RESUMEN
Valproic acid (VPA) has been shown to cause neural tube defects in humans and mice, but its mechanism of action has not been elucidated. We hypothesize that alterations in embryonic antioxidant status and Hoxa2 gene expression play an important role in VPA-induced teratogenesis. A whole embryo culture system was applied to explore the effects of VPA on total glutathione, on glutathione in its oxidized (GSSG) and reduced (GSH) forms [GSSG/GSH ratio] and on Hoxa2 expression in cultured CD-1 mouse embryos during their critical period of organogenesis. Our results show that VPA can (1) induce embryo malformations including neural tube defects, abnormal flexion, yolk sac circulation defects, somite defects, and craniofacial deformities such as fusion of the first and second arches, and (2) alter glutathione homeostasis of embryos through an increase in embryonic GSSG/GSH ratio and a decrease in total GSH content in embryos. Western blot analysis and quantitative real-time RT-PCR show that VPA can inhibit Hoxa2 expression in cultured embryos at both the protein and mRNA level, respectively. The presence of ascorbic acid in the culture media was effective in protecting embryos against oxidative stress induced by VPA and prevented VPA-induced inhibition of Hoxa2 gene expression. Hoxa2 null mutant embryos do not exhibit altered glutathione homeostasis, indicating that inhibition of Hoxa2 is downstream of VPA-induced oxidative stress. These results are first to suggest VPA may, in part, exert its teratogenicity through alteration of the embryonic antioxidant status and inhibition of Hoxa2 gene expression and that ascorbic acid can protect embryos from VPA-induced oxidative stress.
Asunto(s)
Ácido Ascórbico/farmacología , Técnicas de Cultivo de Embriones/métodos , Embrión de Mamíferos/metabolismo , Glutatión/metabolismo , Proteínas de Homeodominio/antagonistas & inhibidores , Homeostasis/efectos de los fármacos , Ácido Valproico/farmacología , Animales , Antioxidantes/metabolismo , Embrión de Mamíferos/anomalías , Embrión de Mamíferos/efectos de los fármacos , Regulación del Desarrollo de la Expresión Génica/efectos de los fármacos , Proteínas de Homeodominio/genética , Proteínas de Homeodominio/metabolismo , Ratones , Modelos Biológicos , Reacción en Cadena de la Polimerasa de Transcriptasa InversaRESUMEN
1. Homeobox (Hox) genes were originally discovered in the fruit fly Drosophila, where they function through a conserved homeodomain as transcriptional regulators to control embryonic morphogenesis. Since then over 1000 homeodomain proteins have been identified in several species. In vertebrates, 39 Hox genes have been identified as homologs of the original Drosophila complex, and like their Drosophila counterparts they are organized within chromosomal clusters. Vertebrate Hox genes have also been shown to play a critical role in embryonic development as transcriptional regulators. 2. Both the Drosophila and vertebrate Hox genes have been shown to interact with various cofactors, such as the TALE homeodomain proteins, in recognition of consensus sequences within regulatory elements of their target genes. These protein-protein interactions are believed to contribute to enhancing the specificity of target gene recognition in a cell-type or tissue- dependent manner. The regulatory activity of a particular Hox protein on a specific regulatory element is highly variable and dependent on its interacting partners within the transcriptional complex. 3. In vertebrates, Hox genes display spatially restricted patterns of expression within the developing CNS, both along the anterioposterior and dorsoventral axis of the embryo. Their restricted gene expression is suggestive of a regulatory role in patterning of the CNS, as well as in cell specification. Determining the precise function of individual Hox genes in CNS morphogenesis through classical mutational analyses is complicated due to functional redundancy between Hox genes. 4. Understanding the precise mechanisms through which Hox genes mediate embryonic morphogenesis requires the identification of their downstream target genes. Although Hox genes have been implicated in the regulation of several pathways, few target genes have been shown to be under their direct regulatory control. Development of methodologies used for the isolation of target genes and for the analysis of putative targets will be beneficial in establishing the genetic pathways controlled by Hox factors. 5. Within the developing CNS various cell adhesion molecules and signaling molecules have been identified as candidate downstream target genes of Hox proteins. These targets play a role in processes such as cell migration and differentiation, and are implicated in contributing to neuronal processes such as plasticity and/or specification. Hence, Hox genes not only play a role in patterning of the CNS during early development, but may also contribute to cell specification and identity.
Asunto(s)
Sistema Nervioso Central/embriología , Sistema Nervioso Central/fisiología , Regulación del Desarrollo de la Expresión Génica/fisiología , Genes Homeobox/fisiología , Animales , HumanosRESUMEN
A series of 3,5-bis(arylidene)-4-piperidones 1 and related N-acryloyl analogues 2 were prepared as candidate cytotoxic agents with a view to discerning those structural features which contributed to bioactivity. A number of the compounds were markedly cytotoxic toward murine P388 and L1210 leukemic cells and also to human Molt 4/C8 and CEM neoplasms. Approximately 40% of the IC50 values generated were lower than the figures obtained for melphalan. In virtually all cases, the N-acyl compounds were significantly more bioactive than the analogues 1. In general, structure-activity relationships revealed that the cytotoxicity of series 1 was correlated positively with the size of the aryl substituents, while in series 2, a -sigma relationship was established. In particular, various angles and interatomic distances were obtained by molecular modeling, and the presence of an acryloyl group on the piperidyl nitrogen atom in series 2 affected the relative locations of the two aryl rings. This observation, along with some differences in distances between various atoms in series 1 and 2, may have contributed to the disparity in cytotoxicity between 1 and 2. The results obtained by X-ray crystallography of representative compounds were mainly in accordance with the observations noted by molecular modeling. Selected compounds interfered with the biosynthesis of DNA, RNA, and protein in murine L1210 cells, while others were shown to cause apoptosis in the human Jurkat leukemic cell line. This study has revealed the potential of these molecules for development as cytotoxic and anticancer agents.
Asunto(s)
Acrilatos/síntesis química , Antineoplásicos/síntesis química , Piperidinas/síntesis química , Acrilatos/química , Acrilatos/farmacología , Animales , Antifúngicos/síntesis química , Antifúngicos/química , Antifúngicos/farmacología , Antineoplásicos/química , Antineoplásicos/farmacología , Aspergillus fumigatus/efectos de los fármacos , Candida albicans/efectos de los fármacos , Cristalografía por Rayos X , Ensayos de Selección de Medicamentos Antitumorales , Humanos , Ratones , Modelos Moleculares , Oxidación-Reducción , Piperidinas/química , Piperidinas/farmacología , ARN/antagonistas & inhibidores , ARN/biosíntesis , Relación Estructura-Actividad , Células Tumorales CultivadasRESUMEN
Coordinated expression of Hoxa2, Hoxd1 and Pax6 proteins were found to coincide with the three developmental stages of the diencephalon, as described for the mouse brain. In the first stage (embryonic day (E) 10-12) Hoxa2, Hoxd1 and Pax6 (an early marker gene of the diencephalon) were expressed as early as E10.5 in prosomeres (p), p2 and p3. All three proteins continue to exhibit overlapping domains of expression at E12.5-13 (beginning of the second stage) when the primitive dense cell layer begins to differentiate into the internal germinal, external germinal and mantle layers. Towards the end of the second stage (E15), Pax6 expression was down-regulated whereas Hoxa2 and Hoxd1 continued to exhibit overlapping domains of expression for both protein and mRNA. Hoxd1 expression decreased significantly in the third stage of diencephalic development (E16-postnatal) such that only Hoxa2 expression persisted in the diencephalon of newborn mice. The temporal and spatial expression of these three proteins imply that coordinated waves of Hoxa2, Hoxd1 and Pax6 expression may be required to provide positional information for the specification of the diencephalon.
Asunto(s)
Diencéfalo/embriología , Regulación del Desarrollo de la Expresión Génica , Proteínas de Homeodominio/genética , Animales , Diencéfalo/química , Proteínas del Ojo , Proteínas de Homeodominio/análisis , Inmunohistoquímica , Hibridación in Situ , Ratones , Ratones Endogámicos , Factor de Transcripción PAX6 , Factores de Transcripción Paired Box , ARN Mensajero/análisis , Proteínas RepresorasRESUMEN
1. Hoxd1 is member of the labial subfamily of Hox genes that has a conserved 60 amino acid homeodomain region. The homeodomain is an important determining factor in the binding of the protein to specific DNA sequence(s). DNA-binding specificity for the Hoxd1 protein has not been determined previously. 2. We have employed a rapid affinity chromatography method to determine optimal DNA binding sequences for the 109 amino acid Hoxd1 peptide, comprising the homeodomain and the entire carboxy terminal region of the Hoxd1 protein. 3. Labial Hox proteins have intrinsically weak DNA-binding activity that has been attributed to the nonbasic residues at positions 2 and 3 in the N-terminal arm of the homeodomain. The presence of the Hoxd1 carboxy terminal region negated the influence of the nonbasic residues and facilitated Hoxd1 DNA-binding specificity. 4. DNA sequences bound to the Hoxd1 peptide-affinity column were separated from a random pool of oligonucleotide sequences by gradient elution and enriched by polymerase chain reaction. Preferred sequences were identified on 5' and 3' of a TAAT core, extending the binding site to T/AT/gTAATTGTA. 5. Stability and specificity of optimal DNA-binding sequence for Hoxd1 homeodomain were determined by equilibrium and kinetic studies. Dissociation coefficient constant (KD) was estimated to be 8.6 x 10(-9) M and the DNA-Hoxd1 homeodomain complex has a half life (t(1/2)) of 12.7 min. 6. A molecular model of Hoxd1 homeodomain-DNA interaction based on the X-ray coordinates of Antennapedia homeodomain-DNA complex has revealed novel interactions of key Hoxd1 residues at the protein-DNA interface.
Asunto(s)
ADN/metabolismo , Proteínas de Homeodominio/metabolismo , Modelos Moleculares , Sondas de Oligonucleótidos/metabolismo , Secuencia de Aminoácidos , Sitios de Unión , Cromatografía de Afinidad/métodos , ADN/química , ADN/genética , Escherichia coli/genética , Escherichia coli/metabolismo , Proteínas de Homeodominio/química , Proteínas de Homeodominio/genética , Datos de Secuencia Molecular , Sondas de Oligonucleótidos/genética , Distribución AleatoriaRESUMEN
Five series of novel compounds were synthesized in order to evaluate the theory of sequential cytotoxicity which seeks to exploit the view that various cancer cells are particularly susceptible to successive attacks by cytotoxic agents. The compounds prepared were various 2-[4-(3-aryl-2-propenoyloxy)phenylmethylene]cyclohexanone s 1 and the related Mannich bases 2. In addition the analogues 3-5 lacking an olefinic bond in the ester group were also synthesized, which were predicted to be less cytotoxic than the compounds of series 1 and 2. The atomic charges at the potential sites for interaction with cellular constituents were determined by molecular modeling calculations. The biodata obtained from murine and human neoplastic cells revealed that the predictions made regarding the viability of the theory were fulfilled in approximately two-thirds of the cases indicating that further investigation of this hypothesis is warranted. In addition, the significant potencies of some of the Mannich bases toward human tumor cell lines, in particular coupled to their selective toxicity toward human leukemic and colon cancer cells, confirms their usefulness in serving as lead molecules for further development. A preliminary investigation into the mode of action of representative compounds revealed their ability to induce apoptosis and inhibit the biosyntheses of ribonucleic acid and proteins.
Asunto(s)
Antineoplásicos/síntesis química , Ciclohexanonas/síntesis química , Bases de Mannich/síntesis química , Animales , Antineoplásicos/química , Antineoplásicos/farmacología , Apoptosis , Cristalografía por Rayos X , Ciclohexanonas/química , Ciclohexanonas/farmacología , Ensayos de Selección de Medicamentos Antitumorales , Humanos , Bases de Mannich/química , Bases de Mannich/farmacología , Ratones , Modelos Moleculares , Proteínas de Neoplasias/antagonistas & inhibidores , Proteínas de Neoplasias/biosíntesis , ARN Neoplásico/antagonistas & inhibidores , ARN Neoplásico/biosíntesis , Relación Estructura-Actividad , Células Tumorales CultivadasRESUMEN
A number of 2-arylidenecyclohexanones 1, 2, 6-bis(arylidene)cyclohexanones 2 and related Mannich bases 3-5 were prepared. Various torsion angles as well as atomic charges on olefinic carbon atoms were determined by molecular modelling on all compounds. These molecules showed cytotoxicity towards murine P388 and L1210 cells as well as to human Molt 4/C8 and CEM T-lymphocytes. The average cytotoxicity of the dienones 2 was more than three times greater than was found with the monoarylidene analogues 1, and, in general, were slightly more cytotoxic than the Mannich bases 3-5. A number of the compounds displayed potency towards a panel of human tumour cell lines and most of the representative compounds in series 2-5 were selectively toxic to colon cancers and leukaemic cells.
Asunto(s)
Antineoplásicos/farmacología , Ciclohexanonas/farmacología , Animales , Antineoplásicos/química , Cristalografía por Rayos X , Ciclohexanonas/química , Ensayos de Selección de Medicamentos Antitumorales , Humanos , Ratones , Estructura Molecular , Células Tumorales CultivadasRESUMEN
Various 2-arylideneindanones 1, 2-arylidenetetralones 2, and 2-arylidenebenzosuberones 3 were synthesized with the aim of determining the relative orientations of the two aryl rings which favored cytotoxicity. Molecular modeling of the unsubstituted compound in each series revealed differences in the spatial arrangements of the two aryl rings, and evaluation of these compounds against P388, L1210, Molt 4/C8, and CEM cells as well as a panel of human tumor cell lines indicated that in general the order of cytotoxicity was 3 > 2 > 1. In particular 2-(4-methoxyphenylmethylene)-1-benzosuberone (3k) had the greatest cytotoxicity, possessing 11 times the potency of the reference drug melphalan when all five screens were considered. Series 3 was considered in further detail. First, excision of the aryl ring fused to the cycloheptanone moiety in series 3 led to some 2-arylidene-1-cycloheptanones 4 which had approximately one-third of the bioactivity of the analogues 3. Second, in some screens cytotoxicity was correlated negatively with the sigma values and positively with the MR constants of the substituents in the arylidene aryl ring of 3. Third, X-ray crystallography of five representative compounds (3i,k-n) revealed differences in the locations of the aryl rings which may have contributed to the variations in cytotoxicity. Finally three members of series 3 inhibited RNA and protein syntheses and induced apoptosis in human Jurkat T cells. This study has revealed that 2-arylidene-1-benzosuberones are a group of useful cytotoxic agents, and in particular 3k serves as a prototypic molecule for subsequent structural modifications.
Asunto(s)
Antineoplásicos/síntesis química , Ciclohexanonas/síntesis química , Indanos/síntesis química , Tetrahidronaftalenos/síntesis química , Animales , Antineoplásicos/química , Antineoplásicos/farmacología , Cristalografía por Rayos X , Ciclohexanonas/química , Ciclohexanonas/farmacología , Ensayos de Selección de Medicamentos Antitumorales , Humanos , Indanos/química , Indanos/farmacología , Modelos Moleculares , Conformación Molecular , Relación Estructura-Actividad , Tetrahidronaftalenos/química , Tetrahidronaftalenos/farmacología , Células Tumorales CultivadasRESUMEN
1. Apoptosis as the mechanism of cell death induced by a new cytotoxic and anticancer agent (N.C.1213) was investigated by morphological and biochemical criteria in human Jurkat T leukemia cells. 2. The effect of N.C.1213 on the survival of Jurkat T, LV-50, H-9, and Molt-3 cells was measured. Jurkat T cells exhibited the highest response, with less than 10% of the cells remaining viable after exposure to 10 microM N.C.1213 for a 24 hr period. All other cell cultures were also affected but to a lesser extent. 3. With the use of a fluorescence microscope, several morphological features characteristic of apoptosis such as condensed chromatin and apoptotic bodies were identified in Jurkat T cells after exposure to N.C.1213 and melphalan. The results indicated that melphalan was more cytotoxic than N.C.1213 as shown by the dye exclusion test. However, N.C.1213 showed a greater apoptotic index than melphalan. The IC50 of N.C.1213 in Jurkat T cells was determined to be 3.5 microM. 4. A DNA ladder (fragmentation of DNA into multimers of approximately 200 base pairs), which is one characteristic feature of apoptosis, was not detected when Jurkat T cells were exposed to N.C.1213. Hence it is probable that the key morphological events in apoptosis observed in the present experimental conditions precede the internucleosomal cleavage of DNA.
Asunto(s)
Antineoplásicos/farmacología , Apoptosis/efectos de los fármacos , Melfalán/toxicidad , Piperidinas/toxicidad , Supervivencia Celular/efectos de los fármacos , Fragmentación del ADN , Humanos , Células Jurkat/citología , Células Jurkat/efectos de los fármacos , Células Jurkat/fisiología , Cinética , Microscopía Fluorescente/métodos , Células Tumorales CultivadasRESUMEN
1. Concentrations of the neurotransmitter amines noradrenaline (NA), dopamine (DA), and 5-hydroxytryptamine (5-HT) and the acid metabolites homovanillic acid (HVA) and 5-hydroxyindoleacetic acid (5-HIAA) were determined in four regions of postmortem brains of demented patients with or without Alzheimer's disease (AD). 2. NA was deficient in the temporal cortex (BA 21) of AD, but not of non-AD, patients. 3. Caudate, in particular, had an impaired dopaminergic system in AD patients, with low HVA levels. 4. In all regions investigated [amygdala, caudate, putamen, temporal cortex (BA 21)] 5-HT was significantly depleted in AD patients, and 5-HIAA was also depleted in amygdala and caudate. 5. These results indicate that neurotransmitter systems other than cholinergic systems are also widely affected in AD and suggest that these deficits may also play an important role in determining the symptomatology of AD.
Asunto(s)
Enfermedad de Alzheimer/metabolismo , Química Encefálica , Neurotransmisores/análisis , Anciano , Anciano de 80 o más Años , Enfermedad de Alzheimer/patología , Amígdala del Cerebelo/química , Núcleo Caudado/química , Demencia/metabolismo , Demencia/patología , Dopamina/análisis , Femenino , Ácido Homovanílico/análisis , Humanos , Ácido Hidroxiindolacético/análisis , Masculino , Persona de Mediana Edad , Neurotransmisores/metabolismo , Norepinefrina/análisis , Putamen/química , Serotonina/análisis , Lóbulo Temporal/químicaRESUMEN
The discovery of mutated, GTPase-deficient alpha subunits of Gs or Gi2 in certain human endocrine tumors has suggested that heterotrimeric G proteins play a role in the oncogenic process. Expression of these altered forms of G alpha s or G alpha i2 proteins in rodent fibroblasts activates or inhibits endogenous adenylyl cyclase, respectively, and causes certain alterations in cell growth. However, it is not clear whether growth abnormalities result from altered cyclic AMP synthesis. In the present study, we asked whether a recently discovered family of G proteins, Gq, which does not affect adenylyl cyclase activity, but instead mediates the activation of phosphatidylinositol-specific phospholipase C harbors transforming potential. We mutated the cDNA for the alpha subunit of murine Gq in codons corresponding to a region involved in binding and hydrolysis of GTP. Similar mutations unmask the transforming potential of p21ras or activate the alpha subunits of Gs or Gi2. Our results show that when expressed in NIH 3T3 cells, activating mutations convert G alpha q into a dominant acting oncogene.
Asunto(s)
Transformación Celular Neoplásica/genética , Proteínas de Unión al GTP/genética , Mutagénesis , Células 3T3 , Animales , Secuencia de Bases , División Celular/genética , ADN , Activación Enzimática , Proteínas de Unión al GTP/fisiología , Fosfatos de Inositol/metabolismo , Ratones , Ratones Desnudos , Datos de Secuencia Molecular , Trasplante de Neoplasias , Fosfatidilinositol Diacilglicerol-Liasa , Fosfoinositido Fosfolipasa C , Hidrolasas Diéster Fosfóricas/metabolismo , TransfecciónRESUMEN
[125I]Iodocyanopindolol binding sites were characterized by autoradiography in the superior cervical ganglia of Wistar-Kyoto (WKY) rats. A high concentration of (-)-[125I]iodocyanopindolol binding sites, characterized as beta-adrenoceptors by (-)-propranolol displacement, was distributed throughout the ganglia and in the postganglionic (internal carotid) nerve. ICI 118,551, a beta 2-selective antagonist, displaced more than 85% of the binding sites, whereas CGP 20712A, a beta 1-selective antagonist, displaced less than 10% of the binding sites, indicating that the beta-adrenoceptors were primarily of the beta 2-subtype. Emulsion autoradiography demonstrated that at least part of the binding sites were associated with principal ganglion cells. Unilateral deafferentation did not modify the number of binding sites in the superior cervical ganglia of WKY or spontaneously hypertensive rat (SHR). These results suggest that at least part of these receptors may correspond to prejunctional beta 2-adrenoceptors originated in principal ganglion cells. The concentration of beta 2-receptors was increased in the superior cervical ganglia of young and adult SHR when compared to age-matched WKY rats (49% and 39%, respectively). There were no differences in beta 2-adrenoceptor number in the stellate ganglia of young and adult WKY and SHR. These results suggest that beta 2-adrenoceptor stimulation may be selectively enhanced in some peripheral sympathetic ganglia in SHR and this could play a role in the development and maintenance of the increased sympathetic activity in this strain.
Asunto(s)
Ganglios Simpáticos/metabolismo , Hipertensión/metabolismo , Receptores Adrenérgicos beta/metabolismo , Antagonistas Adrenérgicos beta/metabolismo , Animales , Autorradiografía , Sitios de Unión , Unión Competitiva , Enalapril/farmacología , Imidazoles/metabolismo , Yodocianopindolol , Pindolol/análogos & derivados , Pindolol/metabolismo , Propanolaminas/metabolismo , Ratas , Ratas Endogámicas SHR , Ratas Endogámicas WKYRESUMEN
A frog brain kainic acid receptor (KAR) was studied using monoclonal and polyclonal antibodies against the affinity-purified receptor. Immunocytochemistry was done on sections of the frog CNS, and the distribution of immunostaining was compared with the distribution of high- and low-affinity 3H-kainic acid (3H-KA) binding sites determined with in vitro receptor autoradiography. These studies showed (1) similar distributions of high- and low-affinity 3H-KA binding sites, (2) identical patterns of immunostaining with the polyclonal antibodies and 2 monoclonal antibodies, and (3) an antibody binding distribution which closely matched that of 3H-KA binding, suggesting that the antibodies recognize the primary KAR in frog brain. In the frog brain, an anteroposterior gradient of immunostaining was observed, with the telencephalon intensely and uniformly immunoreactive. Other areas intensely immunoreactive included the cerebellum, the infundibulum, the tectal and posterior commissures, and the laminar nucleus of the torus semicircularis. The optic tectum showed selective staining of the plexiform layers 3 and 5-7. The pattern of staining was punctate and appeared to be associated with nerve fibers, among them dendritic arborizations. Electron microscopic observations showed staining at the cytoplasmic side of postsynaptic membranes. Extra-synaptic staining was observed as patches on the surface of unmyelinated nerve processes.
Asunto(s)
Anticuerpos Monoclonales , Sistema Nervioso Central/metabolismo , Rana pipiens/metabolismo , Receptores de Neurotransmisores/metabolismo , Animales , Sitios de Unión , Sistema Nervioso Central/ultraestructura , Inmunohistoquímica , Microscopía Electrónica , Receptores de Ácido Kaínico , Coloración y Etiquetado , Colículos Superiores/metabolismo , Distribución TisularRESUMEN
Angiotensin II (ANG II) receptor density was higher in many brain regions of untreated spontaneously hypertensive rats (SHR) compared to untreated Wistar-Kyoto (WKY) animals. Systemic inhibition of angiotensin converting enzyme with enalapril (25 mg/kg, per os for 14 days) produces a large decrease in ANG II receptors localized exclusively in the subfornical organ (SFO) of the SHR, and no alterations in ANG II receptors in the normotensive WKY rats. Selective decrease of ANG II receptors in the SFO of the genetically hypertensive rats with enalapril may be related to its therapeutic efficacy.
Asunto(s)
Encéfalo/fisiopatología , Enalapril/farmacología , Hipertensión/fisiopatología , Sistemas Neurosecretores/fisiopatología , Receptores de Angiotensina/efectos de los fármacos , Órgano Subfornical/fisiopatología , Sistema Vasomotor/fisiopatología , Angiotensina II/sangre , Angiotensina II/fisiología , Inhibidores de la Enzima Convertidora de Angiotensina/metabolismo , Animales , Encéfalo/efectos de los fármacos , Masculino , Ratas , Ratas Endogámicas SHR , Ratas Endogámicas WKY , Receptores de Angiotensina/fisiología , Renina/sangre , Sistema Renina-Angiotensina/efectos de los fármacos , Sistema Renina-Angiotensina/fisiología , Órgano Subfornical/efectos de los fármacos , Sistema Vasomotor/efectos de los fármacosRESUMEN
125I-Polymer standards were calibrated by interpolation of their optical densities in [125I]-brain paste standard curves to obtain dpm/mg protein. There was a linear relationship between the calibrated polymer standards and the dpm/mg polymer, as provided by the manufacturer. One dpm/mg polymer was equivalent to 7.34 +/- 0.22 dpm/mg protein. Receptor quantification in selected rat brain areas with comparison to either brain paste or calibrated polymer standards yielded similar results.
Asunto(s)
Autorradiografía/normas , Encéfalo/metabolismo , Radioisótopos de Yodo/normas , Polímeros/normas , Animales , RatasRESUMEN
In the superior cervical ganglia (SCG) binding site density of angiotensin II (ANG II) was higher in adult spontaneously hypertensive rats (SHR) (571 +/- 29 fmol/mg protein) compared to that in the adult Wistar-Kyoto rat (WKY) (375 +/- 9 fmol/mg protein, P less than 0.05). The ANG II binding density was significantly decreased in the SCG of SHR (-59%, P less than 0.01) and of WKY (-39%, P less than 0.05) after unilateral preganglionic denervation (operated v sham-operated ganglia). Part of the binding sites in the superior cervical ganglia may be present in or be associated to preganglionic nerves, and the number of these sites is higher in SHR.
Asunto(s)
Angiotensina II/metabolismo , Ganglios Simpáticos/anatomía & histología , Animales , Autorradiografía , Sitios de Unión , Desnervación , Ganglios Simpáticos/fisiología , Masculino , Ratas , Ratas Endogámicas SHR , Ratas Endogámicas WKY , Sistema Renina-AngiotensinaRESUMEN
We studied brain angiotensin II (ANG II) receptors by quantitative autoradiography in adult normotensive Wistar-Kyoto (WKY) rats and in spontaneously hypertensive rats (SHR) after treating the rats with the converting-enzyme inhibitor enalapril, 25 mg/kg, po daily for 14 days. Enalapril treatment decreased blood pressure in only SHR, inhibited plasma angiotensin-converting enzyme activity by 85%, and increased plasma ANG I concentration and renin activity in both WKY and SHR. In the untreated SHR animals, ANG II receptor concentrations were higher in the subfornical organ, the area postrema, the nucleus of the solitary tract, and the inferior olive when compared with the untreated WKY rats. Enalapril treatment produced a large decrease in only subfornical organ ANG II receptors of SHR. The selective reversal of the alteration in subfornical organ ANG II receptors in SHR may indicate a decreased central response to ANG II and may be related to the mode of action of angiotensin-converting enzyme inhibitors in this model.
Asunto(s)
Angiotensina II/metabolismo , Encéfalo/metabolismo , Enalapril/farmacología , Sistemas Neurosecretores/metabolismo , Receptores de Angiotensina/metabolismo , Órgano Subfornical/metabolismo , Acetilcolinesterasa/sangre , Angiotensina I/sangre , Animales , Presión Sanguínea/efectos de los fármacos , Masculino , Especificidad de Órganos , Ratas , Ratas Endogámicas SHR , Ratas Endogámicas WKY , Receptores de Angiotensina/efectos de los fármacos , Valores de Referencia , Renina/sangre , Sistema Renina-Angiotensina/efectos de los fármacos , Especificidad de la Especie , Órgano Subfornical/efectos de los fármacosRESUMEN
A regional analysis of brain amine and acid metabolite levels was conducted after chronic administration of the antidepressant tranylcypromine (0.5 mg/kg/day), of a novel fluorinated analog of this compound, and of clorgyline (1.0 mg/kg/day). These compounds were administered to male Sprague-Dawley rats for 28 days by subcutaneous infusion using osmotic minipumps (Alzet 2002). Levels of noradrenaline, 5-hydroxytryptamine, dopamine, and of the acid metabolites 5-hydroxyindole-3-acetic acid, 3,4-dihydroxyphenylacetic acid, and homovanillic acid were measured in the frontal cortex, nucleus accumbens, caudate nucleus, hippocampus, and hypothalamus. After 28 days of drug administration, sustained increases in amines and decreases in their acid metabolites were observed. Regional differences in these effects were minimal. These results are consistent with reports of sustained increases in brain amine concentrations following prolonged administration of other monoamine oxidase inhibitors.
Asunto(s)
Encéfalo/efectos de los fármacos , Clorgilina/farmacología , Neurotransmisores/metabolismo , Propilaminas/farmacología , Tranilcipromina/análogos & derivados , Tranilcipromina/farmacología , Ácido 3,4-Dihidroxifenilacético/metabolismo , Animales , Mapeo Encefálico , Dopamina/metabolismo , Ácido Homovanílico/metabolismo , Ácido Hidroxiindolacético/metabolismo , Masculino , Norepinefrina/metabolismo , Ratas , Ratas Endogámicas , Serotonina/metabolismoRESUMEN
High affinity binding sites for brain natriuretic peptide were characterized in the rat superior cervical ganglia by quantitative autoradiography. In addition, the peptide increased the formation of cyclic GMP in the ganglia in vitro. Brain natriuretic peptide displaced atrial natriuretic peptide from its binding sites. Our results suggest that brain natriuretic peptide and atrial natriuretic peptide may share physiologically active receptors in sympathetic ganglia. Brain natriuretic peptide may modulate the synaptic transmission in sympathetic ganglia, in addition or in conjunction with atrial natriuretic peptide.
Asunto(s)
Factor Natriurético Atrial/metabolismo , Encéfalo/metabolismo , Ganglios Simpáticos/metabolismo , Receptores de Superficie Celular/metabolismo , Animales , Autorradiografía , Unión Competitiva , Radioisótopos de Yodo , Cinética , Masculino , Ratas , Ratas Endogámicas , Receptores del Factor Natriurético AtrialRESUMEN
beta-Adrenoceptors were localized and characterized in valve leaflets of the rat heart. Sixteen micrometer-thick tissue sections containing the mitral and aortic valves were incubated with (-)3-[125I]iodocyanopindolol followed by autoradiography with computerized microdensitometry and comparison with 125I-labeled standards. beta-Adrenoceptors were present in all the valves studied. The selective beta 1-adrenoceptor antagonist CGP 20712 A (100 nM) displaced not more than 20% of the total binding sites, suggesting that most of the beta-adrenoceptors in the valve leaflets are of the beta 2-subtype. Forskolin-binding sites were detected in the mitral valve leaflet by incubation of adjacent tissue sections with [12-3H]forskolin. Our results indicate that catecholamines could regulate the function of the heart valves through stimulation of beta 2-adrenoceptors.