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The primary goal of this study was to explore the social buffering effect that humans offer to goats and dogs with limited exposure to human socialization, particularly in situations involving interactions with unfamiliar humans. A total of 13 dogs and 14 goats were selected for the study, all of which had limited prior socialization with humans. Each animal was placed in a testing room with unfamiliar humans for 15 min. Three experimenters aimed to establish a comfortable environment, encouraging social interaction by offering food to the animals and assessing the animals' willingness to accept food and their response to being approached and petted. If both conditions were satisfied, the animals were classified as "social". If one or none of the conditions were met, the animals were classified as "not social". Cortisol levels were measured by collecting blood samples before and after the test. Non-parametric tests together with a GzLM showed that the effect of human social buffering in goats was different in comparison to dogs: goats exhibited higher cortisol levels after the test, while dogs did not show a significant change. Further analysis demonstrated that "social" goats had a lower likelihood of experiencing significant changes in cortisol levels than dogs. Thus, once human interactions are accepted, both species could benefit from social buffering. In summary, this study enhances our understanding of how dogs and goats respond to social interactions with humans in the social buffering effect.
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Cabras , Hidrocortisona , Humanos , Cães , Animais , Alimentos , ProbabilidadeRESUMO
The reproductive performances of livestock play an essential role in the economic management of the farm. The improvement of semen quantity and quality through the use of food supplements that lack substances which are forbidden in animal feeding, or that may have detrimental effects, is an important goal. Maca (Lepidium meyenii) is a plant that has been used for centuries in the Andes for nutrition and fertility enhancement in humans and animals. The aim of this study was to evaluate the effects of food supplementation of stallions with maca during the breeding season on spermatozoa parameters such as DNA fragmentation and shape, which are two predictive indexes of spermatozoa functionality. For this purpose, ejaculate volume, semen gel-free volume, sperm concentration and motility, total sperm count, sperm DNA fragmentation and sperm head parameters (length, width, perimeter, area, shape factor, roughness) were measured in four stallions. Maca food supplementation in stallions during breeding reduced the percentage of spermatozoa with fragmented DNA, increased significantly sperm concentration and exerted an elongation of the spermatozoa head, a condition that is believed to improve spermatozoa functionality, suggesting that food supplementation of maca could be useful in horse breeding during the breeding season.
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Fragmentação do DNA , Lepidium , Animais , Suplementos Nutricionais , Cavalos , Masculino , Melhoramento Vegetal , EspermatozoidesRESUMO
A nutritional supplement (Ecotrofin™, by Vetoquinol Italia S.r.l) recommended in ruminants feeding to strengthen the physiological condition and improve digestive performance was tested in 20 pluriparae grazing goats divided in two groups (control and treated) to assess its possible effects on milk yield and quality and to assess eventual adverse effects. Animals from both groups also received 400 g/day of corn meal, and the treated group was supplemented with 20 g/head/day of the nutritional supplement. At the doses suggested by the manufacturer, despite a transient increase after 30 days of supplementation, Ecotrofin™ did not show significant effects on milk yield and, although some changes were found in the fatty acids profile, no significant improvement of MUFA and PUFA, as well as of omega-6:omega-3 ratio and CLA content were seen. Therefore, in our experimental conditions the supplementation of diet with Ecotrofin™ did not appear useful to improve goat's performance. A significant effect on kidney health markers (27 vs. 22.5 for urea and 0.83 vs. 0.76 for creatinine, p < 0.05) suggested a beneficial effect on renal function but, since levels fell in the normal ranges in both groups, such hypothesis would need further studies to be addressed.
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Lactação , Leite , Fenômenos Fisiológicos da Nutrição Animal , Animais , Dieta/veterinária , Suplementos Nutricionais , Ácidos Graxos , Feminino , CabrasRESUMO
The present study was conducted to assess the chemical composition of Yellow Maca (Lepidium meyenii) and its biological activity on stallions following oral administration of hypocotyl powder. Maca was subjected to methanolic extraction and the chemical analysis was carried out by LC-MS-QTOF (liquid chromatography-mass spectrometry). Our results showed that Maca contains some effective antioxidants, a high percentage of glucosinolates, and other important components with a high antioxidant capacity. To evaluate the plant biological activity in stallions fed with Maca powder for 60 days, the redox status and some reproductive parameters were investigated. Blood and semen samples were collected at 0, 30, 60, and 90 days from the beginning of this study. Blood samples showed a decrease of the reactive oxygen metabolites, evaluated by d-ROMs test, and an increase of the antioxidant barrier in terms of biological antioxidant potential (BAP test), powerful oxidant capacity (OXY-Adsorbent test), and thiols evaluation (-SHp test). Furthermore, semen samples showed a positive trend during Maca administration in the following parameters: ejaculate volumes and sperm concentrations, total and progressive motility, and acrosome integrity.
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Antioxidantes/administração & dosagem , Lepidium/química , Compostos Fitoquímicos/administração & dosagem , Sêmen/fisiologia , Administração Oral , Animais , Antioxidantes/química , Antioxidantes/farmacologia , Cromatografia Gasosa-Espectrometria de Massas , Glucosinolatos/administração & dosagem , Glucosinolatos/química , Glucosinolatos/farmacologia , Cavalos , Hipocótilo/química , Masculino , Oxirredução/efeitos dos fármacos , Compostos Fitoquímicos/química , Compostos Fitoquímicos/farmacologia , Pós/administração & dosagem , Espécies Reativas de Oxigênio/sangue , Sêmen/efeitos dos fármacos , Motilidade dos Espermatozoides/efeitos dos fármacosRESUMO
Excessive energy intake may evoke complex biochemical processes characterized by inflammation, oxidative stress, and impairment of mitochondrial function that represent the main factors underlying noncommunicable diseases. Because cow milk is widely used for human nutrition and in food industry processing, the nutritional quality of milk is of special interest with respect to human health. In our study, we analyzed milk produced by dairy cows fed a diet characterized by a high forage:concentrate ratio (high forage milk, HFM). In view of the low n-6:n-3 ratio and high content of conjugated linoleic acid of HFM, we studied the effects of this milk on lipid metabolism, inflammation, mitochondrial function, and oxidative stress in a rat model. To this end, we supplemented for 4 wk the diet of male Wistar rats with HFM and with an isocaloric amount (82 kJ, 22 mL/d) of milk obtained from cows fed a diet with low forage:concentrate ratio, and analyzed the metabolic parameters of the animals. Our results indicate that HFM may positively affect lipid metabolism, leptin:adiponectin ratio, inflammation, mitochondrial function, and oxidative stress, providing the first evidence of the beneficial effects of HFM on rat metabolism.
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Ração Animal , Indústria de Laticínios , Suplementos Nutricionais , Inflamação/terapia , Leite/química , Mitocôndrias/fisiologia , Estresse Oxidativo , Ração Animal/análise , Animais , Bovinos , Feminino , Inflamação/prevenção & controle , Masculino , Ratos , Ratos WistarRESUMO
This study aimed to investigate the antidiabetic, antilipidaemic and antioxidant activities of Citrus medica cv Diamante (Rutaceae) hydroalcoholic (CD) peel extract in Zucker diabetic fatty (ZDF) rats. The ability of CD to protect against oxidative stress was investigated by using different in vitro assays and in vivo by using the reactive oxygen metabolites-derived compounds (d-ROMs) test and the biological antioxidant potential test (BAP). Two different doses of CD extract (300 and 600 mg/kg/die) were administered at ZDF rats for 4 weeks. CD reduced cholesterol and triglycerides levels. A dose-dependent effect on body weight and serum glucose levels was observed. A decrease of d-ROMs and an increase of BAP were recorded by using the dose of 600 mg/kg. The extract inhibited lipid peroxidation (IC50 value of 0.23 mg/ml). These findings suggest as an efficient phytotherapeutic approach in combating hyperlipidaemic and hyperglycaemic disorders.
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Glicemia/metabolismo , Citrus/química , Diabetes Mellitus Experimental/sangue , Estresse Oxidativo/efeitos dos fármacos , Extratos Vegetais/farmacologia , Animais , Masculino , Ratos , Ratos ZuckerRESUMO
The study aimed to explore how limited human socialization affects the socio-cognitive abilities and interactions with unfamiliar individuals of a selected group of domesticated dogs and goats. These animals were raised and kept under conditions characterized by limited human socialization, and their behavior was assessed using the "impossible task" paradigm. The study found that dogs, with a history of cooperative interactions and human companionship, exhibited more frequent social engagement with human experimenters in the experimental setting than goats, traditionally domesticated for utilitarian purposes. However, differences in interaction duration and latency were not significant, highlighting the complexity of these interactions. The results suggest that domestication history and behavioral ecology play significant roles in shaping animals' willingness to engage with humans. However, this study acknowledges limitations, such as the specific population studied, and calls for further research with larger and more diverse samples to generalize these findings. Understanding the interplay between domestication history, behavioral ecology, and human socialization could provide insights into the complex factors influencing animal-human interactions and cognitive behaviors, with implications for animal welfare and human-animal relationships.
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In this study, we explored the correlations between circulating levels of oxytocin, cortisol, and different social behaviors toward humans in 26 Italian Red Pied calves (all females, with an average age of 174 ± 24 days) using the impossible task paradigm. This paradigm has proved fruitful in highlighting the effect of socialization on the willingness to interact with humans in several domesticated species. The test consists of the violation of an expectation (recovering food from an experimental apparatus) while a caregiver and a stranger are present. Immediately after the end of the test (less than one minute), blood was collected from the coccygeal vein. Statistics were performed by the Spearman's rank correlation; significant differences were adjusted according to Bonferroni's correction. Cortisol correlates positively (ρ = 0.565; p < 0.05) with the latency of behaviors directed at the caregiver, and the duration of behaviors directed at the apparatus correlates negatively with both the caregiver (ρ = -0.654; p < 0.05) and a stranger (ρ = -0.644; p < 0.05). Contrary to what is reported in the literature on cows, no correlations were found between oxytocin levels and direct behaviors toward the caregiver. This highlights a different behavioral strategy between calves and cows when placed in front of an impossible task.
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Cyclosporin A (CsA) is the prototype of immunosuppressant drugs that have revolutionized the management of all transplantation and autoimmune diseases. Side effects of CsA mainly affecting the kidney but also observed in liver and heart, limit the therapeutic use of this drug after organ transplantation. The renal toxicity of CsA is attributed to reduced renal blood flow which leads to hypoxia-reoxygenation injury accompanied by excessive generation of oxygen-derived free radicals. In several therapeutic protocols, CsA is used in association with corticosteroids to obtain better therapeutic results. Recently, our studies showed that hydrocortisone (HY) has a protective effect on CsA-induced cardiotoxicity. In fact our previous results demonstrated that in rat cardiomyocytes, CsA toxicity is due to a calcium overload, which in turn induce lipid peroxidation and determines oxidative stress-induced cell injury. Treatment with HY effectively inhibits CsA-induced toxicity, decreasing lipid peroxidation as well as calcium intracellular concentration. In this study we evaluated in vivo the effects of CsA, used alone or in association with HY, on some parameters of renal dysfunction (blood urea nitrogen; BUN, creatinine, and cholesterol), malondialdheyde (MDA) levels, antioxidant enzyme catalase (CAT), superoxide dismutase (SOD), glutathione peroxidase (GPx), and apoptosis. CsA administration for 24 days resulted in a marked renal oxidative stress, which significantly deranged the renal functions. Treatment with CsA in association with HY significantly improved the renal dysfunction and renal oxidative status. This study clearly suggests the role of oxidative stress in the pathogenesis of CsA-induced nephrotoxicity.
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Ciclosporina/toxicidade , Hidrocortisona/uso terapêutico , Imunossupressores/toxicidade , Nefropatias/induzido quimicamente , Animais , Apoptose , Pressão Sanguínea/efeitos dos fármacos , Catalase/metabolismo , Colesterol/sangue , Creatinina/sangue , Glutationa Peroxidase/metabolismo , Nefropatias/tratamento farmacológico , Nefropatias/metabolismo , Peroxidação de Lipídeos/efeitos dos fármacos , Masculino , Ratos , Ratos Wistar , Superóxido Dismutase/metabolismo , Ureia/sangueRESUMO
This study explored a possible relationship between the circulating oxytocin, cortisol, and the willingness of dairy cows to engage in social behaviors with humans in an experimental context. The behaviors of twenty-nine cows were recorded during the impossible task paradigm, a procedure aimed at creating a violation of expectancy, in the presence of the caregiver and a stranger. The results showed that serum oxytocin levels were positively correlated with duration and negatively correlated with the latency of the cows' social interactions with the caregiver. This research provides a clear correlation between circulating oxytocin and a willingness to engage in social contact with the caregiver, excluding the possible effect of different cortisol levels on such behavior.
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This study aimed to investigate the efficacy of DìRelaxTM, a nutraceutical formulated to reduce anxiety in dogs, using a randomized controlled trial (RCT) design. The C-BARQ questionnaire, some clinical investigations, and the impossible task test were performed in dogs before and after treatment. The C-BARQ questionnaire is particularly useful for assessing the frequency and severity of problematic behaviors. The impossible task paradigm provides insight into the decision-making processes in the realm of expectancy frustration. Results showed an ameliorative effect on the performances of treated dogs during the solvable phases, with a significant decrease in the time needed to solve the task. No behavioral difference was found between treated and untreated anxious dogs during the unsolvable phase. According to the results from the C-BARQ questionnaire, some of the behaviors appeared to improve. Clinical investigations, including a complete blood cell count and blood chemistry, showed no difference between groups, thus suggesting the safety of the product. In general, this study suggests that DìRelaxTM can be safely administered with no adverse effects and can exercise a beneficial effect on anxious dogs by enhancing their cognitive abilities, but further studies should investigate the best method of administration.
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Hermetia illucens larvae (five days old) were farmed on broiler feed (control diet), a vegetable diet (V100), a 50% of vegetable diet + 50% of butchery wastes (V50 + B50), and a 75% of vegetable diet + 25% of butchery wastes (V75 + B25) to evaluate their suitability. Ten kilograms of substrate and 6000 larvae composed each replicate (nine per group). Larvae were weighed and measured every two days until the 25% developed into prepupae. Larval mortality and growing indexes were calculated. Substrates, larvae, and frass chemical composition were analyzed. Larvae oxidative status and stability were measured in hemolymph and body. The V100 larvae showed the lowest live weight, length, thickness, and growth rate but had low mortality rate and high substrate reduction index and protein conversion ratio. The V100 larvae had similar protein to and lower lipids than the control ones, while the V50 + B50 and V75 + B25 larvae contained higher lipids and lower protein than the others. Despite the vegetable wastes, at different levels, the reactive oxygen species content decreased in hemolymph, and the V100 diet depressed growth performance and should be avoided. The use of butchery wastes combined with vegetable ingredients can be a suitable alternative to balance the high level of lipid and the low content of protein.
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Edible flowers consumption has increased in recent years due to their rich content of healthy phytochemicals. The aim of this study was to analyse the chemical profile of Cucurbita pepo L. flowers, and to explore their antioxidant and hypoglycaemic properties. Moreover, in order to assess in vivo effects, biochemical analysis, Reactive Oxygen Metabolites (d-ROMs) and Biological Antioxidant Potential (BAP) tests were performed on mice serum. High Performance Liquid Chromatography-Diode Array Detection (HPLC-DAD) analyses revealed the presence of (+)-catechin, (-)-epicatechin, rutin, and syringic acid as main constituents. 2,2'-Azino-bis (3-ethylbenzothiazoline-6-sulphonic acid) (ABTS) and Ferric Reducing Antioxidant Power (FRAP) tests showed interesting results. The extract exhibited the strongest inhibitory effect on α-glucosidase (IC50 of 144.77 µg/mL). In vivo results confirmed the hypoglycaemic effects, also affecting lipid metabolism but did not revealed benefits on ROS production. These results may add some information supporting the use of C. pepo flowers as functional foods and/or nutraceuticals.
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Antioxidantes/farmacologia , Cucurbita , Animais , Antioxidantes/isolamento & purificação , Cucurbita/química , Flores/química , Camundongos , Compostos Fitoquímicos/isolamento & purificação , Compostos Fitoquímicos/farmacologia , Extratos Vegetais/farmacologiaRESUMO
The possibility to associate nutraceuticals, as immune-modulating tools, to the treatment of visceral leishmaniosis is a matter of great interest. In this study, we investigated whether the administration of a nutritional supplement (DìLshTM, Dynamopet SRL, Verona, Italy) was able to exert beneficial effects on the inflammatory state and oxidative stress of the dogs naturally infected by Leishmania infantum. To this purpose, specific parameters, namely Tumor Necrosis Factor -alpha (TNF), Interleukin-6 (IL-6), Inteleukin-10 (IL-10), leptin, derivates of Reactive Oxigen Metabolites (d-Roms) and Biological Antioxidant Potential (BAP), as well as the haematological and biochemical profiles of the infected dogs, were investigated upon the treatment with the nutritional supplement and compared with the conventional pharmacological anti-Leishmania therapy. The animals underwent complete clinical examination and blood sample collection before (T0) and 3 months after (T90) the onset of the two treatments. The two treatments showed similar results: significant clinical improvement, ELISA positivity and IgG decrease, an increase in IL-10, and a decrease in IL-6 were observed in animals treated with the nutritional supplement. A decrease in d-Roms and an increase in BAP were also detected in both groups. On the whole, the nutritional supplement possesses anti-inflammatory and antioxidant properties, suggesting that it may support animals' health and be useful to extend the time a drug therapy is needed.
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Throughout their evolutionary history, humans have tried to domesticate a variety of wild terrestrial mammals, resulting in a limited number that has been successfully domesticated. Among these domesticated species, domestic goats (Capra aegagrus hircus) are a useful model species to study the effects of ontogenesis on the socio-cognitive abilities of domestic non-companion animals in their interactions with humans. To this end, the behavioral responses of two groups of goats with a different background of human socialization (high and low socialization) were compared in the impossible task test, an experimental paradigm aimed to study socio-cognitive skills and the tendency to interact with humans. Our results show that, when the task became impossible to solve, goats with a higher level of socialization interacted with the experimenter for a greater amount of time than subjects in the low socialization group, whereas the latter group exhibited increased door directed behavior. Overall, highly socialized goats made more social contact with humans compared to the other group in the impossible task paradigm.
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Aim of the present study was to evaluate the potential beneficial and/or adverse effects of Capsicum annuum L. (cv. Fiesta) extracts at two stage of ripening (immature and mature), and at two dosages (low and high) by evaluation of biochemical profile and oxidative status in CD-1 mice. The extracts were daily administered to mice by oral gavage for 20 days. At the end of the trial, the animals were euthanatized and blood was collected. Evidence of liver damage (increase of AST, ALT and bilirubin) in the group receiving the higher dosage of immature peppers extract were observed. Even if no adverse effects were seen at the lower doses, also no signs of beneficial effects in term of health status, biochemical profile and oxidative status were detected. These results are in contrast with in vitro findings and raise doubts about the possible use of Capsicum annuum L. (cv. Fiesta) as a nutritional supplement.
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Antioxidantes/farmacologia , Capsicum/fisiologia , Extratos Vegetais/efeitos adversos , Extratos Vegetais/farmacologia , Alanina Transaminase/sangue , Animais , Antioxidantes/efeitos adversos , Aspartato Aminotransferases/sangue , Bilirrubina/sangue , Análise Química do Sangue , Capsicum/química , Colesterol/sangue , Relação Dose-Resposta a Droga , Frutas/química , Frutas/fisiologia , Masculino , Camundongos , Camundongos Endogâmicos , Oxidantes/metabolismo , Extratos Vegetais/administração & dosagemRESUMO
A clarification method was proposed to ameliorate the technological quality of fruit juices by preserving bioactive compounds. This study evaluated the in vitro antioxidant and hypoglycemic activities and the in vivo effects of Punica granatum L. natural (NJ) and clarified (CJ) juice by polyvinylidene fluoride (PVDF) hollow fiber membrane. CJ was more active as an antioxidant and as a α-glucosidase inhibitor than NJ. Mice were orally gavaged with water (Control), NJ, and CJ for 28 days. NJ group showed significant decrease of alanine aminotransferase, aspartate amino transferase, and creatine-phosphokinase. CJ administration was associated with urea, creatine-phosphokinase, and triglycerides values significantly lower with respect to the control. Oxidative status was ameliorated with CJ administration, showing a reactive oxygen metabolites (d-ROMs) reduction of 32% and a biological antioxidant potential (BAP) boosting of 23% compared to the control, whereas NJ did not show a similar effect. Results confirmed the beneficial properties of pomegranate juice, showing that membrane clarification may enhance such effects in terms of antioxidant activity.
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The serotonin transporter (SERT) gene has been proposed as a candidate gene responsible for the sudden infant death syndrome (SIDS). In this study, for the first time we obtained a SERT-knockout (KO) mouse model which reproduces SIDS phenotype. SERT-KO mice were generated by mating SERT(Cre/+) heterozygous mice. The SERT-KO mouse embryos at the pre-natal stage E18.5 were lacking of SERT mRNA and protein expression in the heart. A premature death of 75% of SERT-KO mice occurred in the first week after birth. LacZ staining of whole mounts and tissue sections of the heart from SERT(Cre/+);ROSA26R adult mice and E18.5 embryos demonstrated a marked localized expression of SERT in the right ventricle, the conal region, the vasculature, the atrial septum, the ventricular valves, and the sinoatrial node of the conduction system. These data suggest a cardiac phenotype for the sudden death of SERT-KO mice. Histological analysis of heart sections showed that SERT-KO mice develop cardiac fibrosis. Increased collagen accumulation in the myocardium and the valvular and perivascular regions, and enhanced expression of alpha-smooth muscle actin were detected in the heart of SERT-KO mice versus wild-type (WT) mice. Interestingly, higher expression levels of the 5-HT2A receptor and increased levels of phospho-SMAD2/3 and phospho-ERK1/2 were detected in SERT-KO mouse heart versus WT mice. Overall, our findings provide i) new insights into the role of SERT gene in SIDS, and ii) the first in vivo validation of the molecular mechanism involving the activation of TGF-beta1 signalling in the cardiac fibrosis.
Assuntos
Morte Súbita/etiologia , Proteínas da Membrana Plasmática de Transporte de Serotonina/fisiologia , Transdução de Sinais/fisiologia , Fator de Crescimento Transformador beta1/fisiologia , Animais , Animais Recém-Nascidos , Western Blotting , Camundongos , Camundongos Knockout , Proteína Quinase 1 Ativada por Mitógeno/metabolismo , Proteína Quinase 3 Ativada por Mitógeno/metabolismo , Receptor 5-HT2A de Serotonina/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Proteínas da Membrana Plasmática de Transporte de Serotonina/genética , Transdução de Sinais/genética , Proteína Smad2/metabolismo , Proteína Smad3/metabolismo , Fator de Crescimento Transformador beta1/genética , Fator de Crescimento Transformador beta1/metabolismoRESUMO
Both prostate and vestibular glands of mammals contain neuroendocrine cells which synthesize, store and release growth factors including neuropeptides and biogenic amines such as serotonin. An increase of the secretory products by these cells has been correlated to tumour progression and poor prognosis. Serotonin mediates a wide range of physiological functions by binding to multiple receptors on cell surface. However, the entire serotonergic system is mainly regulated by the serotonin transporter SERT which modulates serotonin concentration in extracellular fluid. Primarily located in serotonergic neurons, SERT is also expressed in various cell types in the periphery. In this study, we found a wide distribution of SERT in the parenchymal cells of both the prostate and the vestibular glands of cattle using immunohistochemistry. The expression of SERT mRNA transcripts was assessed by reverse-transcription polymerase chain reaction, thus suggesting that SERT is locally synthesized. Furthermore, Western blotting analysis showed the presence of two isoforms of the protein (70 and 140 kDa), probably corresponding to the high mannose-type SERT and its dimeric form. Our results provide the first evidence for SERT expression in the mammalian genital tract, thus highlighting a new potential target for the therapy of the genital tract cancers.
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Genitália Masculina/metabolismo , Proteínas da Membrana Plasmática de Transporte de Serotonina/metabolismo , Animais , Bovinos , Dimerização , Genitália Masculina/anatomia & histologia , Imuno-Histoquímica , Masculino , Peso Molecular , Próstata/metabolismo , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Proteínas da Membrana Plasmática de Transporte de Serotonina/biossíntese , Proteínas da Membrana Plasmática de Transporte de Serotonina/genética , Distribuição TecidualRESUMO
The aim of this research was to evaluate the possible usefulness of mirrors in improving rabbit behavior in a free-range breeding system. Three groups (each consisting of nine replicates of three animals) were compared: isolated, isolated with mirrors and separated by a wire mesh (possible visual and olfactory contacts). Rabbits allowed to have a visual and olfactory contact showed a significantly higher expression of important natural behaviors (olfactory investigation, gnawing, alertness, stretching, locomotion) compared to the isolated rabbits (with or without mirrors); while rabbits in the mirror group showed higher allo-grooming activity than those isolated and no different locomotion activity than those separated by wire mesh. Thus, mirrors seemed to be able to modify the behavioral repertoire of isolated rabbits by acting on social perception in rabbits reared in small groups in a free-range system. Nevertheless, despite it being advisable to use a combination of different indicators in order to assess the stress level of an animal, the accuracy of serum cortisol, as well as of some secondary stress markers assay, appeared to be limited in this type of breeding.