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1.
Acta Histochem ; 121(1): 7-15, 2019 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-30322809

RESUMO

Nervous control of light emission from deep-sea mesopelagic fishes has been documented for several species. Studies on the nervous control of photophores from deep-sea luminescent fish, are mainly restricted to a pharmacological approach. For example, the light organs, called photophores, isolated from Argyropelecus hemygimnus and Maurolicus muelleri show a much higher sensitivity to adrenaline than to noradrenaline. According to these results and other information in different species, catecholamines are considered as main neurotransmitters triggering bioluminescence in deep-sea fishes. The present work is a study of the nervous control of the isolated photophores from two Stomiid fishes, Chauliodus sloani (the viperfish) and Stomias boa (the dragonfish) with the aim to determine the nature of the nervous control by pharmacological, biochemical and morphological approaches. Results show that, although the photophores of both species are sensitive to catecholamines, adrenaline is present in larger amount than noradrenaline in the light organs of C. sloani. Both catecholamines have different immunoreactive (IR) sites, noradrenaline showing a very diffuse localization as compared to adrenaline in C. sloani. On the contrary, only adrenaline is detected in the photocytes chamber and nerves innervating the photophore in S. boa. Knowing that the majority of dragonfishes exhibit a luminescent chin barbel, we also investigated the presence of catecholamines in this specific tissue in S. boa. Immunohistology reveals the presence of adrenaline within the tissue forming the chin barbel; adrenaline-IR is found in the connective tissue surroundings two group of muscle fibers and blood vessels in the stem but also around the multiple blood vessels located within the barbel bulb. Our results strongly support the adrenergic control of light emission in bioluminescent stomiid fishes.


Assuntos
Catecolaminas/farmacocinética , Luminescência , Neurotransmissores/farmacologia , Animais , Cromatografia Líquida de Alta Pressão , Peixes , Imuno-Histoquímica
2.
J Microbiol Biotechnol ; 27(11): 1907-1915, 2017 Nov 28.
Artigo em Inglês | MEDLINE | ID: mdl-29032653

RESUMO

Lipases are important enzymes with biotechnological applications in dairy, detergent, food, fine chemicals, and pharmaceutical industries. Specifically, hormone-sensitive lipase (HSL) is an intracellular lipase that can be stimulated by several hormones, such as catecholamine, glucagon, and adrenocorticotropic hormone. Bacterial hormone-sensitive lipases (bHSLs), which are homologous to the C-terminal domain of HSL, have α/ß-hydrolase fold with a catalytic triad composed of His, Asp, and Ser. These bHSLs could be used for a wide variety of industrial applications because of their high activity, broad substrate specificity, and remarkable stability. In this review, the relationships among HSLs, the microbiological origins, the crystal structures, and the biotechnological properties of bHSLs are summarized.


Assuntos
Bactérias/enzimologia , Biotecnologia , Microbiologia Industrial , Esterol Esterase/química , Hormônio Adrenocorticotrópico/metabolismo , Sequência de Aminoácidos , Proteínas de Bactérias/química , Domínio Catalítico , Catecolaminas/farmacocinética , Estabilidade Enzimática , Glucagon/metabolismo , Modelos Moleculares , Conformação Proteica , Especificidade por Substrato
3.
Circ Cardiovasc Imaging ; 8(8): e003507, 2015 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-26245765

RESUMO

BACKGROUND: Post-transplant reinnervation is a unique model to study sympathetic neuronal regeneration in vivo. The differential role of subcellular mechanisms of catecholamine handling in nerve terminals has not been investigated. METHODS AND RESULTS: Three different carbon-11-labeled catecholamines were used for positron emission tomography of transport (C-11 m-hydroxyephedrine, HED), vesicular storage (C-11 epinephrine, EPI), and metabolic degradation (C-11 phenylephrine). A 2-day protocol was used, including quantification of myocardial blood flow by N-13 ammonia. Resting myocardial blood flow and EPI, HED and phenylephrine retention were homogeneous in healthy volunteers (n=7). Washout was only observed for phenylephrine (T(1/2) 49±6 min). In nonrejecting, otherwise healthy heart transplant recipients (>1 year after surgery, n=10), resting myocardial blood flow was also homogenous. Regional catecholamine uptake of varying degrees was observed in the anterior left ventricular wall and septum. Overall, 24±19% of left ventricle showed HED uptake levels comparable with healthy volunteers, whereas it was only 8±7% for EPI (P=0.004 versus HED). Phenylephrine washout was not different from healthy volunteers in the area with restored EPI and HED retention (T(1/2) 41±7 min; P>0.05), but was significantly enhanced in the EPI/HED mismatch area (T(1/2) 36±8 min; P=0.008), consistent with inefficient vesicular storage and enhanced metabolic degradation. CONCLUSIONS: Regeneration of subcellular components of sympathetic nerve terminal function does not occur simultaneously. In the reinnervating transplanted heart, a region with normal catecholamine transport and vesicular storage is surrounded by a borderzone, where transport is already restored but vesicular storage remains inefficient, suggesting that vesicular storage is a more delicate mechanism. This observation may have implications for other pathologies involving cardiac autonomic innervation.


Assuntos
Catecolaminas , Transplante de Coração , Coração/inervação , Imagem Molecular/métodos , Regeneração Nervosa , Tomografia por Emissão de Pósitrons , Compostos Radiofarmacêuticos , Sistema Nervoso Simpático/diagnóstico por imagem , Adulto , Estudos de Casos e Controles , Catecolaminas/farmacocinética , Efedrina/análogos & derivados , Epinefrina/análogos & derivados , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Imagem de Perfusão do Miocárdio , Fenilefrina , Valor Preditivo dos Testes , Terminações Pré-Sinápticas/diagnóstico por imagem , Terminações Pré-Sinápticas/metabolismo , Compostos Radiofarmacêuticos/farmacocinética , Sistema Nervoso Simpático/metabolismo , Sistema Nervoso Simpático/fisiopatologia , Resultado do Tratamento
6.
An. R. Acad. Farm ; 80(2): 347-361, abr.-jun. 2014. ilus, graf, tab
Artigo em Inglês | IBECS | ID: ibc-125902

RESUMO

Hormones are expressed during development in unexpected locations and stages, and this fact relates to their distinct functional roles in the embryo. In recent work, we found that the expression of Tyrosine Hydroxylase (TH, first enzyme of the catecholamine synthetic pathway) and the presence of catecholamines, antecede neural innervation in some tissues. We focus this overview on the vertebrate developing heart. TH transcripts were present in early cardiogenesis, and adrenergic as well as dopaminergic receptors were found in the cardiac region of chick embryos. We found direct effects of dopamine on cardiac gene expression and we have advanced in revealing the function of catecholamines on cardiac patterning


Las hormonas están expresadas durante el desarrollo en etapas y localizaciones inesperadas y este hecho se relaciona con sus distintas funciones en el embrión. Recientemente, hemos encontrado que la expresión de la Tirosina Hidroxilasa (TH, el primer enzima de la ruta de síntesis de catecolaminas) y la presencia de catecolaminas, anteceden a la inervación neural en algunos tejidos. Este artículo está centrado en el desarrollo del corazón de vertebrados. Los transcritos de TH se expresan durante la cardiogénesis temprana y se encontraron receptores dopaminérgicos y adrenérgicos en la región cardiaca del embrión de pollo. Hemos demostrado efectos directos de la dopamina sobre la expresión de genes cardiacos y hemos avanzado en caracterizar una función de las catecolaminas sobre la formación del patrón del corazón


Assuntos
Animais , Catecolaminas/farmacocinética , Coração/crescimento & desenvolvimento , Dopamina/farmacocinética , Tirosina 3-Mono-Oxigenase/análise , Vertebrados/crescimento & desenvolvimento , Hormônios/farmacocinética , Neurotransmissores/farmacocinética , Elementos Reguladores de Transcrição
7.
Actual. anestesiol. reanim ; 21(2): 35-44, abr.-jun. 2011. tab
Artigo em Espanhol | IBECS | ID: ibc-97923

RESUMO

La agresión quirúrgica provoca una respuesta endocrinometabólica, siendo el dolor postoperatorio uno de los factores implicados en la producción de dicha respuesta, consecuencia de la activación del sistema simpático y estimulación del eje endocrino hipotálamo-hipofisario-adrenal (HHA). El anestesiólogo puede modificar estas respuestas endocrinas y metabólicas al utilizar los distintos fármacos o técnicas anestésicas durante el acto quirúrgico. En un intento de frenar los efectos de la secreción de catecolaminas, debida al estrés quirúrgico, se han utilizado fármacos como los bloqueantes beta adrenérgicos, con el fin de evitar complicaciones como la isquemia miocárdica perioperatoria. Los resultados han demostrado que los betabloqueantes no disminuyen la respuesta neuroendocrina al estrés, pero sí disminuyen los requerimientos analgésicos, se produce una recuperación de la anestesia más rápida y una mejoría en la estabilidad hemodinámica. En el presente trabajo se hace una revisión sobre los aspectos clínicos derivados de la respuesta neuroendocrina, metabólica, inmunológica e inflamatoria a la agresión quirúrgica(AU)


The surgical aggression provokes an endocrinometabolic response, with the postoperatory pain beeing one of the factors involved in the production of the above mentioned response, consequence of the activation of the simpathetic nervous system and stimulation of the endocrine hypothalamus - hipofisario-adrenal axis (HHA). The anaesthtetist can modify these endocrine and metabolic responses on by using the different drugs or anesthesic techniques during the surgical operation. In an attempt to the limit the effects of the catecolamines secretion due to the surgical stress, the medicaments such as adrenergic betablockers have been iused, in order to avoid complications such perioperatory myocardic ischemia. The results have demonstrated that betablockers do not reduce the neuroendocrine response to the stress, but do reduce the analgesic requirements, tand a faster recovery from the anesthesia is produced, and an improvement in the hemodinámic stability In the present work a review is made on the clinical aspects derived from the neuroendocrine, metabolic, immunological and inflammatory response to the surgical aggression(AU)


Assuntos
Humanos , Masculino , Feminino , Estresse Fisiológico , Transtornos de Estresse Pós-Traumáticos/tratamento farmacológico , Estresse Psicológico/tratamento farmacológico , Dor Pós-Operatória/tratamento farmacológico , Antagonistas Adrenérgicos beta/uso terapêutico , Hemodinâmica , Dor Pós-Operatória/metabolismo , Dor Pós-Operatória/fisiopatologia , Período Pós-Operatório , Glândulas Endócrinas , Catecolaminas/farmacologia , Catecolaminas/farmacocinética , Catecolaminas/uso terapêutico , Receptores de Catecolaminas , Antagonistas Adrenérgicos beta/metabolismo , Antagonistas Adrenérgicos beta/farmacocinética
8.
J Comp Physiol B ; 178(5): 629-36, 2008 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-18274760

RESUMO

Stress assessment favours methods, which do not interfere with an animal's endocrine status. To develop such non-invasive methods, detailed knowledge about the excretion of hormone metabolites in the faeces and urine is necessary. Our study was therefore designed to generate basic information about catecholamine excretion in rats, mice and chickens. After administration of (3)H-epinephrine or (3)H-norepinephrine to male and female rats, mice and chickens, all voided excreta were collected for 4 weeks, 3 weeks or for 10 days, respectively. Peak concentrations of radioactivity appeared in one of the first urinary samples of mice and rats and in the first droppings in chickens 0.2-7.2 h after injection. In rats, between 77.3 and 95.6% of the recovered catecholamine metabolites were found in the urine, while in mice, a mean of 76.3% were excreted in the urine. Peak concentrations in the faeces were found 7.4 h post injection in mice, and after about 16.4 h in rats (means). Our study provides valuable data about the route and the profile of catecholamine excretion in three frequently used species of laboratory animals. This represents the first step in the development of a reliable, non-invasive quantification of epinephrine and norepinephrine to monitor sympatho-adrenomedullary activity, although promising results for the development of a non-invasive method were found only for the chicken.


Assuntos
Catecolaminas/urina , Fezes/química , Animais , Catecolaminas/metabolismo , Catecolaminas/farmacocinética , Galinhas , Epinefrina/metabolismo , Epinefrina/farmacocinética , Epinefrina/urina , Feminino , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Norepinefrina/metabolismo , Norepinefrina/farmacocinética , Norepinefrina/urina , Ratos , Ratos Endogâmicos F344 , Fatores Sexuais , Especificidade da Espécie , Trítio
9.
J Pharmacol Exp Ther ; 324(2): 685-93, 2008 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-17962518

RESUMO

In a previous study performed in the intact adrenal gland (Lim et al., 2002), stimulation with acetylcholine (ACh) or high K(+) concentrations (K(+)) produced greater catecholamine release in spontaneously hypertensive rats (SHR), as compared with normotensive animals. In this study, the time course of secretion was in the range of minutes. Hence, we do not know whether enhanced release is due to greater quantal content and/or distinct kinetics in SHRs and control animals. To get insight into the mechanism involved in such enhanced catecholamine secretory responses, we performed a single-vesicle release study in primary cultures of adrenal chromaffin cells, recorded with amperometry. Cells were stimulated with 2-s pulses of 1 mM ACh or 70 mM K(+). The secretory responses to ACh or K(+) pulses in SHR cells as compared with control cells had the following characteristics: 1) double number of secretory events, 2) 4-fold augmentation of total secretion, 3) cumulative secretion that saturated slowly, 4) 3-fold higher complex events with two to four superimposed spikes that may be explained by faster spike kinetics, 5) about 2- to 3-fold higher event frequency at earlier post stimulation periods, and 6) 2- to 5-fold higher quantal content of simple spikes. We conclude that SHR cells have faster and larger catecholamine release responses, explained by more vesicles ready to undergo exocytosis and greater quantal content of vesicles. This could have relevance to further understand the pathogenic mechanisms involved in the development of high blood pressure, as well as in the identification of new drug targets to treat hypertension.


Assuntos
Catecolaminas/metabolismo , Catecolaminas/farmacocinética , Células Cromafins/metabolismo , Hipertensão/metabolismo , Vesículas Secretórias/metabolismo , Potenciais de Ação/fisiologia , Animais , Pressão Sanguínea/fisiologia , Exocitose/fisiologia , Masculino , Ratos , Ratos Endogâmicos SHR , Ratos Sprague-Dawley
11.
Neurotox Res ; 11(3-4): 261-71, 2007 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-17449463

RESUMO

Although it is well-recognized that catecholamines are generally unable to penetrate the developed blood-brain barrier (BBB) to gain entry into brain, except at circumventricular sites where the BBB is absent or deficient, ontogenetic development of this barrier seems to have escaped systematic study. To explore BBB development, several approaches were used. In the first study rats were treated once on a specific day of postnatal ontogeny, as early as the day of birth, with the neurotoxin 6-hydroxydopamine (6-OHDA; 60 mg/kg), and then terminated in adulthood for regional analysis of endogenous norepinephrine (NE) content of brain. In another study, rats were treated once, on a specific day of postnatal ontogeny, with the BBB-permeable neurotoxin 6-hydroxydopa (6-OHDOPA; 60 mg/kg) following pretreatment with the BBB-impermeable amino acid decarboxylase inhibitor carbidopa (100 mg/kg IP), then terminated in adulthood for regional analysis of endogenous NE content of brain. In the third study rats were treated once, on a specific day of postnatal ontogeny, with the analog [3H]metaraminol, and terminated 1 hour later for determination of regional distribution of tritium in brain. On the basis of [3H]metaraminol distribution and NE depletions after neurotoxin treatments, it is evident that the BBB in neocortex, striatum, cerebellum and other brain regions forms in stages over a period of at least 2 weeks from birth. Moreover, because the BBB consists of several element (physical-, ion-restrictive-, and enzymatic-barrier), the method employed will derive data mainly applicable to the targeted aspect of the barrier, which may or may not necessarily coincide with elements of the barrier that have a different rate of ontogenetic development. Accordingly, it is evident that some aspects of physical- and ion-restrictive elements of the BBB form within approximately the first week after birth in rat neocortex and striatum, while enzymatic elements of the BBB form more than than 2 weeks later. Regardless, the BBB forms at earlier times in forebrain vs hindbrain regions.


Assuntos
Barreira Hematoencefálica/metabolismo , Química Encefálica/efeitos dos fármacos , Catecolaminas/farmacocinética , Adrenérgicos/farmacologia , Fatores Etários , Animais , Animais Recém-Nascidos , Barreira Hematoencefálica/efeitos dos fármacos , Encéfalo/anatomia & histologia , Encéfalo/efeitos dos fármacos , Encéfalo/crescimento & desenvolvimento , Encéfalo/metabolismo , Carbidopa/farmacologia , Dopaminérgicos/farmacologia , Interações Medicamentosas , Feminino , Masculino , Metaraminol/farmacocinética , Oxidopamina/farmacologia , Ratos , Ratos Sprague-Dawley
12.
Best Pract Res Clin Endocrinol Metab ; 20(3): 467-82, 2006 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-16980206

RESUMO

Cushing's syndrome can be exogenous, resulting from the administration of glucocorticoids or adrenocorticotrophic hormone (ACTH), or endogenous, secondary to increased secretion of cortisol or ACTH. Hypertension is one of the most distinguishing features of endogenous Cushing's syndrome, as it is present in about 80% of adult patients and in almost half of children and adolescents patients. Hypertension results from the interplay of several pathophysiological mechanisms regulating plasma volume, peripheral vascular resistance and cardiac output, all of which may be increased. The therapeutic goal is to find and remove the cause of excess glucocorticoids, which, in most cases of endogenous Cushing's syndrome, is achieved surgically. Treatment of Cushing's syndrome usually results in resolution or amelioration of hypertension. However, some patients may not achieve normotension or may require a prolonged period of time for the correction of hypercortisolism. Therefore, therapeutic strategies for Cushing's-specific hypertension (to normalise blood pressure and decrease the duration of hypertension) are necessary to decrease the morbidity and mortality associated with this disorder. The various pathogenetic mechanisms that have been proposed for the development of glucocorticoid-induced hypertension in Cushing's syndrome and its management are discussed.


Assuntos
Síndrome de Cushing/complicações , Hipertensão/complicações , 11-beta-Hidroxiesteroide Desidrogenases/fisiologia , Angiotensina II/fisiologia , Anti-Hipertensivos/uso terapêutico , Aterosclerose/complicações , Catecolaminas/farmacocinética , Síndrome de Cushing/epidemiologia , Síndrome de Cushing/cirurgia , Glucocorticoides/farmacologia , Humanos , Hipertensão/tratamento farmacológico , Hipertensão/epidemiologia , Hipertensão/etiologia , Modelos Biológicos , Sistema Renina-Angiotensina/efeitos dos fármacos , Sistema Renina-Angiotensina/fisiologia , Vasoconstritores/farmacologia , Vasodilatadores/farmacologia
13.
Respiration ; 72(2): 189-96, 2005.
Artigo em Inglês | MEDLINE | ID: mdl-15824530

RESUMO

BACKGROUND: Although aerosolized beta-adrenergic agonists have been used as a therapy for the resolution of pulmonary edema, the mechanisms of catecholamine clearance from the alveolar spaces of the lung are not well known. OBJECTIVE: To determine whether catecholamine clearance from the alveolar spaces is correlated with the fluid transport capacity of the lung. METHODS: Albumin solution containing epinephrine (10(-7)M) or norepinephrine (10(-7)M) was instilled into the alveolar spaces of isolated rat and human lungs. Alveolar fluid clearance rate was estimated by the progressive increase in the albumin concentration over 1 h. Catecholamine clearance rate was estimated by the changes in catecholamine concentration and alveolar fluid volume over 1 h. RESULTS: The norepinephrine clearance rate was faster than the epinephrine clearance rate in the rat and human lungs. In the rat lungs, amiloride (a sodium channel blocker) caused a greater decrease in alveolar fluid clearance and epinephrine clearance rate than propranolol (a nonselective beta-adrenergic antagonist). Although propranolol and phentolamine (an alpha-adrenergic antagonist), and 5-(N-ethyl-N-isoprophyl)amiloride (a Na+/H+ antiport blocker) changed neither the alveolar fluid clearance nor the norepinephrine clearance rate, amiloride and benzamil (a sodium channel blocker) decreased both clearance rates. As in the rat lungs, amiloride decreased alveolar fluid and norepinephrine clearance rates in the human lungs. CONCLUSION: These results indicate that the catecholamine clearance rate from the alveolar spaces is correlated with alveolar fluid clearance in rat and human lungs.


Assuntos
Amilorida/análogos & derivados , Catecolaminas/farmacocinética , Alvéolos Pulmonares/metabolismo , Agonistas alfa-Adrenérgicos/administração & dosagem , Agonistas alfa-Adrenérgicos/farmacocinética , Antagonistas Adrenérgicos alfa/administração & dosagem , Antagonistas Adrenérgicos alfa/farmacocinética , Agonistas Adrenérgicos beta/administração & dosagem , Agonistas Adrenérgicos beta/farmacocinética , Amilorida/administração & dosagem , Amilorida/farmacocinética , Animais , Catecolaminas/administração & dosagem , Epinefrina/administração & dosagem , Epinefrina/farmacocinética , Líquido Extracelular/metabolismo , Humanos , Técnicas In Vitro , Instilação de Medicamentos , Masculino , Norepinefrina/administração & dosagem , Norepinefrina/farmacocinética , Fentolamina/administração & dosagem , Fentolamina/farmacocinética , Propranolol/administração & dosagem , Propranolol/farmacocinética , Alvéolos Pulmonares/citologia , Ratos , Ratos Sprague-Dawley , Bloqueadores dos Canais de Sódio/administração & dosagem , Bloqueadores dos Canais de Sódio/farmacocinética , Trocadores de Sódio-Hidrogênio/antagonistas & inibidores
14.
Pharmacol Rev ; 56(3): 331-49, 2004 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-15317907

RESUMO

This article provides an update about catecholamine metabolism, with emphasis on correcting common misconceptions relevant to catecholamine systems in health and disease. Importantly, most metabolism of catecholamines takes place within the same cells where the amines are synthesized. This mainly occurs secondary to leakage of catecholamines from vesicular stores into the cytoplasm. These stores exist in a highly dynamic equilibrium, with passive outward leakage counterbalanced by inward active transport controlled by vesicular monoamine transporters. In catecholaminergic neurons, the presence of monoamine oxidase leads to formation of reactive catecholaldehydes. Production of these toxic aldehydes depends on the dynamics of vesicular-axoplasmic monoamine exchange and enzyme-catalyzed conversion to nontoxic acids or alcohols. In sympathetic nerves, the aldehyde produced from norepinephrine is converted to 3,4-dihydroxyphenylglycol, not 3,4-dihydroxymandelic acid. Subsequent extraneuronal O-methylation consequently leads to production of 3-methoxy-4-hydroxyphenylglycol, not vanillylmandelic acid. Vanillylmandelic acid is instead formed in the liver by oxidation of 3-methoxy-4-hydroxyphenylglycol catalyzed by alcohol and aldehyde dehydrogenases. Compared to intraneuronal deamination, extraneuronal O-methylation of norepinephrine and epinephrine to metanephrines represent minor pathways of metabolism. The single largest source of metanephrines is the adrenal medulla. Similarly, pheochromocytoma tumor cells produce large amounts of metanephrines from catecholamines leaking from stores. Thus, these metabolites are particularly useful for detecting pheochromocytomas. The large contribution of intraneuronal deamination to catecholamine turnover, and dependence of this on the vesicular-axoplasmic monoamine exchange process, helps explain how synthesis, release, metabolism, turnover, and stores of catecholamines are regulated in a coordinated fashion during stress and in disease states.


Assuntos
Catecolaminas/metabolismo , Animais , Transporte Biológico Ativo/fisiologia , Catecolaminas/química , Catecolaminas/farmacocinética , Humanos , Modelos Biológicos , Estrutura Molecular
15.
Pol J Pharmacol ; 55(4): 573-9, 2003.
Artigo em Inglês | MEDLINE | ID: mdl-14581716

RESUMO

We report that the classical guanylate cyclase inhibitor methylene blue (MB, 1 microM or 10 microM), but not the selective guanylate cyclase inhibitor 1H-[1,2,4]oxidazolo[4,3-a]quinoxalin-1-one (1 microM) or nitric oxide synthase inhibitor, NG-nitro-L-arginine methyl ester (100 microM), causes a shift to the left in the concentration-response curve for noradrenaline in the isolated rat vas deferens preparations. The main objective of our study was to investigate the pharmacological mechanism by which MB increases the sensitivity of the rat vas deferens to noradrenaline. According to the presented results, MB did not change rat vas deferens sensitivity to methoxamine or noradrenaline in the presence of desipramine (0.1 microM). The pre-contracted rat vas deferens relaxation induced by isoproterenol was also not significantly changed by MB (1 microM). Thus, we suggest that MB increases rat vas deferens sensitivity through neuronal uptake inhibition without interfering in either the nitrergic mechanism or guanylate cyclase activity.


Assuntos
Azul de Metileno/farmacologia , Neurônios/metabolismo , Norepinefrina/farmacocinética , Ducto Deferente/efeitos dos fármacos , Animais , Compostos de Bário/farmacologia , Catecolaminas/farmacocinética , Cloretos/farmacologia , Relação Dose-Resposta a Droga , Inibidores Enzimáticos/farmacologia , Guanilato Ciclase/antagonistas & inibidores , Guanilato Ciclase/metabolismo , Técnicas In Vitro , Isoproterenol/farmacologia , Masculino , Metoxamina/farmacocinética , Contração Muscular/efeitos dos fármacos , Relaxamento Muscular/efeitos dos fármacos , NG-Nitroarginina Metil Éster/farmacologia , Óxido Nítrico Sintase/antagonistas & inibidores , Óxido Nítrico Sintase/metabolismo , Oxidiazóis/farmacologia , Quinoxalinas/farmacologia , Ratos , Ducto Deferente/inervação , Ducto Deferente/fisiologia
16.
J Neurosci ; 23(5): 1697-709, 2003 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-12629174

RESUMO

Norepinephrine (NE) transporters (NETs) terminate noradrenergic synaptic transmission and represent a major therapeutic target for antidepressant medications. NETs and related transporters are under intrinsic regulation by receptor and kinase-linked pathways, and clarification of these pathways may suggest candidates for the development of novel therapeutic approaches. Syntaxin 1A, a presynaptic soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) protein, interacts with NET and modulates NET intrinsic activity. NETs colocalize with and bind to syntaxin 1A in both native preparations and heterologous systems. Protein kinase C activation disrupts surface NET/syntaxin 1A interactions and downregulates NET activity in a syntaxin-dependent manner. Syntaxin 1A binds the NH(2) terminal domain of NET, and a deletion of this domain both eliminates NET/syntaxin 1A associations and prevents phorbol ester-triggered NET downregulation. Whereas syntaxin 1A supports the surface trafficking of NET proteins, its direct interaction with NET limits transporter catalytic function. These two contradictory roles of syntaxin 1A on NET appear to be linked and reveal a dynamic cycle of interactions that allow for the coordinated control between NE release and reuptake.


Assuntos
Antígenos de Superfície/metabolismo , Catecolaminas/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Simportadores/metabolismo , Proteínas de Transporte Vesicular , Animais , Antidepressivos/farmacologia , Antígenos de Superfície/genética , Toxinas Botulínicas/farmacologia , Química Encefálica , Catecolaminas/farmacocinética , Células Cultivadas , Ativação Enzimática/efeitos dos fármacos , Ativação Enzimática/fisiologia , Ativadores de Enzimas/farmacologia , Inibidores Enzimáticos/farmacologia , Humanos , Masculino , Proteínas de Membrana/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Proteínas do Tecido Nervoso/antagonistas & inibidores , Proteínas do Tecido Nervoso/genética , Neurônios/citologia , Neurônios/efeitos dos fármacos , Neurônios/metabolismo , Norepinefrina/metabolismo , Norepinefrina/farmacocinética , Proteínas da Membrana Plasmática de Transporte de Norepinefrina , Oligonucleotídeos Antissenso/farmacologia , Técnicas de Patch-Clamp , Ligação Proteica/fisiologia , Estrutura Terciária de Proteína/fisiologia , Transporte Proteico/fisiologia , Ratos , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/metabolismo , Proteínas SNARE , Deleção de Sequência , Simportadores/efeitos dos fármacos , Simportadores/genética , Sinaptossomos/química , Sinaptossomos/metabolismo , Sintaxina 1 , Ducto Deferente/química , Ducto Deferente/metabolismo
17.
J Biol Chem ; 277(10): 7816-23, 2002 Mar 08.
Artigo em Inglês | MEDLINE | ID: mdl-11748228

RESUMO

Previously we observed that rab3 GTPases modulate both the secretion of catecholamines from PC12 neuroendocrine cells and the steady-state accumulation of exogenous norepinephrine (NE) into these cells (Weber, E., Jilling, T., and Kirk, K. L. (1996) J. Biol. Chem. 271, 6963-6971). Here we addressed the mechanisms by which these monomeric GTPases stimulate NE uptake by PC12 cells including their effects on uptake kinetics, their sites of action (secretory granule membrane versus plasma membrane), and the involvement of rab3-interacting proteins in this process. We observed that rab3B stimulated the rate and maximal accumulation of radiolabeled NE into large dense core vesicles within intact PC12 cells. rab3A and rab3B also increased NE uptake into large dense core vesicles in digitonin-permeabilized PC12 cells, which indicates that these GTPases stimulate catecholamine uptake at the level of the secretory granule membrane. In an attempt to identify rab3B targets that may mediate this effect on NE uptake, we found that rab3B interacts directly with phosphoinositide 3-kinase (PI3K) in a GTP-dependent fashion and that PI3K activity was elevated in PC12 cells overexpressing rab3B. Furthermore, two structurally distinct inhibitors of PI3K (wortmannin and LY294002) inhibited NE uptake in intact as well as digitonin-permeabilized PC12 cells, but had no effect on calcium-evoked NE secretion. Our results indicate that rab3 and PI3K positively and coordinately regulate NE uptake in PC12 neuroendocrine cells at least in part by stimulating the secretory vesicle uptake step.


Assuntos
Catecolaminas/farmacocinética , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas rab3 de Ligação ao GTP/metabolismo , Androstadienos/farmacologia , Animais , Western Blotting , Centrifugação com Gradiente de Concentração , Cromonas/farmacologia , Relação Dose-Resposta a Droga , Inibidores Enzimáticos/farmacologia , Glutationa Transferase/metabolismo , Cinética , Morfolinas/farmacologia , Células PC12 , Testes de Precipitina , Ligação Proteica , Transporte Proteico , Ratos , Proteínas Recombinantes de Fusão/metabolismo , Fatores de Tempo , Wortmanina , Proteína rab3A de Ligação ao GTP/metabolismo
18.
Circulation ; 103(14): 1881-6, 2001 Apr 10.
Artigo em Inglês | MEDLINE | ID: mdl-11294807

RESUMO

BACKGROUND: The lack of cardiac catecholamine uptake and storage caused by sympathetic denervation may influence performance of the transplanted heart. Reinnervation, occurring late after transplantation, may partially resolve these effects. In this study, oxidative metabolism and its relation to cardiac work were compared in allografts and normal and failing hearts, and the effects of sympathetic reinnervation were evaluated. METHODS AND RESULTS: Twenty-seven nonrejecting, symptom-free transplant recipients, 11 healthy control subjects, and 10 patients with severe dilated cardiomyopathy underwent PET with (11)C acetate for assessment of oxidative metabolism by the clearance constant k(mono) and radionuclide angiography or MRI for measurement of ventricular function, geometry, and work. Efficiency was estimated noninvasively by a work-metabolic index [WMI=(stroke volumexheart ratexsystolic pressure)/k(mono)]. In 14 of 27 transplants, presence of regional reinnervation was identified with PET and the catecholamine analogue (11)C hydroxyephedrine (extent, 24+/-14% of left ventricle). The WMI was comparable in normal subjects and reinnervated and denervated transplants (6.2+/-2.3 versus 4.9+/-2.0 versus 4.9+/-1.2. 10(6) mm Hg. mL; P=NS) and significantly lower in cardiomyopathy patients (3.0+/-1.3. 10(6) mm Hg. mL; P<0.001). For normal subjects and transplant recipients, the WMI was significantly correlated with afterload (peripheral vascular resistance; r=-0.65, P<0.01), preload (end-diastolic volume; r=0.78, P<0.01), and stroke volume (r=0.81, P<0.01) but not with hydroxyephedrine retention (transplants only; r=0.09, P=NS). CONCLUSIONS: After transplantation, cardiac efficiency is improved compared with failing hearts and comparable to normal hearts. Differences between denervated and reinnervated allografts were not surveyed. Additionally, the dependency on loading conditions and contractility was preserved, suggesting that normal regulatory interactions for efficiency are intact and that sympathetic tone does not play a role under resting conditions.


Assuntos
Transplante de Coração , Coração/fisiologia , Sistema Nervoso Simpático/fisiologia , Idoso , Cardiomiopatias/metabolismo , Cardiomiopatias/fisiopatologia , Catecolaminas/farmacocinética , Feminino , Coração/inervação , Coração/fisiopatologia , Ventrículos do Coração/fisiopatologia , Hemodinâmica , Humanos , Masculino , Pessoa de Meia-Idade , Contração Miocárdica , Miocárdio/metabolismo , Volume Sistólico , Sistema Nervoso Simpático/fisiopatologia , Tomografia Computadorizada de Emissão , Função Ventricular , Função Ventricular Esquerda/fisiologia
19.
Mol Pharmacol ; 59(1): 83-95, 2001 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-11125028

RESUMO

Extracellular concentrations of monoamine neurotransmitters are regulated by a family of high-affinity transporters that are the molecular targets for such psychoactive drugs as cocaine, amphetamines, and therapeutic antidepressants. In Drosophila melanogaster, cocaine-induced behaviors show striking similarities to those induced in vertebrate animal models. Although a cocaine-sensitive serotonin carrier exists in flies, there has been no pharmacological or molecular evidence to support the presence of distinct carrier subtypes for other bioactive monoamines. Here we report the cloning and characterization of a cocaine-sensitive fly dopamine transporter (dDAT). In situ hybridization demonstrates that dDAT mRNA expression is restricted to dopaminergic cells in the fly nervous system. The substrate selectivity of dDAT parallels that of the mammalian DATs in that dopamine and tyramine are the preferred substrates, whereas octopamine is transported less efficiently, and serotonin not at all. In contrast, dDAT inhibitors display a rank order of potency most closely resembling that of mammalian norepinephrine transporters. Cocaine has a moderately high affinity to the cloned dDAT (IC50 = 2.6 microM). Voltage-clamp analysis of dDAT expressed in Xenopus laevis oocytes indicates that dDAT-mediated uptake is electrogenic; however, dDAT seems to lack the constitutive leak conductance that is characteristic of the mammalian catecholamine transporters. The combination of a DAT-like substrate selectivity and norepinephrine transporter-like inhibitor pharmacology within a single carrier, and results from phylogenetic analyses, suggest that dDAT represents an ancestral catecholamine transporter gene. The identification of a cocaine-sensitive target linked to dopaminergic neurotransmission in D. melanogaster will serve as a basis for further dissection of the genetic components of psychostimulant-mediated behavior.


Assuntos
Antidepressivos/farmacologia , Proteínas de Transporte/metabolismo , Catecolaminas/farmacocinética , Glicoproteínas de Membrana , Proteínas de Membrana Transportadoras , Simportadores , Sequência de Aminoácidos , Animais , Transporte Biológico , Proteínas de Transporte/antagonistas & inibidores , Proteínas de Transporte/efeitos dos fármacos , Proteínas de Transporte/genética , Proteínas da Membrana Plasmática de Transporte de Catecolaminas , Cocaína/farmacologia , DNA Complementar/isolamento & purificação , Proteínas da Membrana Plasmática de Transporte de Dopamina , Proteínas de Drosophila , Drosophila melanogaster/efeitos dos fármacos , Drosophila melanogaster/genética , Drosophila melanogaster/metabolismo , Condutividade Elétrica , Eletrofisiologia , Expressão Gênica , Genoma , Taxa de Depuração Metabólica , Dados de Sequência Molecular , Proteínas do Tecido Nervoso/química , Neurotransmissores/metabolismo , Proteínas da Membrana Plasmática de Transporte de Norepinefrina , Octopamina/farmacocinética , Oócitos , Filogenia , Receptores Proteína Tirosina Quinases/química , Homologia de Sequência de Aminoácidos , Transfecção , Xenopus laevis
20.
J Biol Chem ; 274(49): 34535-8, 1999 Dec 03.
Artigo em Inglês | MEDLINE | ID: mdl-10574914

RESUMO

Agonist-dependent activation of the alpha(1)-adrenergic receptor is postulated to be initiated by disruption of an interhelical salt-bridge constraint between an aspartic acid (Asp-125) and a lysine residue (Lys-331) in transmembrane domains three and seven, respectively. Single point mutations that disrupt the charges of either of these residues results in constitutive activity. To validate this hypothesis, we used site-directed mutagenesis to switch the position of these amino acids to observe, if possible, regeneration of the salt-bridge reverses that the constitutive activity of the single point mutations. The transiently expressed switch mutant receptor displayed an altered pharmacological profile. The affinity of selective alpha(1b)-adrenergic receptor antagonists for the switch mutant (D125K/K331D) was no different from the wild-type alpha(1b)-adrenergic receptor, suggesting that both receptors are maintaining similar tertiary structures in the cell membrane. However, there was a significant 4-6-fold decrease in the affinity of protonated amine receptor agonists and a 3-6-fold increase in the affinity of carboxylated catechol derivatives for the switch mutant compared with the wild-type alpha(1b)-adrenergic receptor. This pharmacology is consistent with a reversed charge at position 125 in transmembrane domain three. Interestingly, the ability of either a negatively or positively charged agonist to generate soluble inositol phosphates was similar for both types of receptors. Finally, the switch mutant (D125K/K331D) displayed similar basal signaling activity as the wild-type receptor, reversing the constitutive activity of the single point mutations (D125K and K331D). This suggests an ionic constraint has been reformed in the switch mutant analogous to the restraint previously described for the wild-type alpha(1b)-adrenergic receptor. These results strongly establish the disruption of an electrostatic interaction as an initial step in the agonist-dependent activation of alpha(1)-adrenergic receptors.


Assuntos
Mutação , Receptores Adrenérgicos alfa 1/genética , Receptores Adrenérgicos alfa 1/metabolismo , Agonistas de Receptores Adrenérgicos alfa 1 , Antagonistas de Receptores Adrenérgicos alfa 1 , Aminoácidos/metabolismo , Animais , Células COS , Catecolaminas/farmacocinética , Fosfatos de Inositol/metabolismo , Íons , Mutagênese Sítio-Dirigida , Fenilefrina/farmacocinética , Ligação Proteica
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