RESUMEN

The genetically transformed hairy root line LRT 7.31 obtained by infecting leaf explants of Lopezia racemosa Cav with the Agrobacterium rhizogenes strain ATCC15834/pTDT, was evaluated to identify the anti-inflammatory and cytotoxic compounds reported previously for the wild plant. After several subcultures of the LRT 7.31 line, the bio-guided fractionation of the dichloromethane-methanol (1:1) extract obtained from dry biomass afforded a fraction that showed important in vivo anti-inflammatory, and in vitro cytotoxic activities. Chemical separation of the active fraction allowed us to identify the triterpenes ursolic (1) and oleanolic (2) acids, and (23R)-2α,3ß,23,28-tetrahydroxy-14,15-dehydrocampesterol (3) as the anti-inflammatory principles of the active fraction. A new molecule 3 was characterized by spectroscopic analysis of its tetraacetate derivative 3a. This compound was not described in previous reports of callus cultures, in vitro germinated seedlings and wild plant extracts of whole L. racemosa plants. The anti-inflammatory and cytotoxic activities displayed by the fraction are associated to the presence of compounds 1-3. The present study reports the obtaining of the transformed hairy roots, the bioguided isolation of the new molecule 3, and its structure characterization.

Asunto(s)

Antiinflamatorios/farmacología , Proliferación Celular/efectos de los fármacos , Colesterol/análogos & derivados , Germinación/efectos de los fármacos , Onagraceae/química , Fitosteroles/farmacología , Agrobacterium/química , Agrobacterium/genética , Antiinflamatorios/química , Callo Óseo/efectos de los fármacos , Callo Óseo/crecimiento & desarrollo , Colesterol/química , Colesterol/farmacología , Ácido Oleanólico/química , Ácido Oleanólico/farmacología , Fitosteroles/química , Raíces de Plantas/química , Plantas Modificadas Genéticamente/efectos de los fármacos , Plantones/efectos de los fármacos

RESUMEN

CONTEXT: Agastache mexicana (Kunth) Lint & Epling (Lamiaceae) is a plant used in Mexican traditional medicine for the treatment of hypertension, anxiety and so on. OBJECTIVE: To determine the vasorelaxant effect and functional mode of action of dichloromethane-soluble extract from A. mexicana (DEAm) and isolate the constituents responsible for the pharmacological activity. MATERIALS AND METHODS: Extracts were prepared from the aerial parts of A. mexicana (225.6 g) by successive maceration with hexane, dichloromethane and methanol (three times for 72 h at room temperature), respectively. DEAm (0.01-1000 µg/mL), fractions (at 174.27 µg/mL), acacetin and ursolic acid (UA) (0.5-500 µM) were evaluated to determine their vasorelaxant effect on ex vivo rat aorta ring model. In vivo UA antihypertensive action was determined on spontaneously hypertensive rats. RESULTS AND DISCUSSION: DEAm induced a significant vasorelaxant effect in concentration-dependent and endothelium-independent manners (EC50 = 174.276 ± 5.98 µg/mL) by a calcium channel blockade and potassium channel opening. Bio-guided fractionation allowed to isolate acacetin (112 mg), UA (2.830 g), acacetin/oleanolic acid (OA) (M1) (155 mg) and acacetin/OA/UA (M2) (1.382 g) mixtures, which also showed significant vasodilation. UA significantly diminished diastolic (80 mmHg) and systolic blood pressure (120 mmHg), but heart rate was not modified. CONCLUSION: DEAm produced significant vasorelaxant action by myogenic control cation. The presence of acacetin, OA and UA into the extract was substantial for the relaxant activity of DEAm. In vivo antihypertensive action of UA corroborates the use of A. mexicana as an antihypertensive agent on Mexican folk medicine.

Asunto(s)

Agastache , Cloruro de Metileno/farmacología , Extractos Vegetales/farmacología , Vasodilatación/efectos de los fármacos , Vasodilatadores/farmacología , Animales , Aorta Torácica/efectos de los fármacos , Aorta Torácica/fisiología , Relación Dosis-Respuesta a Droga , Masculino , Técnicas de Cultivo de Órganos , Extractos Vegetales/aislamiento & purificación , Ratas , Ratas Endogámicas SHR , Ratas Wistar , Vasodilatación/fisiología , Vasodilatadores/aislamiento & purificación

RESUMEN

Lopezia racemosa Cav. is a plant used in Mexican traditional medicine to heal inflammatory diseases. From this plant we isolated the novel compound 6-O-palmitoyl- 3-O-ß-D-glucopyranosylcampesterol (1) and 6-O-palmitoyl-3-O-ß-D-glucopyranosyl-ß-sitosterol (2), previously reported to have cytotoxic activity on several cancer cell lines. We evaluated the anti-inflammatory activity of 1 in vivo by mouse ear edema induced with 12-O-tetradecanoylphorbol-13-acetate (TPA) and 57.14% inhibition was observed. The aim of our study was to obtain callus cultures derived from this plant species with the ability to produce the compounds of interest. Callus cultures were initiated on MS basal medium amended with variable amounts of naphthaleneacetic acid (NAA), or 2,4-dichlorophenoxyacetic acid (2,4-D), combined or not with 6-benzylaminopurine (BAP). Ten treatments with these growth regulators were carried out, using in vitro germinated seedlings as source of three different explants: hypocotyl, stem node, and leaf. Highest yield of 1 was observed on callus derived from leaf explants growing in medium containing 1.0 mg/L 2,4-D and 0.5 mg/L BAP. Selected callus lines produced less 1 than wild plants but the in vitro cultured seedlings showed higher production. So we conclude that it could be attractive to further investigate their metabolic potential.

Asunto(s)

Antiinflamatorios/farmacología , Colesterol/análogos & derivados , Inflamación/tratamiento farmacológico , Onagraceae/metabolismo , Fitosteroles/farmacología , Extractos Vegetales/farmacología , Animales , Antiinflamatorios/síntesis química , Células Cultivadas , Colesterol/síntesis química , Colesterol/química , Colesterol/farmacología , Oído/patología , Edema/tratamiento farmacológico , Germinación/fisiología , Masculino , Ratones , Fitoquímicos/farmacología , Fitosteroles/síntesis química , Fitosteroles/química , Semillas/fisiología , Acetato de Tetradecanoilforbol

RESUMEN

OBJECTIVE: To assess the relaxant effect of several organic extracts obtained from Agastache mexicana (A. mexicana), Cochlospermum vitifolium (C. vitifolium), Cordia morelosana (C. morelosana), Lepechinia caulescens (L. caulescens) and Talauma mexicana (T. mexicana) used in Mexican traditional medicine for the treatment of several diseases. METHODS: Extracts were obtained by maceration at room temperature using hexane, dichloromethane and methanol for each plant material. The organic extracts were evaluated ex vivo to determine their relaxant activity on the contractions induced by carbachol (cholinergic receptor agonist, 1 µ mol/L) in isolated rat tracheal rings. RESULTS: A total of 15 extracts were evaluated (three for each species). All test samples showed significant relaxant effect, in a concentration-dependent manner, on the contractions induced by 1 µ mol/L carbachol, with exception of extracts from C. morelosana. Active extracts were less potent than theophylline [phosphodiesterase inhibitor, EC50: (28.79±0.82) µg/mL] that was used as positive control. Concentration-response curves revealed that the extracts with more significant effects were dichloromethanic extracts of T. mexicana [Emax: (103.03±3.32)% and EC50: (159.39±3.72) µg/mL) and C. vitifolium [Emax: (106.58±2.42)% and EC50: (219.54±7.61) µg/mL]. Finally, hexanic and dichloromethanic extracts from A. mexicana were fully effective but less potent than T. mexicana and C. vitifolium. CONCLUSIONS: Less polar extracts obtained from A. mexicana, T. mexicana and C. vitifolium exhibited greater relaxant effect on tracheal rat rings, which allows us to suggest them as sources for the isolation of bioactive molecules with potential therapeutic value in the treatment of asthma.

Asunto(s)

Extractos Vegetales/farmacología , Plantas Medicinales/química , Tráquea/efectos de los fármacos , Animales , Fraccionamiento Químico , Relación Dosis-Respuesta a Droga , Masculino , Medicina Tradicional , México , Ratas , Ratas Wistar , Fármacos del Sistema Respiratorio/farmacología , Tráquea/química

RESUMEN

ETHNOPHARMACOLOGICAL RELEVANCE: Artemisia ludoviciana spp. mexicana (Willd. Ex.) Spring D.D. Keck (Asteraceae), known as "estafiate" is employed for the treatment of diarrhea, dysentery, parasites, abdominal pain, vomiting, stomach ache, and also as antispasmodic agent. The aim of the present study was to evaluate the relaxant effect of hexanic (HEAl), dichloromethanic (DEAl) and methanolic (MEAl) extracts on isolated trachea, ileum and aorta rat rings, and to establish the tracheo-relaxant mode of action of DEAl. MATERIALS AND METHODS: All extracts were investigated based on their capacity of to inhibit the rat ileum spontaneous contraction, to relax contraction induced by noradrenaline (0.1 µM) on endothelium-intact and endothelium-denuded thoracic aorta rat rings, and also to inhibit contraction provoked by carbachol (1 µM) on rat trachea. RESULTS: Organic extracts had no spasmolytic action on ileum strips compared to positive control (papaverine, p<0.05). On the other hand, all extracts induced a significant concentration- and partial endothelium-dependent vasorelaxant activity. Extracts also showed significant relaxant effect on pre-contracted tracheal tissue in a concentration-dependent manner. In last two experiments, DEAl was the most potent and efficient extract; however, it was less potent than papaverine and theophylline, used as positive controls (p<0.05). In tracheal preparation, DEAl shifted to the right, in a parallel manner, the concentration-response curves induced by carbachol (p<0.05). Also, DEAl induced a significant relaxant effect on the contraction produced by potassium chloride (KCl, 80 mM). Pre-incubation with 1-H-[1,2,4]-oxadiazolo-[4,3a]-quinoxalin-1-one (ODQ, 10 µM), indomethacin (10 µM), N(ω)-nitro-L-arginine methyl ester (L-NAME, 10 µM), glibenclamide (10 µM) and 2-aminopyridine (2-AP, 100 µM) did not modify the DEAl-relaxant curves. CONCLUSIONS: Functional experiments suggest that the most active extract, DEAl, induced its relaxant effect by possible muscarinic receptors antagonism and calcium channel blockade in tracheal rings. On the other hand, significant vasorelaxant activity showed by DEAl is partially endothelium-dependent. Finally, spasmolytic activity induced by the extracts in the rat ileum was not significant, which suggests that the antidiarrheic effect of the plant is related to antimicrobial and antiparasitic properties previously described.

Asunto(s)

Artemisia , Relajación Muscular/efectos de los fármacos , Músculo Liso/efectos de los fármacos , Bloqueantes Neuromusculares/farmacología , Parasimpatolíticos/farmacología , Preparaciones de Plantas/farmacología , Vasodilatación/efectos de los fármacos , Animales , Bloqueadores de los Canales de Calcio/farmacología , Relación Dosis-Respuesta a Droga , Endotelio Vascular/efectos de los fármacos , Íleon/efectos de los fármacos , Masculino , Antagonistas Muscarínicos/farmacología , Músculo Liso Vascular/efectos de los fármacos , Bloqueantes Neuromusculares/química , Parasimpatolíticos/química , Componentes Aéreos de las Plantas , Preparaciones de Plantas/química , Plantas Medicinales , Ratas , Ratas Wistar , Solventes/química , Tráquea/efectos de los fármacos

RESUMEN

AIM OF THE STUDY: Ipomoea tyrianthina has been used in Mexican traditional medicine as a mild purgative, for the treatment of nervous disorders, and against tumors. In this study, the effect of convolvulin (an ether-insoluble resin glycoside) from the root of Ipomoea tyrianthina on: Central Nervous System; as spasmolytic and vasodilator; cytotoxic against cancer cell lines is evaluated. MATERIALS AND METHODS: Convolvulin isolated from the root of Ipomoea tyrianthina (IT-EM) was tested on pentylentetrazole induced seizures, pentobarbital-induced hypnosis, release of GABA and glutamic acid, isolated rat aorta and ileum rings, and against Caco-2 and KB cell lines. RESULTS: IT-EM increased the hypnotic effect induced by pentobarbital and the release of GABA in brain cortex of mice, but did not protect mice against pentylenetetrazole-induced convulsions. IT-EM produced a significant vasodilator effect in concentration- and endothelium-dependent manners on isolated rat aorta, but did not inhibit significantly contractions on rat ileum, colon, and jejune rings. IT-EM showed cytotoxic activity against nasopharyngeal carcinoma KB cell line. CONCLUSIONS: Convolvulin (IT-EM) from Ipomoea tyrianthina has sedative effect, vasorelaxant effect in concentration- and endothelium-dependent manners, and cytotoxic activity against nasopharyngeal carcinoma KB cell line.

Asunto(s)

Antineoplásicos/farmacología , Glicósidos/farmacología , Hipnóticos y Sedantes/farmacología , Ipomoea/química , Extractos Vegetales/química , Resinas de Plantas/farmacología , Vasodilatadores/farmacología , Animales , Línea Celular Tumoral , Humanos , Ratones

RESUMEN

AIM OF THE STUDY: Cochlospermum vitifolium is a medicinal plant used for the treatment of diabetes, hepatobilary and cardiovascular illnesses. The aim of current study was to determine the in vivo antihypertensive and in vitro functional vasorelaxant mechanism of methanol extract of Cochlospermum vitifolium (MECv) and naringenin (NG). MATERIALS AND METHODS: Test material was assayed on rat isolated aorta rings test with- and without-endothelium to determine their vasorelaxant mechanism. Also, the in vivo antihypertensive effect was evaluated on spontaneously hypertensive rat (SHR) model. In addition, presence of NG into the extract was confirmed by reverse phase high performance liquid chromatography (RP-HPLC) analysis. RESULTS: MECv (120 mg/kg) and NG (50 and 160 mg/kg) showed acute antihypertensive effects on SHR when systolic and diastolic pressure were decreased at 1 h and 24 h after administration, respectively. Vasorelaxant effect of MECv and NG was shifted to the right when endothelium-intact aortic rings were pre-incubated with L-NAME (10 microM) and ODQ (1 microM). Also, NG relaxant curves were displaced to the right in the presence of tetraethylammonium (TEA, 1 mM) and 2-aminopyridine (2-AP, 100 microM) on endothelium-denuded aortic rings. CONCLUSION: Experiments described above showed that MECv play an important role in hypertension regulation through NO synthesis and may be PGI(2) production and potassium channel activation on excessive endothelial dysfunction conditions. Unfortunately, presence of NG into the extract is not significant on bioactivity of the extract; however, this compound could be tested and evaluated as structural scaffold for future drug design for development of antihypertensive agents.

Asunto(s)

Antihipertensivos/farmacología , Bixaceae/química , Flavanonas/farmacología , Extractos Vegetales/farmacología , Animales , Antihipertensivos/administración & dosificación , Antihipertensivos/aislamiento & purificación , Aorta Torácica/efectos de los fármacos , Aorta Torácica/metabolismo , Cromatografía Líquida de Alta Presión , GMP Cíclico/metabolismo , Relación Dosis-Respuesta a Droga , Endotelio Vascular/efectos de los fármacos , Endotelio Vascular/metabolismo , Flavanonas/administración & dosificación , Flavanonas/aislamiento & purificación , Hipertensión/tratamiento farmacológico , Hipertensión/fisiopatología , Masculino , Óxido Nítrico/metabolismo , Extractos Vegetales/administración & dosificación , Ratas , Ratas Endogámicas SHR , Ratas Wistar , Transducción de Señal/efectos de los fármacos , Vasodilatadores/administración & dosificación , Vasodilatadores/aislamiento & purificación , Vasodilatadores/farmacología

RESUMEN

Mentha pulegium is common known as "poleo" and used for the treatment of diarrhea, headache and cough in Mexican traditional medicine. Organic extracts from aerial parts were evaluated to determine their spasmolytic action on rat isolated ileum test. Hexanic (HEMp), dichloromethanic (DEMp) and methanolic (MEMp) extracts induced a concentration-dependent (0.97 to 1000 microg/mL) antispasmodic effect on spontaneous contractions. DEMp was the most active extract; therefore, spasmolytic mechanism was investigated. This extract (200 microg/mL) induced a significant depression on cumulative concentration-response curve for carbachol and serotonin (P<0.05). Besides, extract decreased and shifted to the right KCl- and CaCl2-induced contraction curves. Moreover, pre-incubation with chlorpromazine (0.001 mM) shifted to the left the relaxant curve. Pre-treatment with L-NAME (1 mM), papaverine (0.01 mM), teophylline (0.01 mM), TEA (1 mM) and glybenclamide (0.1 mM) did not produced any changed of the relaxant curves of DEMp. Findings indicate that dichloromethanic extract of M. pulegium induced its spasmolytic effect through Ca2+-influx blockade, which may explain its traditional use against diarrhea.

Asunto(s)

Íleon/efectos de los fármacos , Íleon/fisiología , Parasimpatolíticos/farmacología , Animales , Masculino , Mentha pulegium , México , NG-Nitroarginina Metil Éster/farmacología , Ratas , Ratas Wistar

RESUMEN

RMELanc-induced relaxation in aortic rings precontracted with NE, 5-HT and KCl. It also reduced NE-induced transient contraction in Ca(2+)-free solution and inhibited contraction induced by increasing external calcium. Nevertheless, the vasorelaxant effect of RMELanc was not reduced by ODQ, 1-alprenolol, TEA, glibenclamide, and 2-AP. Oral administration of 100 mg/kg of RMELanc exhibited a significant decrease in systolic and diastolic blood pressures in SHR rats. HPLC analysis allowed us to detect the presence of 2,7-dihydroxy-3,4,9-trimethoxyphenantrene (1), which induced a significant relaxation effect. Therefore, our results suggest that RMELanc induces vasorelaxant and antihypertensive effects by blockade of Ca(2+) channels.

Asunto(s)

Antihipertensivos/farmacología , Bloqueadores de los Canales de Calcio/farmacología , Músculo Liso Vascular/efectos de los fármacos , Orchidaceae/química , Fenantrenos/farmacología , Vasodilatadores/farmacología , Animales , Aorta , Presión Sanguínea/efectos de los fármacos , Calcio , Cromatografía Líquida de Alta Presión , Masculino , Contracción Muscular/efectos de los fármacos , Fenantrenos/análisis , Raíces de Plantas , Ratas , Ratas Wistar

RESUMEN

Current investigation was undertaken to elucidate the mode of action of tilianin, isolated from Agastache mexicana, as a vasorelaxant agent on in vitro functional rat thoracic aorta test and to investigate the in vivo antihypertensive effect on spontaneously hypertensive rats (SHR). Tilianin (0.002-933 microM) induced significant relaxation in a concentration- and endothelium-dependent and -independent manners in aortic rings pre-contracted with noradrenaline (NA, 0.1 microM), and serotonin (5-HT, 100 microM). Effect was more significant (p < 0.05) in endothelium-intact (+E) aorta rings than when endothelium was removed(E). Pre-treatment with N-nitro-L-arginine methyl ester (L-NAME; 10 microM) or 1-H-[1,2,4]-oxadiazolo-[4,3a]-quinoxalin-1-one (ODQ, 1 microM) produced a significant change of the relaxant response and activity was markedly inhibited, but not by indomethacin (10 microM) or atropine (1 microM). Furthermore, tilianin (130 microM) provoked a significant displacement to the left in the relaxation curve induced by sodium nitroprusside (SNP; 0.32 nM to 0.1 microM). Moreover, tilianin induced significant in vitro NO overproduction (1.49 +/- 0.86 microM of nitrites/g of tissue) in rat aorta compared with vehicle (p < 0.05). In addition, pre-treatment with tetraethylammonium (TEA, 5 mM) and 2-aminopyridine (2-AP, 0.1 microM) shifted to the right the relaxant curve induced by tilianin (p < 0.05). Finally, a single oral administration of tilianin (50 mg/kg) exhibited a significant decrease in systolic and diastolic blood pressures (p < 0.05) in SHR model. Results indicate that tilianin mediates relaxation mainly by an endothelium-dependent manner,probably due to NO release, and also through an endothelium-independent pathway by opening K+ channels, both causing the antihypertensive effect.

Asunto(s)

Agastache/química , Antihipertensivos/uso terapéutico , GMP Cíclico/fisiología , Flavonoides/uso terapéutico , Glicósidos/uso terapéutico , Activación del Canal Iónico/efectos de los fármacos , Óxido Nítrico/fisiología , Canales de Potasio/fisiología , Vasodilatadores/uso terapéutico , Relación Dosis-Respuesta a Droga , Flavonoides/aislamiento & purificación , Glicósidos/aislamiento & purificación , Humanos , Medicina Tradicional , Metanol , México , Extractos Vegetales/aislamiento & purificación , Extractos Vegetales/farmacología , Plantas Medicinales , Canales de Potasio/efectos de los fármacos

RESUMEN

The aim of the present study was to evaluate the possible mechanism of the vasorelaxant action of methanol extract from Laelia autumnalis (MELa) in isolated rat aortic rings, and to establish its antihypertensive activity in vivo. MELa (0.15-->50 microg/mL) induced relaxation in aortic rings pre-contracted with KCl (80 mM), showing an IC50 value of 34.61+/-1.41 microg/mL and E max value of 85.0+/-4.38% (in endothelium-intact rings) and an IC50 value of 45.11+/-4.17 microg/mL and E max value of 80.0+/-12.1% (in endothelium-denuded rings). Serotonin (5-HT, 1 x 10(-4) M) provoked sustained contraction, which was markedly inhibited by MELa (0.15-->50 microg/mL) in a concentration-dependent and endothelium-independent manner. Pretreatment with MELa (15, 46, 150, 300 and 1500 microg/mL) also inhibited contractile responses to norepinephrine (NE 1 x 10(-11) M to 1 x 10(-5.5) M). In endothelium-denuded rings, the vasorelaxant effect of MELa was reduced partially by ODQ (1 microM), but not by tetraethylammonium (5 microM), glibenclamide (10 microM), and 2-aminopyridine (100 microM). The extract also reduced NE-induced transient contraction in Ca2+-free solution, and inhibited contraction induced by increasing external calcium in Ca2+-free medium plus high KCl (80 mM). The antihypertensive effect of MELa was determined in spontaneously hypertensive rats (SHR). A single oral administration of the extract (100 mg/kg) exhibited a significant decrease in systolic and diastolic blood pressure and heart rate (p<0.05) in SHR rats. Our results suggest that MELa induces relaxation in rat aortic rings through an endothelium-independent pathway, involving blockade of Ca2+ channels and a possible cGMP enhanced concentrations and also causes an antihypertensive effect.

Asunto(s)

Antihipertensivos/farmacología , Bloqueadores de los Canales de Calcio/farmacología , Músculo Liso Vascular/efectos de los fármacos , Orchidaceae/química , Animales , Aorta Torácica/efectos de los fármacos , Presión Sanguínea/efectos de los fármacos , Calcio/farmacología , Técnicas In Vitro , Masculino , Contracción Muscular/efectos de los fármacos , Relajación Muscular/efectos de los fármacos , Norepinefrina/farmacología , Extractos Vegetales/farmacología , Cloruro de Potasio/farmacología , Ratas , Ratas Wistar , Retículo Sarcoplasmático/efectos de los fármacos , Retículo Sarcoplasmático/metabolismo , Serotonina/farmacología

RESUMEN

The present study was undertaken to elucidate the mode of action of methanol extract from aerial parts of L. caulescens (TC-MELc) as spasmolytic agent on in vitro rat ileum test, and investigate the possible antibacterial activity of different extracts from the plant. TC-MELc induced a concentration-dependent (0.001 to 100microg/mL) antispasmodic effect on spontaneous contractions. TC-MELc also (IC50 11.2microg/mL) induced a marked depression on cumulative concentration-response curve for carbachol (Emax=2.3+/-0.3g vs. 0.66+/-0.1g) and serotonin (Emax=1.1+/-0.3g vs. -0.01+/-0.09g). Besides, extract decreased and displaced to the right KCl and CaCl2 concentration-response curves. Moreover, TC-MELc (11.2microg/mL) provoked a total relaxation when ileum strips were contracted with carbachol (1microM) in calcium-free Krebs solution. Pre-treatment with l-NAME (10microM) produced a significant change of the relaxant response and activity was markedly inhibited. Additionally, hexanic (HELc), dichloromethanic (DELc) and methanolic (MELc) extracts from aerial parts were studied to determine their antibacterial activity. DELc showed antibacterial activity on all bacterial strains assayed (

Asunto(s)

Bloqueadores de los Canales de Calcio/farmacología , Óxido Nítrico/biosíntesis , Parasimpatolíticos/farmacología , Extractos Vegetales/farmacología , Plantas Medicinales , Animales , Antibacterianos/farmacología , Relación Dosis-Respuesta a Droga , Masculino , México , Ratas , Ratas Wistar