nico com losartana, um bloqueador de receptores AT1, sobre o comportamento de ansiedade, depress?o e mem?ria de ratas ovariectomizadas.

Programa de P?s-Gradua??o em Ci?ncias Biol?gicas. N?cleo de Pesquisas em Ci?ncias Biol?gicas, Pr?-Reitoria de Pesquisa de P?s Gradua??o, Universidade Federal de Ouro Preto.A priva??o de horm?nios ovarianos est? associada a disfun??es em mulheres na p?s-menopausa. Estudos em roedores t?m revelado que a priva??o hormonal induz altera??es no hipocampo e am?gdala, e tamb?m na atividade do sistema renina angiotensina (SRA). A alta express?o de receptores AT1 no hipocampo e na am?gdala sugere o envolvimento do SRA na modula??o de respostas emocionais e cognitivas. De fato, tem-se notado o efeito neuroprotetor dos bloqueadores de receptores de AT1 (BRAs), no entanto, at? onde sabemos, n?o h? estudos avaliando os efeitos dos BRAs sobre as respostas cognitivas e emocionais resultantes da deple??o de horm?nios ovarianos. Assim, o objetivo deste trabalho ? avaliar o efeito da priva??o de horm?nios ovarianos e do tratamento cr?nico com o BRA losartana sobre o comportamento do tipo ansiedade, depress?o e mem?ria de ratas ovariectomizadas. Inicialmente avaliamos o efeito do ciclo estral e de 3 semanas de ovariectomia (OVX) em ratas Fischer sobre o comportamento do tipo ansiedade e p?nico avaliados no labirinto em T elevado (LTE), campo aberto (CA) e paradigma claro/escuro (PCE). Adiante, ratas Long Evans Sham e OVX foram tratadas com ?gua natural ou losartana durante 10 semanas. A partir da quinta semana as ratas foram avaliadas no labirinto em cruz elevado (LCE), CA, reconhecimento de novo objeto (RNO), amea?a iminente (AI) e prefer?ncia por sacarose (PS). O enc?falo foi removido na 10? semana para avalia??o da express?o g?nica de AT1 no hipocampo. Nossos resultados mostram que o ciclo estral n?o influenciou o comportamento do tipo ansiedade e p?nico. J? as ratas OVX apresentaram aumento da lat?ncia de esquiva inibit?ria (Esq1: p = 0,038; Esq2: p = 0,032, teste Mann-Whitney) e redu??o da fuga no LTE (Fuga 1: p = 0,012, teste t de Student), e menor tempo no compartimento claro do PCE (p = 0,0417, teste t de Student), que s?o respostas indicativas de ansiedade. A ovariectomia tamb?m induziu comportamento do tipo ansiedade no LCE e d?ficit de mem?ria avaliada no RNO. O tratamento com losartana foi capaz de prevenir tais altera??es no grupo OVX, exibindo respostas semelhantes aos grupos Sham-Ve?culo e Sham-Losartana (tempo: F (1, 52) = 7,78; p = 0,007; entradas: F (1, 52) = 5; p = 0,02; IR: F (1, 55) = 4,99; p = 0,03, two-way ANOVA). O tratamento aumentou a express?o de RNAm de AT1 no hipocampo (F (1, 23) = 4,74; p = 0,04, two-way ANOVA) independente da cirurgia. Conclu?mos que o tratamento com losartana foi capaz de prevenir comportamento do tipo ansiedade e d?ficit de mem?ria de curto prazo induzidos pela priva??o de horm?nios ovarianos em ratas ovariectomizadas.The loss of ovarian hormones is associated with the onset dysfunctions in postmenopausal women. Studies in rodents has reported that hormonal induces changes the hippocampus and amygdala, and also in renin angiotensin system (RAS) activity. The high expression of AT1 receptors in the hippocampus and amygdala suggests the involvement of RAS in the modulation of emotional and cognitive responses. Indeed, the neuroprotective effect of AT1 receptor blockers (ARBs) has been noted, however, in our knowledge, there are no studies assessing the effects of BRAs on the cognitive and emotional responses resulting from depletion of ovarian hormones. Thus, the aim of this study is to assess the effect of losartan chronic treatment on the anxiety-, depression-like behavior and short-term memory induced by loss of ovarian hormones in ovariectomized rats. Firstly, we assessed the effect of estrus cycle phase and 3 weeks of ovariectomy (OVX) on anxiety- and panic-like behavior of Fischer rats assessed in the elevated T maze (ETM), open field (OF) and dark/light paradigm (DLP). Afterwards, Long Evans rats Sham and OVX were treated with water or losartan along 10 weeks. Five weeks after post-surgery, rats were assessed in the elevated plus maze (EPM), OF, novel object recognition (NOR), looming threat (LT) and sucrose preference (SP) tests. Brain was collected in the 10th week for AT1 gene expression the hippocampus. Our results reveals that the cycle did not influence anxiety- and panic-like behavior. On the other hand, OVX group showed increased inhibitory avoidance latency (Av1: p = 0.038; Av2: p = 0.032, Mann-Whitney test) and reduced escape latency in the ETM (Esc1: p = 0.012, Student's t test) and time exploring the light compartment of DLP (p = 0.0417, Student's t test), which are indicative responses of anxiety. Ovariectomy also induced anxiety-like behavior in EPM and declarative memory deficit assessed in NOR. Losartan treatment was able to prevent anxiety-like behavior and memory deficit in the OVX group, who exhibited responses similar to the Sham-Vehicle and Sham-Losartan groups (time: F (1, 52) = 7.78, p = 0.007; entries: F (1, 52) = 5, p = 0.02, IR: F (1, 55) = 4.99, p = 0.03, two-way ANOVA). The treatment increased the expression of AT1 mRNA in the hippocampus (F (1, 23) = 4,74; p = 0,04, two-way ANOVA) independently of the surgery. We conclude that the losartan chronic treatment was able to prevent anxiety-like behavior and short-term memory deficit induced by loss of ovarian hormone in ovariectomized rats

New insights on amygdala : basomedial amygdala regulates the physiological response to social novelty.

The amygdala has been associated with a variety of functions linked to physiological, behavioral and endocrine responses during emotional situations. This brain region is comprised of multiple sub-nuclei. These subnuclei belong to the same structure, but may be involved in different functions, thereby making the study of each sub-nuclei important. Yet, the involvement of the basomedial amygdala (BMA) in the regulation of emotional states has yet to be defined. Therefore, the aim of our study was to investigate the regulatory role of the BMA on the responses evoked during a social novelty model and whether the regulatory role depended on an interaction with the dorsomedial hypothalamus (DMH). Our results showed that the chemical inhibition of the BMA by the microinjection of muscimol (c-aminobutyric acid (GABAA) agonist) promoted increases in mean arterial pressure (MAP) and heart rate (HR), whereas the chemical inhibition of regions near the BMA did not induce such cardiovascular changes. In contrast, the BMA chemical activation by the bilateral microinjection of bicuculline methiodide (BMI; GABAA antagonist), blocked the increases in MAP and HR observed when an intruder rat was suddenly introduced into the cage of a resident rat, and confined to the small cage for 15 min. Additionally, the increase in HR and MAP induced by BMA inhibition were eliminated by DMH chemical inhibition. Thus, our data reveal that the BMA is under continuous GABAergic influence, and that its hyperactivation can reduce the physiological response induced by a social novelty condition, possibly by inhibiting DMH neurons

Tobacco-free cigarette smoke exposure induces anxiety and panic-related behaviours in male wistar rats.

Smokers, who generally present with lung damage, are more anxious than non-smokers and have an associated augmented risk of panic. Considering that lung damage signals specific neural pathways that are related to affective responses, the aim of the present study was to evaluate the influence of pulmonary injury on anxiety and panic-like behaviours in animals exposed to cigarette smoke with and without tobacco. Male Wistar rats were divided into the following groups: a control group (CG); a regular cigarette group (RC); and a tobacco-free cigarette (TFC) group. Animals were exposed to twelve cigarettes per day for eight consecutive days. The animals were then exposed to an elevated T-maze and an open field. The RC and TFC groups presented increases in inflammatory cell inflow, antioxidant enzyme activity, and TBARS levels, and a decrease in the GSH/GSSG ratio was observed in the TFC group. Exposure to RC smoke reduced anxiety and panic-related behaviours. On the other hand, TFC induced anxiety and panic-related behaviours. Thus, our results contradict the concept that nicotine is solely accountable for shifted behavioural patterns caused by smoking, in that exposure to TFC smoke causes anxiety and panic-related behaviours.Cigarette smoke exposure is associated with anxiety states. Smokers are more anxious than non-smokers1, while cigarette smoking cessation is associated with increased levels of anxiety and stress, as the nicotine in cigarettes has been shown to have anxiolytic effects2. Moreover, smoking is also associated with an augmented risk of panic attacks, and quitting smoking could help reduce this risk3. Importantly, in a study conducted by Amaring and colleagues, it was reported that 72% of panic disorder patients declared that they were regular smokers at the onset of their disease4. Cigarette smoke is also one of the several agents and environmental factors that can trigger oxidative stress and pulmonary damage5. Cigarette smoke causes cellular recruitment, lipid peroxidation, production of inflammatory mediators, and oxidative stress6?11. For instance, studies in mice have shown that exposure to short-term cigarette smoke evokes an increase in inflammatory cell inflow and oxidative damage6,9. In general, exposure to pollutants induces pulmonary inflammation through the generation of oxidative stress12,13, defined as the imbalance in reactive oxygen species production, to the detriment of the antioxidant defence systems14. Importantly, exposure to ambient air particles not only induces pulmonary inflammation but also behavioural disorders both in humans and in mice15. Currently, the majority of anxiety studies associated with cigarette smoking have focused on the anxiolytic effects of nicotine2. However, it has been shown that lung damage can induce central nervous system responses by activating specific neuronal pathways16,17, which include those linked to affective responses, such as anxiety and panic18. This raises the question of whether the anxiety and panic-type behaviour associated with smoking might be related not only to the nicotine or to tobacco?s other constituents but also to lung damage

Influência do estradiol nos transtornos de ansiedade em ratas fischer submetidas à restrição alimentar.

Programa de Pós-Graduação em Ciências Biológicas. Núcleo de Pesquisas em Ciências Biológicas, Pró-Reitoria de Pesquisa de Pós Graduação, Universidade Federal de Ouro Preto.A restrição alimentar (RA) severa provoca carência energética e nutricional, levando o indivíduo à desnutrição, e este desequilíbrio acarreta inúmeras consequências bioquímicas, fisiológicas e emocionais, tais como ansiedade, pânico e danos à memória. A RA também altera o eixo hipotálamo-hipófise-ovariano, que é a via que regula a produção de hormônios ovarianos, ocasionando redução da síntese e liberação de estradiol pelos ovários. Estudos já mostraram a presença de alta densidade de receptores de estradiol do tipo β (ERβ), que quando ativados resultam em efeitos ansiolíticos, no núcleo dorsal da rafe (NDR), indicando uma relação entre o estradiol, o NDR e os transtornos de ansiedade generalizada (TAG) e pânico (TP). O NDR é uma importante estrutura encefálica relacionada a fisiopatologia dos TAG e TP. Assim, o nosso objetivo foi investigar a influência do estradiol no NDR sobre os TAG e TP em ratas submetidas à restrição alimentar. Para isso, ratas Fischer (210 ± 10g) foram divididas em controle (C) e RA e alojadas individualmente. Durante 14 dias, as C receberam ração ad libitum e os RA receberam 40% do consumo médio dos C. No 7° dia foram anestesiados com ketamina-xilazina para inserção de cânula-guia no NDR. Ao final do protocolo dietético, cada animal foi submetido a dois diferentes ensaios nos quais ou veículo (100 nL) ou estradiol (0,5 pmol/100 nL) foi microinjetado no NDR. Após 20 minutos, as ratas foram testadas no Labirinto em T Elevado (LTE), um aparato que permite avaliar comportamento de ansiedade (esquiva inibitória) e pânico (fuga), e em seguida foram colocadas no campo aberto (arena retangular que avalia a atividade locomotora do animal). No dia seguinte as ratas foram testadas novamente no LTE para avaliação da memória. Nossos resultados mostraram que as ratas RA possuíam o útero mais leve em relação às controle, sugerindo que a RA alimentar reduz a concentração de hormônios ovarianos, entre eles o estradiol. Nós também observamos que as RA levaram mais tempo para sair do braço fechado do LTE do que as C durante a esquiva (F1,63 = 10,34; p = 0,0021), o que indica um comportamento mais ansioso pelo grupo RA. Ao microinjetarmos estradiol no NDR das ratas RA, notou-se que o tempo de latência de saída do braço fechado diminuiu em relação àquelas que receberam veículo (F2,65 = 3,86; p=0,0002), sugerindo que o estradiol agiu como um agente ansiolítico revertendo o comportamento ansioso apresentado no experimento anterior. Não encontramos diferença entre os grupos no tempo de latência de fuga. Em relação à atividade locomotora, as ratas RA mostraram-se menos ativas que as C, independente da administração de veículo ou estradiol no NDR. Diante disso, sugerimos que a RA reduz os níveis de estradiol, diminuindo a concentração disponível deste XVI hormônio para atuar nos receptores presentes no NDR, o que altera o controle desta via, levando ao comportamento de ansiedade nas ratas.Food restriction (FR) causes severe energy and nutritional deficiency, leading the individual to malnutrition, and this imbalance results in countless biochemical, physiological and emotional consequences such as anxiety, panic and damage to memory. FR also alters the hypothalamic-pituitary-ovarian axis, which is the pathway that regulates the production of the ovarian hormones, leading to reduced synthesis and release of estradiol by the ovaries. Studies have shown the presence of high density estradiol β receptors (ERβ) in the dorsal raphe nucleus (DRN) indicating a relationship among estradiol, the DRN and generalized anxiety disorder (GAD) and panic disorder (PD). The DRN is an important brain structure related to the pathophysiology of GAD and PD. Thus, the aim of our study was to investigate the influence of estradiol in DRN on GAD and PD in female rats subjected to food restriction. For that, female Fischer rats (210 ± 10g) were divided into control (C) and FR and housed individually. For 14 days, the C received food ad libitum and FR received 40% of the average consumption of the C. On the 7th day the rats were anesthetized with ketamine-xylazine for insertion of the cannula guide in the DRN. At the end of the dietary protocol, each animal was subjected to two tests in which vehicle (100 nL) or estradiol (0.5 pmol / 100 nl) was microinjected in the DRN. After 20 minutes, the rats were tested in the Elevated T Maze (ETM), an apparatus designed to measure anxiety behavior (inhibitory avoidance) and panic (escape), and then were placed in the open field (rectangular arena that measures the locomotor activity). The following day the rats were retested in the ETM for the evaluation of memory. Our results show that FR had a lower weight of the rat uterus than the control, suggesting that FR reduces the concentration of ovarian hormones including estradiol. We have also found that the FR animals showed increased avoidance latences (F1,63 = 10.34, p = 0.0021), which indicates a more anxious behavior by the FR group. After the estradiol microinjetion in the DRN of FR rats, we noted that the latency to leave the closed arm decreased compared to those animals receiving vehicle (F2,65 = 3.86; p = 0.0002), suggesting that estradiol acted as an anxiolytic agent reversing the anxious behavior presented in the previous experiment. We found no difference between groups in the escape latency time. Regarding the locomotor activity of the FR rats were less active than C, regardless of vehicle or estradiol administration in the DRN Therefore, we suggest that FR reduces estradiol levels, reducing the available concentration of this hormone to act on receptors in the DRN, which changes the control of this pathway, leading to anxiety behavior in rats

High fat diet induced-obesity facilitates anxiety-like behaviors due to GABAergic impairment within the dorsomedial hypothalamus in rats.

Overweight and obesity are conditions associated with an overall range of clinical health consequences, and they could be involved with the development of neuropsychiatric diseases, such as generalized anxiety disorder (GAD) and panic disorder (PD). A crucial brain nuclei involved on the physiological functions and behavioral responses, especially fear, anxiety and panic, is the dorsomedial hypothalamus (DMH). However, the mechanisms underlying the process whereby the DMH is involved in behavioral changes in obese rats still remains unclear. The current study further investigates the relation between obesity and generalized anxiety, by investigating the GABAA sensitivity to pharmacological manipulation within the DMH in obese rats during anxiety conditions. Male Wistar rats were divided in two experimental groups: the first was fed a control diet (CD; 11% w/w) and second was fed a high fat diet (HFD; 45% w/w). Animals were randomly treated with muscimol, a GABAA agonist and bicuculline methiodide (BMI), a GABAA antagonist. Inhibitory avoidance and escape behaviors were investigated using the Elevated T-Maze (ETM) apparatus. Our results revealed that the obesity facilitated inhibitory avoidance acquisition, suggesting a positive relation between obesity and the development of an anxiety-like state. The injection of muscimol (an anxiolytic drug), within the DMH, increased the inhibitory avoidance latency in obese animals (featuring an anxiogenic state). Besides, muscimol prolonged the escape latency and controlling the possible panic-like behavior in these animals. Injection of BMI into the DMH was ineffective to produce an anxiety-like effect in obese animals opposing the results observed in lean animals. These findings support the hypotheses that obese animals are susceptible to develop anxiety-like behaviors, probably through changes in the GABAergic neurotransmission within the DMH

Estrogen receptor ? activation within dorsal raphe nucleus reverses anxiety-like behavior induced by food restriction in female rats.

Severe food restriction (FR), as observed in disorders like anorexia nervosa, has been associated to the reduction of estrogen levels, which in turn could lead to anxiety development. Estrogen receptors, mainly ER? type, are commonly found in the dorsal raphe nucleus (DRN) neurons, an important nucleus related to anxiety modulation and the primary source of serotonin (5-HT) in the brain. Taking together, these findings suggest an involvement of estrogen in anxiety modulation during food restriction, possibly mediated by ER? activation in serotonergic DRN neurons. Thus, the present study investigated the relationship between food restriction and anxiety-like behavior, and the involvement of DRN and ER? on the modulation of anxiety-like behaviors in animals subjected to FR. For that, female Fischer rats were grouped in control group, with free access to food, or a FR group, which received 40% of control intake during 14 days. Animals were randomly treated with 17?-estradiol (E2), DPN (ER? selective agonist), or their respective vehicles, PBS and DMSO. Behavioral tests were performed on Elevated T-Maze (ETM) and Open Field (OF). Our results suggest that FR probably reduced the estrogen levels, since the remained in the non-ovulatory cycle phases, and their uterine weight was lower when compared to control group. The FR rats showed increased inhibitory avoidance latency in theETM indicating that FR is associated with the development of an anxiety-like state. The injections of both E2 and DPN into DRN of FR animals had an anxiolytic effect. Those data suggest thatanxiety-like behavior induced by FR could be mediated by a reduction of ER? activation in the DRN neurons, probably due to decreased estrogen levels

Association of high-fat diet with neuroinflammation, anxiety-like defensive behavioral responses, and altered thermoregulatory responses in male rats.

Overweight and obesity are a worldwide pandemic affecting billions of people. These conditions have been associated with a chronic low-grade inflammatory state that is recognized as a risk factor for a range of somatic diseases as well as neurodevelopmental disorders, anxiety disorders, trauma- and stressor-related disorders, and affective disorders. We previously reported that the ingestion of a high-fat diet (HFD; 45% fat kcal/g) for nine weeks was capable of inducing obesity in rats in association with increased reactivity to stress and increased anxiety-related defensive behavior. In this study, we conducted a nine-week diet protocol to induce obesity in rats, followed by investigation of anxiety-related defensive behavioral responses using the elevated T-maze (ETM), numbers of FOS-immunoreactive cells after exposure of rats to the avoidance or escape task of the ETM, and neuroinflammatory cytokine expression in hypothalamic and amygdaloid nuclei. In addition, we investigated stress-induced cutaneous thermoregulatory responses during exposure to an open-field (OF). Here we demonstrated that nine weeks of HFD intake induced obesity, in association with increased abdominal fat pad weight, increased anxiety-related defensive behavioral responses, and increased proinflammatory cytokines in hypothalamic and amygdaloid nuclei. In addition, HFD exposure altered avoidance- or escape task-induced FOSimmunoreactivity within brain structures involved in control of neuroendocrine, autonomic, and behavioral responses to aversive stimuli, including the basolateral amygdala (BLA) and dorsomedial (DMH), paraventricular (PVN) and ventromedial (VMH) hypothalamic nuclei. Furthermore, rats exposed to HFD, relative to control dietfed rats, responded with increased tail skin temperature at baseline and throughout exposure to an open-field apparatus. These data are consistent with the hypothesis that HFD induces neuroinflammation, alters excitability of brain nuclei controlling neuroendocrine, autonomic, and behavioral responses to stressful stimuli, and enhances stress reactivity and anxiety-like defensive behavioral responses