Effect of tryptophan enriched diets on aggression in hierarchical groups of juvenile Atlantic salmon (Salmo salar)

Aggression in fish hierarchies often leads to one dominant individual monopolizing resources (food/shelter), thus increasing their fitness. Meanwhile individuals in subordinate ranks, endure high stressful conditions, which often lead to disease and higher mortality rates. Levels of serotonin have been linked to agonistic behavior. Enriching fish diets with tryptophan (TRP), the natural precursor for serotonin, has been used in several vertebrate species in order to control aggression levels. Twelve hierarchies consisting of 4 fish in each were studied. Six groups were given a TRP enriched diet and six groups a control diet. Aggressive acts performed and those received were observed during 7 days, 15 min per day, in order to establish hierarchies. Whole brains were excised and immediately frozen from each fish at the end of the trial, in order to analyze the monoamine levels by means of HPLC chromatography. TRP groups were found to have a higher overall aggression compared to control groups. Dividing the groups into their hierarchical ranks, middle rank individuals were overall more aggressive, displaying also more aggressive attacks towards dominants in TRP groups, compared with control groups. These results were opposite to what was expected. Serotonergic activity, measured as the [5-HIAA]/ [5HT] ratio, was higher in more aggressive individuals, which is also opposite from what has been found before. High amounts of received aggression were found to be positively correlated with high [5-HIAA]/ [5HT] ratios. This is the first time (as far as we know) that the effect of a TRP enriched diet on fish hierarchical groups has been studied. Our contradictory results show how complicated the social interactions and mechanisms in fish hierarchies are, since previous studies have only looked at fish interacting in pair-wise contests. We propose that under true hierarchical conditions, other factors are involved in the overall control of aggression; therefore, attempting to control agonistic behavior in groups of fish with TRP may not be sufficient. This could be an important finding that needs to be studied further, especially in the context of fish welfare in intensive aquaculture

In Vitro Culture of the Anxiolytic Plant, Souroubea Sympetala

A protocol for the in vitro culture of the anxiolytic medicinal plant Souroubea sympetala (Marcgraviaceae) was developed, representing one of the first in vitro cultures for the family. This species was previously very difficult to cultivate from seed or cuttings. Methods included (1) the improvement of seed germination by axenic culture (2) development of regenerative cultures in vitro, then cultivation under greenhouse and finally field conditions and (3) creation of cell suspensions. Phytochemical analysis was undertaken by liquid chromatography coupled to mass spectrometry (HPLC-MS). The percentage of seed germination was improved from 2% to 59% in axenic culture and the full development of the seedling with its apical shoot and root took twenty-four days. The best seedling development was obtained in Gamborg B5 culture medium. Most friable callus formation, (66.7%) was obtained in the Murashige and Skoog medium supplemented with naphthalene acetic acid (1 mg · L–1) and kinetin (0.5 mg · L–1) from which viable cell cultures were developed. Analysis identified 4 main triterpenes with both in vitro plants and greenhouse grown plants derived from them. The triterpenes were betulinic acid, ursolic acid, alpha-amyrin and beta-amyrin. The betulinic acid found in greenhouse plants was comparable to wild plants. The cell suspension cultures had much lower levels of betulinic acid than plants and are not at present a viable source of this anxiolytic triterpene. In conclusion the method provides healthy plants for cultivation of this new medicinal plant and consequently harvesting of wild plants is not required.Se ha desarrollado un protocolo para el cultivo in vitro de la planta medicinal ansiolítica Souroubea sympetala (Marcgraviaceae), que representa uno de los primeros cultivos in vitro de la familia. Anteriormente, esta especie era muy difícil de cultivar a partir de semillas o esquejes. Los métodos aplicados incluyeron (1) la mejora de la germinación de semillas mediante cultivo axénico (2) el desarrollo de cultivos regenerativos in vitro, luego el cultivo en condiciones de invernadero y finalmente de campo y (3) la creación de suspensiones celulares. El análisis fitoquímico se llevó a cabo mediante cromatografía líquida acoplada a espectrometría de masas (HPLC-MS). El porcentaje de germinación de las semillas mejoró del 2% al 59% en cultivo axénico y el desarrollo completo de la plántula con su brote apical y su raíz tardó veinticuatro días. El mejor desarrollo de plántulas se obtuvo en el medio de cultivo Gamborg B5. La mayor formación de callo friable, (66,7%) se obtuvo en el medio Murashige y Skoog suplementado con ácido naftaleno acético (1 mg - L-1) y kinetina (0,5 mg - L-1) a partir de los cuales se desarrollaron cultivos celulares viables. Los análisis identificaron 4 triterpenos principales tanto en las plantas in vitro como en las cultivadas en invernadero. Los triterpenos eran ácido betulínico, ácido ursólico, alfa-amirina y beta-amirina. El ácido betulínico encontrado en las plantas de invernadero era comparable al de las plantas silvestres. Los cultivos de suspensión celular tenían niveles de ácido betulínico muy inferiores a los de las plantas y no constituyen en la actualidad una fuente viable de este triterpeno ansiolítico. En conclusión, el método proporciona plantas sanas para el cultivo de esta nueva planta medicinal y Por consiguiente, no es necesario recolectar plantas silvestres.University of OttawaInstituto de Investigación y Servicios Forestales (INISEFOR

A botanical extract of Souroubea sympetala and its active principle, betulinic acid, attenuate the cortisol response to a stressor in rainbow trout, Oncorhynchus mykiss

The capacity of the traditional anxiolytic plant, Souroubea sympetala, and its bioactive principle, betulinic acid (BA), to lower cortisol levels in rainbow trout, Oncorhynchus mykiss, was investigated. Effects at the level of the cortisol-producing interrenal cells were investigated using an in vitro head kidney preparation. Head kidney tissue incubated with S. sympetala extract or BA produced significantly less cortisol in response to adrenocorticotropic hormone (ACTH) than cells incubated with ACTH alone; a 50% reduction in the cortisol response was predicted to occur at 43 μg mL− 1 extract or 71 μg mL− 1 BA. To investigate whether effects on interrenal cell cortisol production translated into effects at the whole-animal level, the plasma cortisol response to an acute stressor, confinement in a net, was assessed. Fish fed a commercial trout pellet diet supplemented with 1 mg kg− 1 fish BA for six days exhibited a significantly lower plasma cortisol response to the acute stressor than fish fed the unsupplemented diet. Collectively, these results demonstrate that S. sympetala and BA act on the interrenal cells of the hypothalamic-pituitary-interrenal (HPI) axis in rainbow trout to significantly attenuate the cortisol response to an acute stressor. These findings suggest that BA is of potential use in mitigating effects of exposure to the stressors commonly encountered in an aquaculture setting. Statement of relevance • Souroubea sympetala extract and betulinic acid reduced ACTH-stimulated cortisol production by head kidney tissue in vitro, with little or no evidence of cytotoxicity.• The cortisol response to an acute stressor was attenuated in rainbow trout fed a commercial diet supplemented with betulinic acid.• The cortisol-reducing properties of the S. sympetala extract and betulinic acid may be useful in mitigating effects of stress in aquaculture, particularly since, as natural products, they may raise less public concern with respect to human consumption.Se investigó la capacidad de la planta ansiolítica tradicional, Souroubea sympetala, y su principio bioactivo, el ácido betulínico (BA), para reducir los niveles de cortisol en la trucha arco iris, Oncorhynchus mykiss. Los efectos a nivel de las células interrenales productoras de cortisol se investigaron utilizando una preparación de riñón de cabeza in vitro. El tejido del riñón de la cabeza incubado con extracto de S. sympetala o BA produjo significativamente menos cortisol en respuesta a la hormona adrenocorticotrópica (ACTH) que las células incubadas con ACTH sola; Se predijo que se produciría una reducción del 50% en la respuesta del cortisol con 43 μg mL - 1 extracto o 71 μg mL - 1 BA. Para investigar si los efectos sobre la producción de cortisol de las células interrenales se traducían en efectos a nivel de animal completo, se evaluó la respuesta del cortisol plasmático a un factor estresante agudo, el confinamiento en una red. Los peces alimentados con una dieta comercial de gránulos de trucha suplementada con 1 mg kg - 1 pescado BA durante seis días exhibieron una respuesta de cortisol plasmático significativamente menor al factor estresante agudo que los peces alimentados con la dieta sin suplementos. En conjunto, estos resultados demuestran que S. sympetala y BA actúan sobre las células interrenales del eje hipotalámico-pituitario-interrenal (HPI) en la trucha arco iris para atenuar significativamente la respuesta del cortisol a un factor de estrés agudo. Estos hallazgos sugieren que la BA tiene un uso potencial para mitigar los efectos de la exposición a los factores estresantes que se encuentran comúnmente en un entorno de acuicultura. Declaración de relevancia • El extracto de Souroubea sympetala y el ácido betulínico redujeron la producción de cortisol estimulada por ACTH en el tejido renal de la cabeza in vitro, con poca o ninguna evidencia de citotoxicidad. • La respuesta del cortisol a un factor estresante agudo se atenuó en la trucha arco iris alimentada con una dieta comercial complementada con ácido betulínico. • Las propiedades reductoras de cortisol del extracto de S. sympetala y el ácido betulínico pueden ser útiles para mitigar los efectos del estrés en la acuicultura, sobre todo porque, como productos naturales, pueden generar menos preocupación pública con respecto al consumo humano.Foi investigada a capacidade da planta ansiolítica tradicional, Souroubea sympetala, e seu princípio bioativo, ácido betulínico (BA), para baixar os níveis de cortisol na truta arco-íris, Oncorhynchus mykiss. Os efeitos ao nível das células interrenais produtoras de cortisol foram investigados usando uma preparação de rim de cabeça in vitro. O tecido renal da cabeça incubado com extrato de S. sympetala ou BA produziu significativamente menos cortisol em resposta ao hormônio adrenocorticotrófico (ACTH) do que as células incubadas com ACTH sozinho; uma redução de 50% na resposta do cortisol foi prevista para ocorrer no extrato de 43 μg mL - 1 ou 71 μg mL - 1 BA. Para investigar se os efeitos na produção de cortisol das células interrenais se traduziam em efeitos no nível do animal inteiro, a resposta do cortisol plasmático a um estressor agudo, confinamento em uma rede, foi avaliada. Os peixes alimentados com uma dieta comercial de pellet de truta suplementada com 1 mg kg-1 de BA para peixes por seis dias exibiram uma resposta de cortisol plasmático significativamente menor ao estressor agudo do que os peixes alimentados com a dieta não suplementada. Coletivamente, esses resultados demonstram que S. sympetala e BA atuam nas células interrenais do eixo hipotálamo-hipófise-interrenal (HPI) na truta arco-íris para atenuar significativamente a resposta do cortisol a um estressor agudo. Esses achados sugerem que o BA é de uso potencial na mitigação dos efeitos da exposição aos estressores comumente encontrados em um ambiente de aqüicultura. Declaração de relevância • O extrato de Souroubea sympetala e o ácido betulínico reduziram a produção de cortisol estimulado por ACTH pelo tecido renal da cabeça in vitro, com pouca ou nenhuma evidência de citotoxicidade. • A resposta do cortisol a um estressor agudo foi atenuada na truta arco-íris alimentada com uma dieta comercial suplementada com ácido betulínico. • As propriedades redutoras de cortisol do extrato de S. sympetala e ácido betulínico podem ser úteis na mitigação dos efeitos do estresse na aquicultura, particularmente porque, como produtos naturais, podem causar menos preocupação pública com relação ao consumo humanoUniversity of Ottawa, CanadáUniversidad Nacional, Costa RicaEscuela de Ciencias Ambientale

Indigenous ex situ conservation of Q'eqchi' Maya medicinal plant resources at the Itzamma Garden— Indian Creek, Belize, Central America

The Itzamma Garden and Medicinal Plant Project is a collaborative effort focused on identifying traditionally important plants used by the Q'eqchi' Maya of southern Belize. The Garden represents a cornerstone of the project where these plant resources (~130 in total, of which 102 have been identified to the species level) are cultivated ex situ as a means of providing primary healthcare by traditional healers. Here, we present a comprehensive agro-ecological survey of the Garden describing the layout and cultivation scenarios, development challenges encountered, and associated implementation strategies for improving site practices. A general recommendation for the adaptation of this strategy to other localities is the thorough depiction of ecological features based on interactions with local practitioners, for example, leading to the effective cultivation of plants, the enhancement of soil, and the surveillance of crop nutritional status and medicinal potency. We consider that the 'ethnobotanical garden approach' could provide a collaborative 'working model' for rural development, especially indigenous communities interfacing with their local agro-ecosystems

Anxiolytic activity and active principles of Piper amalago (Piperaceae), a medicinal plant used by the Q\u27eqchi\u27 Maya to treat susto, a culture-bound illness

Ethnopharmacological relevance: The medicinal plant, Piper amalago L. (Piperaceae), is used traditionally by Q\u27eqchi\u27 Maya healers for the treatment of susto a culture-bound syndrome. Previous research suggests that susto symptoms may be a manifestation of anxiety. The objectives were to characterize the effect of ethanolic extract of P. amalago in behavioral assays of anxiety at doses representative of traditional use and to isolate active principles. Materials and methods: Rats treated orally with low dose ethanolic extracts of P. amalago leaves (8-75mg/kg) were tested in several behavioral paradigms including the elevated plus maze (EPM), social interaction (SI), and conditioned emotional response (CER) tests, and compared to diazepam, a positive control. The active anxiolytic principle was isolated by bioassay guided isolation using an in vitro GABAA competitive binding assay. Results: Extracts had significant anxiolytic activity in all behavioral tests, with the strongest activity in the SI and the CER paradigms. In an in vitro GABAA competitive binding assay, a 66.5µg/mL concentration of P. amalago ethanol extract displaced 50% of the GABAA-BZD receptor ligand [(3)H]-Flunitrazepam. Bioassay-guided fractionation identified a furofuran lignan, a molecule with structural similarity to yangambin, with high affinity for the GABAA-BZD receptor as the principle bioactive. Conclusion: The results suggest that the ethnobotanical use of this plant may have a pharmacological basis in its anxiolytic activity, as demonstrated in animal behaviour tests

Antifungal Saponins from the Maya Medicinal Plant Cestrum schlechtendahlii G. Don (Solanaceae)

Herbario Juvenal Valerio RodríguezBioassay-guided fractionation of the crude extract (80% EtOH) of the leaves of Cestrum schlechtendahlii, a plant used by Q'eqchi' Maya healers for treatment of athlete's foot, resulted in the isolation and identification of two spirostanol saponins (1 and 2). Structure elucidation by MS, 1D-NMR, and 2D-NMR spectroscopic methods identified them to be the known saponin (25R)-1β,2α-dihydroxy-5α-spirostan-3-β-yl-O-α-l-rhamnopyranosyl-(1 → 2)-β-d-galactopyranoside (1) and new saponin (25R)-1β,2α-dihydroxy-5α-spirostan-3-β-yl-O-β-d-galactopyranoside (2). While 2 showed little or no antifungal activity at the highest concentration tested, 1 inhibited growth of Saccharomyces cerevisiae (minimum inhibitory concentration (MIC) of 15–25 μM), Candida albicans, Cryptococcus neoformans, and Fusarium graminearum (MIC of 132–198 μM).El fraccionamiento guiado por bioensayo del extracto crudo (80% EtOH) de las hojas de Cestrum schlechtendahlii, una planta utilizada por los curanderos mayas Q'eqchi 'para el tratamiento del pie de atleta, resultó en el aislamiento e identificación de dos saponinas de espirostanol (1 y 2 ). La elucidación de la estructura mediante métodos espectroscópicos de MS, 1D-NMR y 2D-NMR los identificó como la saponina conocida (25R) -1β, 2α-dihidroxi-5α-espirostan-3-β-il-O-α-l-ramnopiranosil- (1 → 2) -β-d-galactopiranósido (1) y nueva saponina (25R) -1β, 2α-dihidroxi-5α-espirostan-3-β-il-O-β-d-galactopiranósido (2). Mientras que 2 mostraron poca o ninguna actividad antifúngica a la concentración más alta probada, 1 inhibió el crecimiento de Saccharomyces cerevisiae (concentración mínima inhibitoria (MIC) de 15-25 μM), Candida albicans, Cryptococcus neoformans y Fusarium graminearum (MIC de 132-198 μM) ).O fracionamento guiado por bioensaio do extrato bruto (80% de EtOH) das folhas de Cestrum schlechtendahlii, uma planta usada pelos curandeiros Q'eqchi 'Maya para o tratamento do pé de atleta, resultou no isolamento e identificação de duas saponinas espirostanol (1 e 2 ) A elucidação da estrutura por métodos espectroscópicos de MS, 1D-NMR e 2D-NMR os identificou como sendo a saponina conhecida (25R) -1β, 2α-dihidroxi-5α-espirostan-3-β-il-O-α-l-ramnopiranosil- (1 → 2) -β-d-galactopiranosídeo (1) e nova saponina (25R) -1β, 2α-dihidroxi-5α-espirostan-3-β-il-O-β-d-galactopiranosídeo (2). Enquanto 2 mostraram pouca ou nenhuma atividade antifúngica na concentração mais alta testada, 1 inibiu o crescimento de Saccharomyces cerevisiae (concentração inibitória mínima (MIC) de 15–25 μM), Candida albicans, Cryptococcus neoformans e Fusarium graminearum (MIC de 132–198 μM )University of Ottawa, CanadaInstituto de Ecología, MéxicoUniversidad Nacional, Costa RicaBelize Indigenous Training Institute, BelizeAgriculture and Agri-Food Canada, CanadaCarleton University, CanadaEscuela de Ciencias Ambientale

Antifungal Saponins from the Maya Medicinal Plant Cestrum schlechtendahlii G. Don (Solanaceae)

Bioassay-guided fractionation of the crude extract (80% EtOH) of the leaves of Cestrum schlechtendahlii, a plant used by Q'eqchi' Maya healers for treatment of athlete's foot, resulted in the isolation and identification of two spirostanol saponins (1 and 2). Structure elucidation by MS, 1D-NMR, and 2D-NMR spectroscopic methods identified them to be the known saponin (25R)-1β,2α-dihydroxy-5α-spirostan-3-β-yl-O-α-l-rhamnopyranosyl-(1 → 2)-β-d-galactopyranoside (1) and new saponin (25R)-1β,2α-dihydroxy-5α-spirostan-3-β-yl-O-β-d-galactopyranoside (2). While 2 showed little or no antifungal activity at the highest concentration tested, 1 inhibited growth of Saccharomyces cerevisiae (minimum inhibitory concentration (MIC) of 15-25 μM), Candida albicans, Cryptococcus neoformans, and Fusarium graminearum (MIC of 132-198 μM)