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Context: Valeriana pavonii Poepp. & Endl. (Caprifoliaceae), is a plant used in traditional medicine as a tranquilizer in Colombia. Valerian extracts have been widely used since ancient times for their sedative and anxiolytic properties; however, the way its active metabolites, including iridoids, interact on their respective targets is not fully understood. Aims: To isolate and identificate active iridoid esters from V. pavonii. Perform in vitro inhibition assays and computational analyses to study their possible interaction on the benzodiazepine site of the GABAA receptor. Methods: Two compounds were obtained from dichloromethane and petroleum ether fractions of V. pavonii, respectively, by chromatographic techniques. The structural elucidation was performed by NMR and spectroscopic analyses. In vitro inhibition assays of the binding of 3H-flunitrazepam (3H-FNZ) for the benzodiazepine binding site of the GABAA receptor (BDZ-bs of the GABAA receptor) were carried out. Results: Two iridoid esters, hydrine-type valepotriates (compounds 1 and 2), were reported for the first time in V. pavonii. Both iridoids, 1 and 2, inhibited the binding of 3H-FNZ on the BDZ-bs of the GABAA receptor (40% at 300 µM). Docking studies and MMGBSA calculations revealed that these compounds exhibited molecular interactions with crucial residues of the benzodiazepine site, similar to those observed for drugs like flunitrazepam, diazepam, and flumazenil. Conclusions: These findings contribute to understanding the in vivo activity of extracts of Valeriana pavonni on the central nervous system, which showed promising effects, especially as anticonvulsants, sedative-hypnotics, and antidepressants, through the modulation of the GABAergic system by hydrine-type valepotriates and its derivatives.

The complex nature of neurodegenerative diseases (NDDs), such as Alzheimer's disease (AD) and Parkinson's disease (PD) calls for multidirectional treatment. Restoring neurotransmitter levels by combined inhibition of cholinesterases (ChEs) and monoamine oxidases (MAOs, MAO-A and MAO-B), in conjunction with strategies to counteract amyloid β (Aβ) aggregation, may constitute a therapeutically strong multi-target approach for the treatment of NDDs. Chalcones are a subgroup of flavonoids with a broad spectrum of biological activity. We report here the synthesis of 2′-hydroxychalcones as MAO-A and MAO-B inhibitors. Compounds 5c (IC50 = 0.031 ± 0.001 μM), 5a (IC50 = 0.084 ± 0.003 μM), 2c (IC50 = 0.095 ± 0.019 μM) and 2a (IC50 = 0.111 ± 0.006 μM) were the most potent, selective and reversible inhibitors of human (h)MAO-B isoform. hMAO-B inhibitors 1a, 2a and 5a also inhibited murine MAO-B in vivo in mouse brain homogenates. Molecular modelling rationalised the binding mode of 2′-hydroxychalcones in the active site of hMAO-B. Additionally, several derivatives inhibited murine acetylcholinesterase (mAChE) (IC50 values from 4.37 ± 0.83 μM to 15.17 ± 6.03 μM) and reduced the aggregation propensity of Aβ. Moreover, some derivatives bound to the benzodiazepine binding site (BDZ-bs) of the γ-aminobutyric acid A (GABAA) receptors (1a and 2a with Ki = 4.9 ± 1.1 μM and 5.0 ± 1.1 μM, respectively), and exerted sedative and/or anxiolytic like effects on mice. The biological results reported here on 2′-hydroxychalcones provide an extension to previous studies on chalcone scaffold and show them as a potential treatment strategy for NDDs and their associated comorbidities.

Folkloric or galenic preparations of valerian roots and rhizomes have been used as sedatives/anxiolytics and sleep inducers since ancient times. “Valerianas” are plants that naturally grow in our region. Although some of them are used in folk medicine, they lack scientific information. We performed a comparative study of the phytochemical composition and the potential in vivo effects of ethanolic extracts of argentine valerian species: Valeriana carnosa Sm., V. clarionifolia Phil. and V. macrorhiza Poepp. ex DC., from “Patagonia Argentina”; V. ferax (Griseb.) Hock and V. effusa Griseb., from the central part of our country, and V. officinalis (as the reference plant). All these plants were rich in phenolic compounds, evidenced the presence of ligands for the benzodiazepine binding site of the GABAA receptor and were able to induce sedation as assessed by loss-of-righting reflex assays (500 mg/kg, i.p.). Mice treated with V. macrorhiza, V. carnosa and V. ferax extracts showed reduced exploratory behaviors while V. clarionifolia produced anxiolytic-like activities (500 mg/kg, i.p.) in the Hole board test. Oral administrations (300 mg/kg and 600 mg/kg, p.o.) evidenced sedative effects for V. ferax and anxiolytic-like properties for V. macrorhiza, V. carnosa and V. clarionifolia extracts. Our native valerian species are active on the CNS, validating its folkloric use as anxiolytic/sedative and sleep enhancers., Facultad de Ciencias Exactas, Instituto de Estudios Inmunológicos y Fisiopatológicos

Anxiety disorders, depression and pain are highly prevalent pathologies. Their pharmacotherapy is associated with unwanted side effects; hence there is a clinical need to develop more effective drugs with fewer adverse reactions. Chalcones are one of the major classes of naturally occurring compounds. Chalcones and their derivatives have a huge importance in medicinal chemistry, displaying a wide range of pharmacological activities including anti-inflammatory, antimicrobial, antioxidant, cytotoxic and antitumor actions. The aim of this work was to evaluate chalcone effects on different targets involved in these pathologies. We have synthesized a series of simple chalcone derivatives taking common structural requirements described in literature related to their anxiolytic-like, antidepressant-like and/or antinociceptive properties into account. Furthermore, their potential in vitro effects towards different targets involved in these pathologies were evaluated. We have obtained twenty chalcones with moderate to high yields and assessed their ability to bind distinctive receptors, from rat brain homogenates, by displacement of labelled specific ligands: [ 3 H] FNZ (binding site of benzodiazepines/GABA A ), [ 3 H] 8-OH-DPAT (serotonin 5-HT 1A ) and [ 3 H] DAMGO (μ-opioid). Those compounds that showed the better in vitro activities were evaluated in mice using different behavioural tasks. In vivo results showed that 5′-methyl-2′-hydroxychalcone (9) exerted anxiolytic-like effects in mice in the plus maze test. While chalcone nuclei (1) revealed antidepressant-like activities in the tail suspension test. In addition, the novel 5′-methyl-2′-hydroxy-3′-nitrochalcone (12) exhibited antinociceptive activity in acute chemical and thermal nociception tests (writhing and hot plate tests). In conclusion, chalcones are thus promising compounds for the development of novel drugs with central nervous system (CNS) actions. Fil: Higgs, Josefina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Química y Físico-Química Biológicas "Prof. Alejandro C. Paladini". Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Instituto de Química y Físico-Química Biológicas; Argentina Fil: Wasowski, Cristina Lucia N.. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Química y Físico-Química Biológicas "Prof. Alejandro C. Paladini". Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Instituto de Química y Físico-Química Biológicas; Argentina Fil: Marcos, Alejandra Lucía. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Química y Físico-Química Biológicas "Prof. Alejandro C. Paladini". Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Instituto de Química y Físico-Química Biológicas; Argentina Fil: Jukic, Marko. University Of Ljubljana; Eslovenia Fil: Pavan, Carlos Humberto. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Química y Físico-Química Biológicas "Prof. Alejandro C. Paladini". Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Instituto de Química y Físico-Química Biológicas; Argentina Fil: Gobec, Stanislav. University Of Ljubljana; Eslovenia Fil: de Tezanos Pinto, Felicitas. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Química y Físico-Química Biológicas "Prof. Alejandro C. Paladini". Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Instituto de Química y Físico-Química Biológicas; Argentina Fil: Colettis, Natalia Claudia. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Química y Físico-Química Biológicas "Prof. Alejandro C. Paladini". Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Instituto de Química y Físico-Química Biológicas; Argentina Fil: Marder, Nora Mariel. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Química y Físico-Química Biológicas "Prof. Alejandro C. Paladini". Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Instituto de Química y Físico-Química Biológicas; Argentina

Epilepsy is recognized as one of the most common and serious neurological disorder affecting 1–2% of the world׳s population. The present study demonstrates that systemic administration of 3-butyl-5,5-dimethyl-1,2,3-oxathiazolidine-4-one-2,2-dioxide (DIOXIDE), a synthetic compound bioisoster of trimethadione and phenytoin (classical anticonvulsants), elicits a dose dependent anticonvulsant response in mice submitted to the subcutaneous pentylenetetrazole seizure test (scPTZ). Among various factors supposed to play role in epilepsy, oxidative stress and reactive species have strongly emerged. The protection exerted by DIOXIDE over the extent of brain oxidative damage produced by PTZ was determined, by measuring the levels of lipid peroxidation and reduced glutathione and the activity of Na+/K+-ATPase. Psychiatric disorders represent frequent comorbidities in persons with epilepsy. In this report, the potential anxiolytic and antidepressant activities of DIOXIDE were evaluated in several widely used models for assessing anxiolytic and antidepressant activities in rodents. Although DIOXIDE did not evidence anxiolytic activity at the doses tested, it revealed a significant antidepressant-like effect. Preliminary studies of its mechanism of action, by means of its capacity to act via the GABAA receptor (using the [3H]flunitrazepam binding assay in vitro and the picrotoxin test in vivo) and the Na+ channel (using the alkaloid veratrine, a voltage-Na+ channel agonist) demonstrated that the anticonvulsant effect is not likely related to the GABAergic pathway and the antidepressant-like effect could be due to its Na+ channel blocking properties. The results for DIOXIDE suggested it as a new anticonvulsant–antioxidant and antidepressant compound that deserves further development. Fil: Pastore, Valentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Química y Físico-Química Biológicas "Prof. Alejandro C. Paladini". Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Instituto de Química y Físico-Química Biológicas; Argentina Fil: Wasowski, Cristina Lucia N.. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Química y Físico-Química Biológicas "Prof. Alejandro C. Paladini". Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Instituto de Química y Físico-Química Biológicas; Argentina Fil: Higgs, Josefina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Química y Físico-Química Biológicas "Prof. Alejandro C. Paladini". Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Instituto de Química y Físico-Química Biológicas; Argentina Fil: Mangialavori, Irene Cecilia. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Química y Físico-Química Biológicas "Prof. Alejandro C. Paladini". Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Instituto de Química y Físico-Química Biológicas; Argentina Fil: Bruno Blanch, Luis Enrique. Universidad Nacional de La Plata. Facultad de Ciencias Exactas. Departamento de Ciencias Biológicas. Cátedra de Química Medicinal; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina Fil: Marder, Nora Mariel. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Química y Físico-Química Biológicas "Prof. Alejandro C. Paladini". Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Instituto de Química y Físico-Química Biológicas; Argentina

We report herein the design and optimization of a novel series of sulfamides and sulfamates derived from amino esters with anticonvulsant properties. The structures were designed based on the pharmacophoric pattern previously proposed, with the aim of improving the anticonvulsant action. The compounds were obtained by a new synthetic procedure with microwave assisted heating and the use of adsorbents in the isolation process. All the derivatives showed protection against the maximal electroshock seizure test (MES test) in mice at the lowest dose tested (30 mg/kg) but they did not show significant protection against the chemical induced convulsion by pentylenetetrazole. These results verify the ability of the computational model for designing new anticonvulsants structures with anti-MES activity. Additionally, we evaluated the capacity of the synthesized structures to bind to the benzodiazepine binding site (BDZ-bs) of the γ-aminobutiric acid receptor (GABAA receptor). Some of them showed medium to low affinity for the BDZ-bs. Fil: Villalba, Maria Luisa. Universidad Nacional de La Plata. Facultad de Ciencias Exactas. Departamento de Ciencias Biológicas. Cátedra de Química Medicinal; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina Fil: Enrique, Andrea Verónica. Universidad Nacional de La Plata. Facultad de Ciencias Exactas. Departamento de Ciencias Biológicas. Cátedra de Química Medicinal; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina Fil: Higgs, Josefina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Química y Físico-Química Biológicas "Prof. Alejandro C. Paladini". Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Instituto de Química y Físico-Química Biológicas; Argentina Fil: Castaño, Rocío A.. Universidad Nacional de La Plata. Facultad de Ciencias Exactas. Departamento de Ciencias Biológicas. Cátedra de Química Medicinal; Argentina Fil: Goicoechea, Sofia. Universidad Nacional de La Plata. Facultad de Ciencias Exactas. Departamento de Ciencias Biológicas. Cátedra de Química Medicinal; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina Fil: Taborda, Facundo. Universidad Nacional de La Plata. Facultad de Ciencias Exactas. Departamento de Ciencias Biológicas. Cátedra de Química Medicinal; Argentina Fil: Gavernet, Luciana. Universidad Nacional de La Plata. Facultad de Ciencias Exactas. Departamento de Ciencias Biológicas. Cátedra de Química Medicinal; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina Fil: Lick, Ileana Daniela. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Centro de Química Inorgánica "Dr. Pedro J. Aymonino". Universidad Nacional de La Plata. Facultad de Ciencias Exactas. Centro de Química Inorgánica "Dr. Pedro J. Aymonino"; Argentina Fil: Marder, Nora Mariel. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Química y Físico-Química Biológicas "Prof. Alejandro C. Paladini". Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Instituto de Química y Físico-Química Biológicas; Argentina Fil: Bruno Blanch, Luis Enrique. Universidad Nacional de La Plata. Facultad de Ciencias Exactas. Departamento de Ciencias Biológicas. Cátedra de Química Medicinal; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina

The cytotoxic activity of 2'-nitroflavone was evaluated in different haematological cancer cell lines and its mechanism of action was further studied in HL-60 cells. 2'-Nitroflavone arrested the cell cycle at the G(2)/M phase and induced an apoptotic response characterized by an increase in the sub-G1 fraction of cells, a typical DNA ladder fragmentation, chromatin condensation and the detection of cells stained with Annexin V. Apoptosis was dependent on the activation of at least caspase-8, caspase-9 and caspase-3. The involvement of the death receptor pathway was indicated by the upregulation of both the tumour necrosis factor-related apoptosis-inducing ligand (TRAIL) and its death receptor (DR5). We also showed that 2'-nitroflavone increased the expression levels of Bax and induced the release of cytochrome C to cytosol, suggesting the participation of the mitochondria-dependent pathway. When mitogen-activated protein kinases pathways were studied, it was found that p38 and c-Jun NH(2)-terminal kinase (JNK) pathways were activated by 2'-nitroflavone in HL-60 cells, whereas the phosphorylation levels of extracellular signal-regulated kinases (ERK) 1/2 decreased significantly. In addition, whereas both pharmacological inhibition of JNK and downregulation of JNK expression by RNA interference reduced the nitroflavone growth-inhibitory activity and the apoptotic effect, contrasting results were obtained when the ERK1/2 pathway was inhibited, and no effect was observed in the presence of a specific inhibitor of p38 mitogen-activated protein kinase. These findings show for the first time the antitumour action of 2'-nitroflavone in haematological cancer cell lines and suggest that both JNK and ERK1/2 cascades are involved in the apoptotic response induced by 2'-nitroflavone in HL-60 cells.

A set of sulfamides designed, synthesized and evaluated against maximal electroshock seizure (MES) and pentilenetetrazol (PTZ) tests with promising results, were tested for their affinity for the benzodiazepine binding site of the GABAA receptor. The most active compounds, N,N′-dicyclohexylsulfamide (7) and N,N′-diphenethylsulfamide (10), competitively inhibited the binding of [3H]-flunitrazepam to the benzodiazepine binding site with Ki ± SEM values of 27.7 ± 4.5 μM (n = 3) and 6.0 ± 1.2 μM (n = 3), respectively. The behavioral actions of these sulfamides, i.p. administered in mice, were examined in the plus-maze, hole-board and locomotor activity assays. Compound 7 exhibited anxiolytic-like effects in mice evidenced by a significant increase of the parameters measured in the hole-board test (at 1 and 3 mg/kg) and the plus-maze assay (at 1 and 3 mg/kg). Compound 10 evidenced anxiolytic activity in the plus-maze and the hole-board tests at 1 mg/kg. Locomotor activity of mice was not modified by compound 7 or 10 at the doses tested. Flumazenil, a non selective benzodiazepine binding site antagonist, was able to completely reverse the anxiolytic-like effects of these sulfamides, proving that the GABAA receptor is implicated in this action. Anxiety represents a major problem for people with epilepsy. The use of anxiolytic and anticonvulsant sulfamides would be beneficial to individuals who suffer from both disorders. Fil: Wasowski, Cristina Lucia N.. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Química y Físico-Química Biológicas "Prof. Alejandro C. Paladini". Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Instituto de Química y Físico-Química Biológicas; Argentina Fil: Gavernet, Luciana. Universidad Nacional de la Plata. Facultad de Ciencias Exactas. Departamento de Ciencias Biológicas. Cátedra de Química Medicinal; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina Fil: Barrios, Ivana Analia. Universidad Nacional de la Plata. Facultad de Ciencias Exactas. Departamento de Ciencias Biológicas. Cátedra de Química Medicinal; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina Fil: Villalba, Maria Luisa. Universidad Nacional de la Plata. Facultad de Ciencias Exactas. Departamento de Ciencias Biológicas. Cátedra de Química Medicinal; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina Fil: Pastore, Valentina. Universidad Nacional de La Plata. Facultad de Ciencias Exactas. Departamento de Ciencias Biológicas. Cátedra de Química Medicinal; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina Fil: Samaja, Gisela Anabel. Universidad Nacional de la Plata. Facultad de Ciencias Exactas. Departamento de Ciencias Biológicas. Cátedra de Química Medicinal; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina Fil: Enrique, Andrea Verónica. Universidad Nacional de la Plata. Facultad de Ciencias Exactas. Departamento de Ciencias Biológicas. Cátedra de Química Medicinal; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina Fil: Bruno Blanch, Luis Enrique. Universidad Nacional de la Plata. Facultad de Ciencias Exactas. Departamento de Ciencias Biológicas. Cátedra de Química Medicinal; Argentina Fil: Marder, Nora Mariel. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Química y Físico-Química Biológicas "Prof. Alejandro C. Paladini". Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Instituto de Química y Físico-Química Biológicas; Argentina

Fil: Wasowski, Cristina Lucia N.. Consejo Nacional de Investigaciones Cientificas y Tecnicas. Oficina de Coordinacion Administrativa Houssay. Instituto de Quimica y Fisico-Quimica Biologicas "Prof. Alejandro C. Paladini". Universidad de Buenos Aires. Facultad de Farmacia y Bioquimica. Instituto de Quimica y Fisico-Quimica Biologicas; Argentina

Using the guide of a competitive assay for the benzodiazepine binding site in the γ-aminobutyric acid type A receptor (GABAA), two active diterpenes were isolated from the aerial parts of Aloysia virgata (Ruíz & Pavón) A.L. Jussieu var. platyphylla (Briquet) Moldenke. These compounds, identified as (16R)-16,17,18-trihydroxyphyllocladan-3-one (1) and (16R)-16,17-dihydroxyphyllocladan-3-one (2) on the basis of spectral data, competitively inhibited the binding of [3H]-FNZ to the benzodiazepine binding site with Ki ± S.E.M. values of 56 ± 19 μM and 111 ± 13 μM, respectively. The behavioral actions of these diterpenes, intraperitoneally (i.p.) administered in mice, were examined in the plus-maze, holeboard, locomotor activity and light/dark tests. Compound 1 exhibited anxiolytic-like effects in mice evidenced by a significant increase of the parameters measured in the holeboard test (the number of head dips at 0.3 mg/kg and 3 mg/kg, the rears at 1 mg/kg and the time spent head-dipping at 3 mg/kg), in the plus-maze assay (the percentage of open arm entries at 1 mg/kg) and in the light/dark test (the time in light and the number of transitions at 1 mg/kg). Compound 2 augmented the number of rearings in the holeboard apparatus (at 0.3 mg/kg and 1 mg/kg) and the locomotor activity (at 1 mg/kg). These results reveal the presence of neuroactive compounds in Aloysia virgata. Fil: Wasowski, Cristina Lucia N.. Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Departamento de Química Analítica y Fisicoquímica; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina Fil: Marder, Nora Mariel. Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Departamento de Química Analítica y Fisicoquímica; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina

Introducción. El fraccionamiento fitoquímico de Valeriana pavonii, especie vegetal nativa utilizada tradicionalmente en Colombia con fines tranquilizantes, condujo al aislamiento e identificación de la isovaleramida, uno de los principios responsables de su actividad sobre el sistema nervioso central como anticonvulsivo. Objetivo. Reportar la identificación de la isovaleramida, metabolito de V. pavonii activo sobre el sistema nervioso central. Materiales y métodos. La purificación de la isovaleramida se realizó mediante técnicas cromatográficas. Su estrucutra se determinó por experimentos de resonancia magnética y espectrometría de masas. Se emplearon las pruebas de convulsión máxima inducida eléctricamente en ratones como ensayo farmacológico in vivo y el ensayo in vitro de unión al sitio de las benzodiacepinas sobre el receptor GABA-A. Resultados. En el modelo de convulsión máxima inducida eléctricamente en ratones, la isovaleramida, aislada de la fracción más activa de V. pavonii, confirió un índice de protección de 90% en una dosis de 100 mg/kg, por vía oral, comparable al agente de referencia utilizado: fenitoína sódica (20 mg/kg, por vía oral, 100%) y superior al control (vehículo, 20%). En el ensayo in vitro, la isovaleramida presentó un 42% de inhibición del sitio de unión de flunitracepam con tritio. Conclusión. La isovaleramida es uno de los principios activos anticonvulsivos de V. pavonii, por primera vez reportado en esta especie. Estos resultados dan soporte al uso tradicional de V. pavonii y a su interés como fuente de principios útiles en terapéutica. Introduction. Fractioning of an extract of Valeriana pavonii, a native species used in Colombian folk medicine as tranquilizer, led to the isolation and identification of isovaleramide, one of the active constituents responsible for its central nervous system activity as anticonvulsant. Objective. Description of the isolation and identification of isovaleramide, an active principle on central nervous system from Valeriana pavonii. Materials and methods. The purification of isovaleramide was carried out by chromatographic techniques. Its structural elucidation was determined by nuclear magnetic resonance and mass spectrometry. Maximal electroshock seizure was used as in vivo pharmacological test, additionally in vitro GABA-A/BDZ-binding site studies were performed. Results. Isovaleramide was isolated from the most active fraction of Valeriana pavonii. This compound, at 100 mg/Kg, p.o, evidenced a 90% index protection against the maximal electroshock seizure in mice (MES), comparable to the reference agent: sodium phenytoin (20 mg/kg, p.o, 100%). In the in vitro assay, isovaleramide (300 µM) exhibited a 42% of inhibition of the binding of 3H-FNZ to its sites. Conclusion. Isovaleramide is one of the active anticonvulsant constituents of Valeriana pavonii, for the first time reported in this species. These results support the traditional use of Valeriana pavonii and its interest as a therapeutic source. Fil: Giraldo, Sara Emilia. Universidad Nacional de Colombia; Colombia Fil: Rincón, Javier. Universidad Nacional de Colombia; Colombia Fil: Puebla, Pilar. Universidad de Salamanca; España Fil: Marder, Nora Mariel. Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina Fil: Wasowski, Cristina Lucia N.. Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina Fil: Vergel, Nadezdha. Universidad Nacional de Colombia; Colombia Fil: Guerrero, Mario Francisco. Universidad Nacional de Colombia; Colombia

Pharmacological assay guided purification of an ethanol extract of Tilia petiolaris DC. inflorescences resulted in the isolation and identification of isoquercitrin (ISO), quercetin 3-O-glucoside-7-O-rhamnoside (QUE) and kaempferol 3-O-glucoside-7-O-rhamnoside (KAE). The behavioral actions of these glycosylated flavonoids were examined in the hole board, locomotor activity and thiopental-induced loss of righting reflex tests in mice. QUE (10 and 30 mg/kg) and KAE (30 mg/kg), intraperitoneally (i.p.) administered to mice, reduced all the parameters measured in the hole board test, but ISO (30 mg/kg) only reduced the number of rearings. Meanwhile QUE at 30 mg/kg i.p. also decreased the ambulatory locomotor activity and increased the sodium thiopental-induced time of loss of the righting reflex suggesting a clear depressant action. The above results demonstrate the occurrence of neuroactive flavonoid glycosides in Tilia.

6,3'-Dinitroflavone (6,3'-DNF) is a synthetic flavone derivative that exerts anxiolytic effects in the elevated plus maze. Based on the finding that this effect is blocked by Ro15-1788 (ethyl-8-fluoro-5,6-dihydro-5-methyl-6-oxo-4H-imidazo[1,5-a][1,4]benzodiazepine-3-carboxylate) which is a specific antagonist at the benzodiazepine binding site of GABA(A) receptors we investigated the interaction of 6,3'-DNF with several recombinant GABA(A) receptor subtypes. Inhibition of [(3)H]flunitrazepam binding to recombinant GABA(A) receptors in transiently transfected HEK293 cells indicated that 6,3'-DNF exhibited the highest affinity for GABA(A) receptors composed of alpha1beta2gamma2 subunits and a 2-20 fold lower affinity for homologous receptors containing alpha2, alpha3, or alpha5 subunits. Two-electrode voltage-clamp experiments in Xenopus oocytes indicated that 6,3'-DNF does not induce chloride flux in the absence of GABA, but exerts low efficacy inverse agonistic modulatory effects on GABA-elicited currents in the GABA(A) receptor subtypes alpha1beta2gamma2 and alpha5beta2gamma2. In the subtypes alpha2beta2gamma2, alpha3beta2gamma2, alpha4beta2gamma2, alpha6beta2gamma2 or alpha4beta2delta and alpha4beta3delta, 6,3'-DNF exerts either none or very low efficacy positive modulatory effects. In contrast, 100 nM Ro15-1788 exhibited weak to moderate partial agonistic effects on each receptor investigated. These data indicate that Ro15-1788 only can antagonize the weak inverse agonist effects of 6,3'-DNF on alpha1beta2gamma2 and alpha5beta2gamma2 receptors, but will enhance the weak agonistic effects on the other receptor subtypes investigated. The possible mechanism of the Ro15-1788 sensitive anxiolytic effect of 6,3'-DNF is discussed.

Polygala sabulosa, a folk medicine, presents dihydrostyryl-2-pyrones (DST) and styryl-2-pyrones (STY), compounds structurally similar to kavalactones. Our previous study showed that the ethyl acetate fraction (EA) and these constituents present anxiolytic-like, hypno-sedative, and anticonvulsant effects in mice.This study investigated the role of benzodiazepine binding site (BDZ-bs) in the central effects of either EA or three DST (1, 2, and 3) and three STY (4, 5, and 7), using in vivo and in vitro assays.In the elevated plus-maze (EPM), flumazenil (FMZ), a BDZ antagonist, partially blocked the anxiolytic-like effect of DST-3 or STY-4 and STY-7, but not DST-1. Using electroencephalogram (EEG), EA protected against pentylenetetrazole (PTZ)-induced convulsion in rats, an effect partially blocked by FMZ, suggesting the participation of the BDZ-bs in this action. EA also protected against the maximal electroshock (MES)-induced convulsions in mice, a profile distinct from diazepam (DZP). DST and STY compounds inhibited the [(3)H]-flunitrazepam ([(3)H]-FNZ) binding to BDZ-bs in rat cortical synaptosomes with K (i) higher than 100 microM (DST-1), 41.7 microM (DST-2), 35.8 microM (DST-3), 90.3 microM (STY-4), 31.0 microM (STY-5) and 70.0 microM (STY-7). In the saturation assay, DST-3 and STY-7 competitively inhibited the binding of [(3)H]-FNZ to BDZ-bs with a significant decrease in apparent affinity (K (d)) and no change in maximal binding (B (max)).The present data support a partial BDZ-bs mediation of the anxiolytic-like and anticonvulsant effects of EA of P. sabulosa and its main isolated constituents, DST and STY.

The pharmacological effects on the central nervous system (CNS) of a range of available flavonoid glycosides were explored and compared to those of the glycosides 2S-hesperidin and linarin, recently isolated from valeriana. The glycosides 2S-neohesperidin, 2S-naringin, diosmin, gossipyn and rutin exerted a depressant action on the CNS of mice following i.p. injection, similar to that found with 2S-hesperidin and linarin. We demonstrate in this work that these behavioural actions, as measured in the hole board, thiopental induced sleeping time and locomotor activity tests, are unlikely to involve a direct action on gamma-aminobutyric acid type A (GABA(A)) receptors. The corresponding aglycones were inactive, pointing to the importance of the sugar moieties in the glycosides in their CNS depressant action following systemic administration. The pharmacological properties of the flavonoid glycosides studied here, in addition to our previous results with hesperidin and linarin, opens a promising new avenue of research in the field.

The aim of the present work is to demonstrate the putative sedative and anxiolytic-like effects of a hydro-ethanolic extract obtained from the aerial parts of Aloysia polystachya (Verbenaceae) in male mice using several behavioural assays. Groups of male mice orally treated with doses of 1.0, 10.0 and 100.0 mg/kg of the extract did not show any significant alteration of their locomotor activity, body temperature or motor coordination. The same treatment increased the duration of the sleeping time induced by 30.0 mg/kg i.p. of sodium pentobarbital. However, the sleeping time induced by ethyl ether was not modified by the oral administration of the extract, not confirming the putative sedative effect of the plant. The ethanolic extract also significantly increased the percentage of both entries (1.0 and 100.0 mg/kg) and the time spent (10.0 and 100.0 mg/kg) into the open arms of the elevated plus maze (EPM). Nevertheless, the binding of 3H-flunitrazepam (3H-FNZ) to the benzodiazepine binding site (BDZ-bs), in washed crude synaptosomal membranes from rat cerebral cortex, was not affected by the semi-purified components from Aloysia polystachya. These results indicate an anxiolytic-like profile of action for the extract of Aloysia polystachya without sedative side effect, being this activity probably mediated by other mechanism than BDZ-bs modulation at the GABAA receptors.

We have recently reported the presence of the anxiolytic flavone 6-methylapigenin (MA) and of the sedative and sleep-enhancing flavanone glycoside 2S (−) hesperidin (HN) in Valeriana officinalis and Valeriana wallichii . MA, in turn, was able to potentiate the sleep-inducing properties of HN. The present paper reports the identification in V. officinalis of the flavone glycoside linarin (LN) and the discovery that it has, like HN, sedative and sleep-enhancing properties that are potentiated by simultaneous administration of valerenic acid (VA). These effects should be taken into account when considering the pharmacological actions of valeriana extracts.

The antiproliferative activity of several natural and synthetic flavonoids and some related compounds was evaluated in vitro against a cell line derived from a human cervical carcinoma (WISH cells). According to their activities, the most potent derivatives were 2'-nitroflavone (14), 2',6-dinitroflavone (15) and the n-buthyl ester of caffeic acid (29). When these compounds were tested in the presence of recombinant human interferon-alpha2b (rhIFN-alpha2b), a cytokine exhibiting an antimitogenic action on WISH cells, an additive effect on cell growth inhibition was observed. Time course studies of the antiproliferative action exerted by the active derivatives or the rhIFN-alpha2b suggested that these compounds induced cell death.

Valerian is an ancient tranquillizing drug obtained from the underground organs of several Valeriana species. Its active principles were assumed to be terpenoids in the form of valepotriates and/or as components of the essential oil. However, unknown active compounds were not discarded and synergic effects were suspected. We have recently isolated 6-methylapigenin (MA) from Valeriana wallichii and proved that it is a benzodiazepine binding site (BDZ-bs) ligand [Planta Med. 68 (2002) 934]. The present paper is the first report of the presence of 2S(−)-hesperidin in valeriana and describes that it has sedative and sleep-enhancing properties. MA, in turn, was found to have anxiolytic properties and was able to potentiate the sleep-enhancing properties of hesperidin (HN). MA and HN are new members of the growing family of natural flavonoids with activity on the CNS, and their properties suggest that they are promising drug leads in the field.

6-Chloro-3'-nitroflavone integrates a list of nearly 70 flavone derivatives synthesized in our laboratories. The effects of 6-chloro-3'-nitroflavone on the benzodiazepine binding sites (BDZ-BSs) of the GABA(A) receptor were examined in vitro and in vivo. 6-Chloro-3'-nitroflavone inhibited the [3H]flunitrazepam ([3H]FNZ) binding to rat cerebral cortex membranes with a Ki of 6.68 nM and the addition of GABA to extensively washed membranes did not modify its affinity for the BDZ-BSs (GABA-shift = 1.16+/-0.12). The binding assays performed in rat striatal and cerebellar brain membranes showed that this compound has similar affinity to different populations of BDZ-BSs. Electrophysiological experiments revealed that 6-chloro-3'-nitroflavone did not affect GABA(A)-receptors (GABA(A)-Rs) responses recorded in Xenopus oocytes expressing alpha1beta2gamma2s subunits, but blocked the potentiation exerted by diazepam (DZ) on GABA-activated chloride currents. In vivo experiments showed that 6-chloro-3'-nitroflavone did not possess anxiolytic, anticonvulsant, sedative, myorelaxant actions in mice or amnestic effects in rats; however, 6-chloro-3'-nitroflavone antagonized diazepam-induced antianxiety action, anticonvulsion, short-term, and long-term amnesia and motor incoordination. These biochemical, electrophysiological, and pharmacological results suggest that 6-chloro-3'-nitroflavone behaves as an antagonist of the BDZ-BSs.

6-Methyl-3'-bromoflavone inhibited [(3)H]flunitrazepam binding to the benzodiazepine binding site of the GABA(A) receptor (BDZ-bs) with Ki values between 10 and 50 nM in different brain regions. The GABA ratio of 1.03 for [(3)H]flunitrazepam binding to cerebral cortex, 0.76 for cerebellum, 0.7 for hippocampus, 0.7 for striatum, and 0.8 for spinal cord indicated an antagonistic or weak inverse agonistic profile of 6-methyl-3'-bromoflavone on BDZ-bs. Unlike classical benzodiazepines, it had no anticonvulsant, anxiolytic, myorelaxant, sedative, amnestic or motor incoordination effects. However, it antagonized the muscle relaxant, the sedative effect, and the changes in locomotor activity induced by diazepam. Taken together, these findings suggest that 6-methyl-3'-bromoflavone has an antagonistic profile on the BDZ-bs.

Flavonoids isolated from plants used as tranquilizers in folkloric medicine have a selective affinity, for central benzodiazepine receptors (BDZ-Rs) and some of them possess a pharmacological profile compatible with a partial agonist action. Synthetic derivatives of the common flavone nucleus, give rise to high affinity ligands when electronegative groups are introduced in carbons 6 and/or 3'. Representative compounds such as 6,3'-dinitroflavone and, 6-bromo-3'-nitroflavone exhibit a high affinity for the BDZ-Rs (Ki = 1.5 to 30 nM) and have anxiolytic effects not associated with myorelaxant, sedative or amnesic actions. These compounds or similar ones, could lead to improved therapeutic drugs in the treatment of anxiety.

Salvia guaranitica St. Hil. is a traditional medicinal plant used in Latin America as sedative. We have recently demonstrated the presence of cirsiliol in its extracts and found that this flavonoid is a competitive low affinity benzodiazepine receptor ligand (Marder et al., 1996). This report describes the pharmacological properties of Salvia guaranitica extracts and of its active principle, cirsiliol. A partially purified fraction of this plant, administered intraperitoneally in mice (in a dose equivalent to 3 g of the fresh plant), exhibited sedative and hypnotic effects as measured in the hole board and in the pentobarbital-induced sleep tests, respectively. On the other hand, this fraction had no anxiolytic or myorelaxant effects. In the pentobarbital-induced sleep test, cirsiliol (2-10mg/kg, i. p.) exhibited a dose-dependent hypnotic action. In contrast, it did not produce myorelaxant (up to 30mg/kg) or anticonvulsant (up to 10mg/kg) effects. Cirsiliol was found to be more potent in displacing (3)H Zolpidem binding (K(i) = 20 LiM) than (3)H flunitrazepam binding (K(i) = 200 μM) to benzodiazepine receptors from rat cerebral cortex. It is concluded that Salvia guaranitica extracts and its active principle cirsiliol, possess sedative and hypnotic properties; cirsiliol produces these effects probably acting on the so-called type I benzodiazepine receptor.

The synthesis and biochemical characterization of 6-bromo-3′-nitroflavone ( 1 ) is presented. Compound 1 has higher affinity for cerebellar and cerebral cortical than for striatal, hippocampal, or spinal cord benzodiazepine receptors (BDZ-Rs). In the cerebral cortex it recognizes two populations of binding sites (K i s 1.2 nM and 15.5 nM, respectively), and at doses of 0.01 to 0.3 mg/kg, ip produces anxiolytic effects in mice.

Benzodiazepines (BDZs) are the most widely prescribed class of psychoactive drugs in current therapeutic use, despite the important unwanted side-effects that they produce such as sedation, myorelaxation, ataxia, amnesia, ethanol and barbiturate potentiation and tolerance. Searching for safer BDZ-receptor (BDZ-R) ligands we have recently demonstrated the existence of a new family of ligands which have a flavonoid structure. First isolated from plants used as tranquilizers in folkloric medicine, some natural flavonoids have shown to possess a selective and relatively mild affinity for BDZ-Rs and a pharmacological profile compatible with a partial agonistic action. In a logical extension of this discovery various synthetic derivatives of those compounds, such as 6,3'-dinitroflavone were found to have a very potent anxiolytic effect not associated with myorelaxant, amnestic or sedative actions. This dinitro compound, in particular, exhibits a high affinity for the BDZ-Rs (Ki = 12-30 nM). Due to their selective pharmacological profile and low intrinsic efficacy at the BDZ-Rs, flavonoid derivatives, such as those described, could represent an improved therapeutic tool in the treatment of anxiety. In addition, several flavone derivatives may provide important leads for the development of potent and selective BDZ-Rs ligands.

6,3′-Dinitroflavone is a synthetic flavone derivative with high affinity for central benzodiazepine receptors that has anxiolytic effects. Here, we describe its biochemical and pharmacological characterization. 6,3′-Dinitroflavone inhibited differentially [3H]flunitrazepam binding to central benzodiazepine receptors in several brain regions, showing a lower Ki value in the cerebellum (central benzodiazepine receptor type I-enriched area), and a higher Ki value in the spinal cord and in the dentate gyrus (central benzodiazepine receptor type II-enriched area). When i.p. injected in mice, 6,3′-dinitroflavone had a potent anxiolytic effect in the elevated plus maze test. This effect was blocked by the specific central benzodiazepine receptor antagonist, Ro 15-1788. 6,3′-Dinitroflavone did not exhibit anticonvulsant or myorelaxant effects in mice or amnestic effects in rats. Moreover, it abolished the myorelaxant effect of diazepam. On the other hand, 6,3′-dinitroflavone possessed a mild sedative action only at doses 100–300-fold greater than the anxiolytic one. Based on these findings, we suggest that 6,3′-dinitroflavone has a benzodiazepine partial agonist profile, with low selectivity for central benzodiazepine receptor types I and II.

The pharmacotherapy for the treatment of pain is an active area of investigation. There are effective drugs to treat this problem, but there is also a need to find alternative treatments free of undesirable side effects. In the present work the capacity of a series of flavonoids to bind to the m opioid receptor was evaluated. The most active compound, 3,3-dibromoflavanone (31), a synthetic flavonoid, presented a significant inhibition of the binding of the selective m opioid ligand [3 H]DAMGO, with a Ki of 0.846 0.263 mM. Flavanone 31 was further synthesized using a simple and cheap procedure with good yield. Its in vivo effects in mice, after acute treatments, were studied using antinociceptive and behavioral assays. It showed no sedative, anxiolytic, motor incoordination effects or inhibition of the gastrointestinal transit in mice at the doses tested. It evidenced antinociceptive activity on the acetic acid-induced nociception, hot plate and formalin tests (at 10 mg/kg and 30 mg/kg). The results showed that the 5-HT2 receptor and the adrenoceptors seem unlikely to be involved in its antinociceptive effects. Naltrexone, a nonselective opioid receptors antagonist, totally blocked compound 31 antinociceptive effects on the hot plate test, but naltrindole (d opioid antagonist) and nor-binaltorphimine (k opioid antagonist) did not. These findings demonstrated that 3,3-dibromoflavanone (31), at doses that did not interfere with the motor performance, exerted clear dose dependent antinociception when assessed in the chemical and thermal models of nociception in mice and it seems that its action is related to the activation of the m opioid receptor. Fil: Higgs, Josefina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Química y Fisicoquímica Biológicas; Argentina Fil: Wasowski, Cristina Lucia N.. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Química y Fisicoquímica Biológicas; Argentina Fil: Loscalzo, Leonardo Martin. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Química y Fisicoquímica Biológicas; Argentina Fil: Marder, Nora Mariel. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Química y Fisicoquímica Biológicas; Argentina

The synthesis and anticonvulsant activity of novel heterocycles N-derivative-1,2,3-oxathiazolidine-4-one-2,2-dioxides, bioisosteres of trimethadione (TMD, oxazolidine-2,4-dione) and phenytoin (PHE), are described. TMD is an anticonvulsant drug widely used against absences seizures in the early 80's and PHE is an antiepileptic drug with a wide spectrum activity. The intermediates of synthesis of N-derivative-1,2,3-oxathiazolidine-4-one-2,2-dioxides, α-hydroxyamides, were obtained using microwave assisted synthesis. Anticonvulsant screening was performed in mice after intraperitoneal administration in the maximal electroshock seizure test (MES) and subcutaneous pentylenetetrazole seizures test (scPTZ). These new compounds showed a wide spectrum activity and were no neurotoxic in the RotoRod test. α-Hydroxyamides and N-derivative-1,2,3-oxathiazolidine-4-one-2,2-dioxides were 3-4700 times more potent than valproic acid in the MES test. Quantification of anticonvulsant protection was calculated (ED(50)) for the most active candidates; α-hydroxyamides 3a-c and 3e, and N-derivative-oxathiazolidine-4-one-2,2-dioxides 5a-c with ED(50) values of 9.1, 53.9, 44.6, 25.2, 15.1, 91.1 and 0.06mg/kg, respectively, in the MES test.

This paper extended the evaluation of the depressant and antinociceptive activities of hesperidin in order to determine its effectiveness by the intraperitoneal and oral routes, its pharmacological interaction with diverse pathways of neurotransmission and the role of its aglycone, hesperetin. The capacity of hesperidin and hesperetin to bind to μ-opioid receptor and their actions on μ-opioid receptor co-expressed with GIRK1/GIRK2 channels (G protein-activated inwardly rectifying K+ channels) in Xenopus laevis oocytes were also determined. Hesperidin exhibited a depressant activity in the hole board and locomotor activity tests, antinociceptive activities in the abdominal writhing and hot plate tests and no motor incoordination in the inverted screen and rotarod assays, only by the intraperitoneal route. Hesperetin did not show any effects in vivo in mice in these models, but in vitro it displaced the [3H]DAMGO binding with low-affinity and inhibited inward currents through the expressed GIRK1/2 channels. Although hesperidin actions in vivo demonstrated to be mediated by an opioid mechanism of action, it failed to directly bind to and activate the μ-opioid receptor or produce any change on inward GIRK1/2 currents in vitro. However, it should be considered that hesperidin may be metabolized, possibly resulting in crucial changes in its biological activity.

The aim of this work was to evaluate if the intraperitoneal administration of the natural compound hesperidin, in a sedative dose, and neo-hesperidin, a hesperidin structural analog that exerts minor sedative effect, were able to induce changes in intracellular signaling cascades in different areas of the brain. The systemic administration of hesperidin produced a marked reduction in the phosphorylation state of extracellular signal-regulated kinases 1/2 (ERK 1/2), but not of Ca(+2)/calmodulin-dependent protein kinase II alpha subunit (alphaCaMKII), in the cerebral cortex, cerebellum and hippocampus. In contrast, neo-hesperidin did not markedly affect the activity of ERK 1/2 in both the cortex and the cerebellum. Taken together, these results demonstrated that intracellular signalling involving a selective decrease in ERK1/2 activation accompanied the depressant action of hesperidin. Even more, the low sedative action of neo-hesperidin correlates with a negligible decrease in phosphorylation state of ERK 1/2 (pERK 1/2), suggesting that low levels of pERK 1/2 in CNS could be a marker of sedative efficacy of flavonoids.

Previous reports from our laboratory described the sedative activity of hesperidin (hesperetin-7-rhamnoglucoside). This property is greatly increased when the glycoside is injected jointly with diazepam and this interaction has been shown to be synergistic. In the present work the generality of the synergistic phenomenon is proved, since potentiation also occurs with several other benzodiazepines, namely alprazolam, bromazepam, midazolam and flunitrazepam. In order to advance in the study of the mechanism of action of hesperidin, the possible participation of several brain receptors, which are implicated in the control of numerous behavioral and physiological functions, was explored by investigating the effects of a variety of their antagonists on hesperidin actions. The results showed that the 5-HT2 receptor and the alpha1-adrenoceptor seem unlikely to be involved in the behavioral effects of hesperidin. Naltrexone, a nonselective antagonist of opioid receptors, totally blocked hesperidin effects on locomotion, and partially antagonized hesperidin-induced decreased exploration in the hole board test. Nor-binaltorphimine, a selective kappa opioid receptor antagonist, was able to partially block hesperidin effects on locomotor activity. Furthermore, hesperidin-induced antinociception was partially blocked by naltrexone, and potentiated by co-administration with alprazolam. Hence, the participation of the opioid system in the sedative, antinociceptive and potentianting effects of hesperidin with benzodiazepines in mice is highly probable. Our results suggest a possible beneficial use of the association of hesperidin with benzodiazepines, not only to improve human sedative therapy, but also in the management of pain.

It has been recently reported the presence in Valeriana of the flavone 6-methylapigenin and the flavanone glycoside hesperidin. The apigenin derivative is a ligand for the benzodiazepine binding site in the gamma-aminobutyric acid receptor type A (GABA(A)) and has anxiolytic properties. Hesperidin has sedative and sleep-enhancing properties but is not a ligand for the benzodiazepine binding site. 6-Methylapigenin is able to potentiate the sleep-enhancing effect of hesperidin. In this work we demonstrate that this property is shared with various GABA(A) receptor ligands, among them the agonist diazepam, which was used to study the potentiation as measured in the hole board test. Isobolar analysis of the results showed the interaction being synergistic. We discarded pharmacokinetic effects or a direct action of hesperidin on the benzodiazepine binding site. A possible use of hesperidin properties to decrease the effective therapeutic doses of benzodiazepines is suggested.

Using the guidance by a competitive assay for the benzodiazepine binding site in the GABA(A) receptor, active compounds were isolated from the rhizomes and roots of Valeriana wallichii DC. The UV, NMR and mass spectral data permitted the identification of 6-methylapigenin. This flavonoid has a Ki = 495 nM for the BDZ-bs and a GABA ratio of 1.6-2.0, which suggests possible agonistic properties. The calculated percentage of 6-methylapigenin in the crude drug is in the range: 0.013% to 0.0013%.

6-Bromo-3'-nitroflavone is a synthetic flavone derivative that selectively recognizes benzodiazepine receptors and has potent anxiolytic-like effects. Here, we describe in detail its pharmacological characterization. When i.p. injected in mice, 6-bromo-3'-nitroflavone (0.01-0.3 mg/kg) had an anxiolytic-like effect in the elevated plus-maze test. This effect was blocked by the specific benzodiazepine receptor antagonist, flumazenil. In addition, it exhibited anxiolytic-like actions when given orally (1 mg/kg). 6-Bromo-3'-nitroflavone did not exhibit myorelaxant effects (up to 30 mg/kg, i.p.). Unlike diazepam, this flavonoid produced no anterograde amnesia in a one-trial inhibitory avoidance learning. On the other hand, 6-bromo-3'-nitroflavone possessed mild anticonvulsant activity (0.1 mg/kg, i.p.) and provoked sedative-depressant actions only at doses 100-1000 times higher than those producing anxiolytic-like effects. 6-Bromo-3'-nitroflavone (0.1-1 mM) produced a lower potentiation of gamma-amino-butyric acid (GABA)-stimulated 36Cl- influx (126-138%) in comparison to diazepam (0.1 mM: 166%) in cerebral cortical membrane vesicles. Taken together, these findings suggest that 6-bromo-3'-nitroflavone has anxiolytic-like action possibly behaving as a partial agonist of the benzodiazepine receptors.

Benzodiazepines are the most widely prescribed class of psychoactive drugs in current therapeutic use, despite the important unwanted side effects that they produce, such as sedation, myorelaxation, ataxia, amnesia, and ethanol and barbiturate potentiation and tolerance. They exert their therapeutic effects via binding to the benzodiazepine binding site of gamma-aminobutyric acid (GABA) type A receptors, and allosterically modulating the chloride flux through the ion channel complex. First isolated from plants used as tranquilizers in folkloric medicine, some natural flavonoids have been shown to possess selective affinity for the benzodiazepine binding site with a broad spectrum of central nervous system effects. Since the initial search for alternative benzodiazepine ligands amongst the flavonoids, a list of successful synthetic derivatives has been generated with enhanced activities. This review provides an update on research developments that have established the activity of natural and synthetic flavonoids on GABA type A receptors. Flavonoids are prominent drugs in the treatment of mental disorders, and can also be used as tools to study modulatory sites at GABA type A receptors and to develop GABA type A selective agents further. Fil: Wasowski, Cristina Lucia N.. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Química y Físico-Química Biológicas "Prof. Alejandro C. Paladini". Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Instituto de Química y Físico-Química Biológicas; Argentina Fil: Marder, Nora Mariel. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Química y Físico-Química Biológicas "Prof. Alejandro C. Paladini". Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Instituto de Química y Físico-Química Biológicas; Argentina