RESUMEN

Secondary metabolites derived from benzoic acid (BA) are of central importance in the interactions of plants with pests, pathogens, and symbionts and are potentially important in plant development. Peroxisomal ß-oxidation has recently been shown to contribute to BA biosynthesis in plants, but not all of the enzymes involved have been defined. In this report, we demonstrate that the peroxisomal ATP-binding cassette transporter COMATOSE is required for the accumulation of benzoylated secondary metabolites in Arabidopsis (Arabidopsis thaliana) seeds, including benzoylated glucosinolates and substituted hydroxybenzoylcholines. The ABNORMAL INFLORESCENCE MERISTEM protein, one of two multifunctional proteins encoded by Arabidopsis, is essential for the accumulation of these compounds, and MULTIFUNCTIONAL PROTEIN2 contributes to the synthesis of substituted hydroxybenzoylcholines. Of the two major 3-ketoacyl coenzyme A thiolases, KAT2 plays the primary role in BA synthesis. Thus, BA biosynthesis in Arabidopsis employs the same core set of ß-oxidation enzymes as in the synthesis of indole-3-acetic acid from indole-3-butyric acid.

Asunto(s)

Transportadoras de Casetes de Unión a ATP/metabolismo , Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Ácido Benzoico/metabolismo , Complejos Multienzimáticos/metabolismo , Semillas/metabolismo , Transportadoras de Casetes de Unión a ATP/genética , Adenosina Trifosfatasas , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Benzoilcolina/análogos & derivados , Benzoilcolina/química , Benzoilcolina/metabolismo , Colina/química , Colina/metabolismo , Cinamatos/metabolismo , Regulación de la Expresión Génica de las Plantas , Glucosinolatos/metabolismo , Complejos Multienzimáticos/genética , Mutación , Oxidación-Reducción , Canales de Potasio con Entrada de Voltaje/genética , Canales de Potasio con Entrada de Voltaje/metabolismo , Ácido Salicílico/metabolismo , Metabolismo Secundario , Semillas/genética

RESUMEN

Acetylcholinesterases (AChEs) have been estimated in the infective juveniles (IJs) of eight different strains of heterorhabditid nematodes. The enzyme content ranged from 45.6 to 421.3 units/10(5) IJs with specific activity 34.0 to 82.6 units/mg protein. The isoenzyme patterns revealed the existence of two-slow-moving isoforms. Heterorhabditis bacteriophora AChE1A has been purified from the IJs of the heterorhabditid nematode strain of the highest enzymatic activity to homogeneity by ammonium sulfate precipitation, gel filtration on Sephacryl S-200 and DEAE-Sepharose. The specific activity of the purified enzyme was 1378.1 units/mg protein with purification fold 17.5 over crude extract. The enzyme has a pH optimum at 7.5. The optimum temperature for enzyme activity and stability was 35 degrees C. The activation energy was calculated to be 9.0 kcal/mol. The enzyme hydrolyzes acetylthiocholine (AcSCh), propionylthiocholine (PrSCh), S-butyrylthiocholine (BuSCh) and benzoylthiocholine (BzSCh) iodides with relative rate 100, 74.6, 41.7 and 22.2%, respectively. It displayed an apparent Michaelis-Menten behavior in the concentration range from 0.1 to 2 mM for the three former substrates with Km values 0.27, 0.42 and 0.59 mM, respectively. H. bacteriophora ChE1A is an AChE since it hydrolyzed AcSChI at higher rate than the other substrates and displayed excess substrate inhibition with AcSChI at concentrations over 2 mM. It was inhibited by eserine and BW284C51, but not by iso-OMPA. Its biochemical properties were compared with those reported for different species of insects as target hosts for heterorhabditid nematodes and animal parasitic nematodes.

Asunto(s)

Acetilcolinesterasa/aislamiento & purificación , Acetilcolinesterasa/metabolismo , Estadios del Ciclo de Vida/fisiología , Rhabditoidea/enzimología , Acetiltiocolina/metabolismo , Animales , Benzoilcolina/análogos & derivados , Benzoilcolina/metabolismo , Butiriltiocolina/metabolismo , Hidrólisis , Especificidad por Sustrato , Tiocolina/análogos & derivados , Tiocolina/metabolismo

RESUMEN

Catalytic parameters of human butyrylcholinesterase (BuChE) for hydrolysis of homologous pairs of oxo-esters and thio-esters were compared. Substrates were positively charged (benzoylcholine versus benzoylthiocholine) and neutral (phenylacetate versus phenylthioacetate). In addition to wild-type BuChE, enzymes containing mutations were used. Single mutants at positions: G117, a key residue in the oxyanion hole, and D70, the main component of the peripheral anionic site were tested. Double mutants containing G117H and mutations on residues of the oxyanion hole (G115, A199), or the pi-cation binding site (W82), or residue E197 that is involved in stabilization of tetrahedral intermediates were also studied. A mathematical analysis was used to compare data for BuChE-catalyzed hydrolysis of various pairs of oxo-esters and thio-esters and to determine the rate-limiting step of catalysis for each substrate. The interest and limitation of this method is discussed. Molecular docking was used to analyze how the mutations could have altered the binding of the oxo-ester or the thio-ester. Results indicate that substitution of the ethereal oxygen for sulfur in substrates may alter the adjustment of substrate in the active site and stabilization of the transition-state for acylation. This affects the k2/k3 ratio and, in turn, controls the rate-limiting step of the hydrolytic reaction. Stabilization of the transition state is modulated both by the alcohol and acyl moieties of substrate. Interaction of these groups with the ethereal hetero-atom can have a neutral, an additive or an antagonistic effect on transition state stabilization, depending on their molecular structure, size and enantiomeric configuration.

Asunto(s)

Butirilcolinesterasa/metabolismo , Acilación , Sustitución de Aminoácidos , Benzoilcolina/análogos & derivados , Benzoilcolina/metabolismo , Butirilcolinesterasa/genética , Glicolatos/metabolismo , Humanos , Hidrólisis , Cinética , Organofosfatos/metabolismo , Fenilacetatos/metabolismo , Relación Estructura-Actividad , Especificidad por Sustrato , Tiocolina/análogos & derivados , Tiocolina/metabolismo

RESUMEN

The interactions of a photoreactive analogue of benzoylcholine, 4-azido-2,3,5,6-tetrafluorobenzoylcholine (APFBzcholine), with nicotinic acetylcholine receptors (nAChRs) were studied using electrophysiology and photolabeling. APFBzcholine acted as a low-efficacy partial agonist, eliciting maximal responses that were 0.3 and 0.1% of that of acetylcholine for embryonic mouse and Torpedo nAChRs expressed in Xenopus oocytes, respectively. Equilibrium binding studies of [3H]APFBzcholine with nAChR-rich membranes from Torpedo electric organ revealed equal affinities (K(eq) = 12 microM) for the two agonist binding sites. Upon UV irradiation at 254 nm, [3H]APFBzcholine was photoincorporated into the nAChR alpha, gamma, and delta subunits in an agonist-inhibitable manner. Photolabeled amino acids in the agonist binding sites were identified by Edman degradation of isolated, labeled subunit fragments. [3H]APFBzcholine photolabeled gammaLeu-109/deltaLeu-111, gammaTyr-111, and gammaTyr-117 in binding site segment E as well as alphaTyr-198 in alpha subunit binding site segment C. The observed pattern of photolabeling is examined in relation to the predicted orientation of the azide when APFBzcholine is docked in the agonist binding site of a homology model of the nAChR extracellular domain based upon the structure of the snail acetylcholine binding protein.

Asunto(s)

Benzoilcolina/análogos & derivados , Agonistas Nicotínicos/química , Etiquetas de Fotoafinidad/química , Receptores Nicotínicos/química , Receptores Nicotínicos/fisiología , Animales , Benzoilcolina/química , Benzoilcolina/farmacología , Electrofisiología , Receptores Nicotínicos/efectos de los fármacos , Torpedo

RESUMEN

The rate-limiting step for hydrolysis of the positively charged oxoester benzoylcholine (BzCh) by human butyrylcholinesterase (BuChE) is deacylation (k(3)), whereas it is acylation (k(2)) for hydrolysis of the homologous thioester benzoylthiocholine (BzSCh). Steady-state hydrolysis of BzCh and BzSCh by wild-type BuChE and its peripheral anionic site mutant D70G was investigated at different hydrostatic pressures, which allowed determination of volume changes associated with substrate binding, and the activation volumes for the chemical steps. A differential nonlinear pressure-dependence of the catalytic parameters for hydrolysis of both substrates by both enzymes was shown. Nonlinearity of the plots may be explained in terms of compressibility changes or rate-limiting changes. To distinguish between these two possibilities, enzyme phosphorylation by diisopropylfluorophosphate (DFP) in the presence of substrate (BzSCh) under pressure was studied. There was no pressure dependence of volume changes for DFP binding or for phosphorylation of either wild-type or D70G. Analysis of the pressure dependence for steady-state hydrolysis of substrates, and for phosphorylation by DFP provided evidence that no enzyme compressibility changes occurred during the catalyzed reactions. Thus, the nonlinear pressure dependence of substrate hydrolysis reflects changes in the rate-limiting step with pressure. Change in rate-determining step occurred at a pressure of 100 MPa for hydrolysis of BzCh by wild-type and at 75 MPa for D70G. For hydrolysis of BzSCh the change occurred at higher pressures because k(2) << k(3) at atmospheric pressure for this substrate. Elementary volume change contributions upon initial binding, productive binding, acylation and deacylation were calculated from the pressure differentiation of kinetic constants. This analysis shed light on the molecular events taking place along the hydrolysis pathways of BzCh and BzSCh by wild-type BuChE and the D70G mutant. In addition, volume change differences between wild-type and D70G provided new evidence that residue D70 in the peripheral site controls hydration of the active site gorge and the dynamics of the water molecule network during catalysis. Finally, a steady-state kinetic study of the oxyanion hole mutant (G117H) showed that substitution of the ethereal sulfur for oxygen in the substrate alters the final adjustment of substrate in the active site and stabilization of the acylation transition state.

Asunto(s)

Benzoilcolina/análogos & derivados , Benzoilcolina/metabolismo , Butirilcolinesterasa/metabolismo , Sustitución de Aminoácidos , Presión Atmosférica , Sitios de Unión , Butirilcolinesterasa/química , Butirilcolinesterasa/genética , Catálisis , Activación Enzimática , Humanos , Hidrólisis , Isoflurofato/metabolismo , Cinética , Modelos Moleculares , Fosforilación , Unión Proteica , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo

RESUMEN

A special component is isolated from Semen sinapis Albae (white mustard seed), a traditional Chinese medicine. According to the physical and chemical investigation and spectroscopic identification, this component can be known as p-hydroxybenzoylcholine bisulfate, a choline base. This component in the drug is also determined by RP-HPLC. A reversed-phase C(18) column is used to separate the p-hydroxybenzoylcholine with an eluent of methanol-0.05 mol/L monopotassium phosphate solution (30:70) (adjusted by phosphoric acid to pH 3.6) at the flow rate of 0.5 mL/min. Detection is carried out with a UV detector operated at 285 nm, and the column temperature is 25 degrees C. It reveals that there is 0.021% (w/w) of p-hydroxybenzoylcholine bisulfate in Semen sinapis Albae and 0.037% (w/w) in stir-baked Semen sinapis Albae.

Asunto(s)

Benzoilcolina/análisis , Benzoilcolina/aislamiento & purificación , Medicina Tradicional China , Sinapis/química , Benzoilcolina/análogos & derivados , Benzoilcolina/química , Medicamentos Herbarios Chinos/química , Espectroscopía de Resonancia Magnética , Estructura Molecular , Sensibilidad y Especificidad

RESUMEN

[(3)H]4-[(3-trifluoromethyl)-3H-diazirin-3-yl]benzoylcholine (TDBzcholine) was synthesized and used as a photoaffinity probe to map the orientation of an aromatic choline ester within the agonist binding sites of the Torpedo nicotinic acetylcholine receptor (nAChR). TDBzcholine acts as a nAChR competitive antagonist that binds at equilibrium with equal affinity to both agonist sites (K(D) approximately 10 microM). Upon UV irradiation (350 nm), nAChR-rich membranes equilibrated with [(3)H]TDBzcholine incorporate (3)H into the alpha, gamma, and delta subunits in an agonist-inhibitable manner. The specific residues labeled by [(3)H]TDBzcholine were determined by N-terminal sequence analysis of subunit fragments produced by enzymatic cleavage and purified by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and/or reversed-phase high-performance liquid chromatography. For the alpha subunit, [(3)H]TDBzcholine photoincorporated into alphaCys-192, alphaCys-193, and alphaPro-194. For the gamma and delta subunits, [(3)H]TDBzcholine incorporated into homologous leucine residues, gammaLeu-109 and deltaLeu-111. The photolabeling of these amino acids suggests that when the antagonist TDBzcholine occupies the agonist binding sites, the Cys-192-193 disulfide and Pro-194 from the alpha subunit Segment C are oriented toward the agonist site and are in proximity to gammaLeu-109/deltaLeu-111 in Segment E, a conclusion consistent with the structure of the binding site in the molluscan acetylcholine binding protein, a soluble protein that is homologous to the nAChR extracellular domain.

Asunto(s)

Aminoácidos/análisis , Azirinas/metabolismo , Benzoilcolina/análogos & derivados , Benzoilcolina/metabolismo , Colina/metabolismo , Canales Iónicos/metabolismo , Agonistas Nicotínicos/metabolismo , Antagonistas Nicotínicos/metabolismo , Etiquetas de Fotoafinidad/metabolismo , Receptores Nicotínicos/metabolismo , Acetilcolina/metabolismo , Secuencia de Aminoácidos , Aminoácidos/metabolismo , Venenos de Anfibios/metabolismo , Animales , Azirinas/farmacología , Benzoilcolina/farmacología , Sitios de Unión , Unión Competitiva , Bungarotoxinas/metabolismo , Membrana Celular/metabolismo , Colina/análogos & derivados , Colina/farmacología , Radioisótopos de Yodo , Datos de Secuencia Molecular , Antagonistas Nicotínicos/farmacología , Fragmentos de Péptidos/metabolismo , Isoformas de Proteínas/metabolismo , Subunidades de Proteína/análisis , Subunidades de Proteína/metabolismo , Torpedo , Tritio , Rayos Ultravioleta , Xenopus

RESUMEN

[(3)H]4-Benzoylbenzoylcholine (Bz(2)choline) was synthesized as a photoaffinity probe for the Torpedo nicotinic acetylcholine receptor (nAChR). [(3)H]Bz(2)choline acts as an nAChR competitive antagonist and binds at equilibrium with the same affinity (K(D) = 1.4 microm) to both agonist sites. Irradiation at 320 nm of nAChR-rich membranes equilibrated with [(3)H]Bz(2)choline results in the covalent incorporation of [(3)H]Bz(2)choline into the nAChR gamma- and delta-subunits that is inhibitable by agonist, with little specific incorporation in the alpha-subunits. To identify the sites of photoincorporation, gamma- and delta-subunits, isolated from nAChR-rich membranes photolabeled with [(3)H]Bz(2)choline, were digested enzymatically, and the labeled fragments were isolated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and/or reversed-phase high performance liquid chromatography. For the gamma-subunit, Staphylococcus aureus V8 protease produced a specifically labeled peptide beginning at gammaVal-102, whereas for the delta-subunit, endoproteinase Asp-N produced a labeled peptide beginning at deltaAsp-99. Amino-terminal sequence analysis identified the homologous residues gammaLeu-109 and deltaLeu-111 as the primary sites of [(3)H]Bz(2)choline photoincorporation. This is the first identification by affinity labeling of non-reactive amino acids within the acetylcholine-binding sites, and these results establish that when choline esters of benzoic acid are bound to the nAChR agonist sites, the para substituent is selectively oriented toward and in proximity to amino acids gammaLeu-109/deltaLeu-111.

Asunto(s)

Benzoilcolina/análogos & derivados , Antagonistas Nicotínicos/metabolismo , Etiquetas de Fotoafinidad/metabolismo , Receptores Nicotínicos/química , Benzoilcolina/metabolismo , Unión Competitiva , Bungarotoxinas/metabolismo

RESUMEN

Effects of rat and human calcitonin gene-related peptide (r alpha CGRP and h beta CGRP, respectively) upon uterine contractile force were investigated using uterine horns from nonpregnant rats, r alpha CGRP and h beta CGRP were equipotent (pD2 = 8.85-9.09) in inhibiting spontaneous and electrically evoked uterine contractions. r alpha CGRP was relatively ineffective in inhibiting potassium-induced contractures of preparations from stilbestrol-pretreated rats. The use of selective antagonists established that r alpha CGRP did not release prostanoids, or release or act at receptors for catecholamines and histamine. The effects of the peptides were not significantly modulated by estrogen levels since pD2 values were similar (8.56-8.86) in field-stimulated preparations from rats in proestrus/estrus or metestrus/diestrus.

Asunto(s)

Péptido Relacionado con Gen de Calcitonina/farmacología , Contracción Uterina/efectos de los fármacos , Animales , Atropina/farmacología , Benzoilcolina/análogos & derivados , Benzoilcolina/farmacología , Depresión Química , Dietilestilbestrol/farmacología , Estimulación Eléctrica , Estro , Femenino , Humanos , Indometacina/farmacología , Isoproterenol/farmacología , Potasio/farmacología , Ranitidina/farmacología , Ratas , Proteínas Recombinantes/farmacología

RESUMEN

Flounder (Platichthys flesus) muscle contains two types of cholinesterases, that differ in molecular form and in substrate specificity. Both enzymes were purified by affinity chromatography. About 8% of cholinesterase activity could be attributed to collagen-tailed asymmetric acetylcholinesterase sedimenting at 17S, 13S and 9S, which showed catalytic properties of a true acetylcholinesterase. 92% of cholinesterase activity corresponded to an amphiphilic dimeric enzyme sedimenting at 6S in the presence of Triton X-100. Treatment with phospholipase C yielded a hydrophilic form and uncovered an epitope called the cross-reacting determinant, which is found in the hydrophilic form of a number of glycosyl-phosphatidylinositol-anchored proteins. This enzyme showed catalytic properties intermediate to those of acetylcholinesterase and butyrylcholinesterase. It hydrolyzed acetylthiocholine, propionylthiocholine, butyrylthiocholine and benzoylthiocholine. The Km and the maximal velocity decreased with the length and hydrophobicity of the acyl chain. At high substrate concentrations the enzyme was inhibited. The p(IC50) values for BW284C51 and ethopropazine were between those found for acetylcholinesterase and butylcholinesterase. For purified detergent-soluble cholinesterase a specific activity of 8000 IU/mg protein, a turnover number of 2.8 x 10(7) h-1, and 1 active site/subunit were determined.

Asunto(s)

Colinesterasas/aislamiento & purificación , Colágeno/metabolismo , Glucolípidos/metabolismo , Músculos/enzimología , Fosfatidilinositoles/metabolismo , Acilación , Animales , Benzoilcolina/análogos & derivados , Sitios de Unión , Western Blotting , Catálisis , Centrifugación por Gradiente de Densidad , Cromatografía de Afinidad , Reacciones Cruzadas , Electroforesis en Gel de Poliacrilamida , Endopeptidasa K , Peces , Glicosilfosfatidilinositoles , Cinética , Polietilenglicoles , Serina Endopeptidasas , Especificidad por Sustrato , Tiocolina/análogos & derivados , Fosfolipasas de Tipo C

Asunto(s)

Benzoilcolina/análogos & derivados , Butirilcolinesterasa/análisis , Colina/análogos & derivados , Colinesterasas/análisis , Benzoilcolina/síntesis química , Humanos , Indicadores y Reactivos , Cinética , Especificidad por Sustrato

RESUMEN

We report the evaluation of a new commercially available assay system for determination of the catalytic activity of serum cholinesterase (EC 3.1.1.8) and application of the method to a centrifugal fast analyzer. Serum cholinesterase activity is determined at 30 degrees C using para-hydroxybenzoylcholine as substrate. This reaction is coupled to a second reaction using para-hydroxybenzoate hydroxylase (EC 1.14.13.2) as coupling enzyme. Enzyme activity is measured kinetically by monitoring the decrease in absorbance at 340 nm of NADPH in the second reaction. The procedure is precise and the results obtained from normal and pathological sera show good correlation with those obtained by the alternative procedures employing propionylthiocholine, butyrylthiocholine and benzoylcholine as substrates. The reference range for 700 healthy subjects was estimated to be 140-345 U/L (95% central range, determined non-parametrically), with significant difference between males and females (155-353 U/L for men and 134-323 U/L for women, p less than 0.001).

Asunto(s)

Acetilcolinesterasa/sangre , Pruebas Enzimáticas Clínicas/métodos , Acetilcolinesterasa/genética , Acetilcolinesterasa/normas , Adolescente , Adulto , Anciano , Benzoilcolina/análogos & derivados , Catálisis , Niño , Inhibidores de la Colinesterasa/farmacología , Pruebas Enzimáticas Clínicas/normas , Dibucaína/farmacología , Femenino , Humanos , Masculino , Persona de Mediana Edad , Fenotipo , Valores de Referencia , Especificidad por Sustrato

RESUMEN

A simple and reproducible method for the determination of serum pseudo-cholinesterase activity was developed by making use of a stable substrate, p-hydroxybenzoylcholine, with p-hydroxybenzoate hydroxylase as a linked enzyme. The method is based on spectrophotometric measurement of the decrease of NADPH. p-Hydroxybenzoate released from p-hydroxybenzoylcholine is hydroxylated by the action of p-hydroxybenzoate hydroxylase in the presence of NADPH and O2 to produce 3,4-dihydroxybenzoate and NADP+. This method is superior to the conventional methods in that this substrate is extremely stable up to pH 9.0, which is close to the optimum pH for the assay (pH 8.0). Serum interference was resolved by the use of p-hydroxybenzoate hydroxylase as a linked enzyme. The Km value of pseudocholinesterase for p-hydroxybenzoylcholine is 1 X 10(-5) M. The results of our method and Garry's method (Clin. Chem. 11, 91-96, 1965) correlated well (r = 0.962). The within-run and between-run C.V. values were 2.1 and 2.7, respectively.

Asunto(s)

Butirilcolinesterasa/sangre , Colinesterasas/sangre , 4-Hidroxibenzoato-3-Monooxigenasa , Benzoilcolina/análogos & derivados , Benzoilcolina/síntesis química , Humanos , Concentración de Iones de Hidrógeno , Cinética

Asunto(s)

Colinesterasas/sangre , Acetilcolina , Oxidorreductasas de Alcohol , Animales , Benzoilcolina/análogos & derivados , Colorimetría , Humanos , Métodos , Especificidad por Sustrato , Tiocolina

RESUMEN

Two radioiodinated analogs of benzoylcholine were investigated as possible myocardial-imaging agents. O-([2-125I]Iodobenzoyl)-choline and N-([2-125I]iodobenzoyl)cholamine were prepared by nucleophilic substitution of sodium [125I]iodide for stable iodine in O-(N,N]dimethylaminoethyl)-2-iodobenzoate and N-(N',N'-dimethylaminoethyl)-2-iodobenzamide, respectively, and by methylation with methyl iodide. The in vivo distribution of each compound in mice was determined as a function of time. Favorable heart-to-blood and heart-to-lung ratios were obtained with N-([2-125I]iodobenzoyl)cholamine.

Asunto(s)

Benzoilcolina/análogos & derivados , Colina/análogos & derivados , Corazón/diagnóstico por imagen , Animales , Benzoilcolina/metabolismo , Radioisótopos de Yodo , Marcaje Isotópico , Ratones , Cintigrafía , Distribución Tisular

RESUMEN

1. The interaction of serotonin precursor L-tryptophan with the pressor responses of the anaesthetized rat to the intravenous injection of clonidine, adrenaline and angiotensin has been studied. 2. Pretreatment of rats with L-tryptophan (100 mg/kg) depressed the pressor response to clonidine but had no effect on the responses elicited by adrenaline or angiotensin. 3. The L-tryptophan-induced depression of the clonidine response was prevented by pretreating rats with either Rö 4-4602, carbidopa, BW 172C58, methysergide or by pithing. 4. Intravenous infusions of serotonin depressed the pressor responses to clonidine, adrenaline and angiotensin in both intact anaesthetized and pithed rats. 5. It is concluded that the depressant action of L-tryptophan is dependent on its conversion within the periphery to serotonin. This action is also dependent on or mediated by the sympathetic nervous system.

Asunto(s)

Presión Sanguínea/efectos de los fármacos , Clonidina/antagonistas & inhibidores , Triptófano/farmacología , Angiotensina II/farmacología , Animales , Benserazida/farmacología , Benzoilcolina/análogos & derivados , Benzoilcolina/farmacología , Carbidopa/farmacología , Epinefrina/farmacología , Masculino , Metisergida/farmacología , Ratas , Serotonina/farmacología