RESUMO
Pineal gland secretes the hormone melatonin at night with a circadian rhythm. The synthesis and secretion of melatonin are stimulated at night by norepinephrine released by sympathetic postganglionic neurons projecting from the superior cervical ganglia. Norepinephrine simultaneously activates α- and ß-adrenoceptors, triggering melatonin synthesis.To study the regulation of melatonin production and secretion, it is very convenient to use an ex vivo preparation. Thus, it is possible to keep intact pineal glands in culture and to study the actions of agonists, antagonists, modulators, toxic agents, etc., in melatonin synthesis. Artificial melatonin synthesis stimulation in vitro is usually achieved by using a ß-adrenergic agonist alone or in association with an α-adrenergic agonist. In this chapter, the methodology of cultured pineal glands will be described. Several papers were published by our group using this methodology, approaching the role played in melatonin synthesis control by angiotensin II and IV, insulin, glutamate, voltage-gated calcium channels, anhydroecgonine methyl ester (AEME, crack-cocaine product), monosodium glutamate (MSG), signaling pathways like NFkB, pathophysiological conditions like diabetes, etc.
Assuntos
Cocaína , Insulinas , Melatonina , Glândula Pineal , Agonistas alfa-Adrenérgicos/metabolismo , Agonistas alfa-Adrenérgicos/farmacologia , Agonistas Adrenérgicos beta/farmacologia , Angiotensina II/metabolismo , Canais de Cálcio/metabolismo , Ritmo Circadiano/fisiologia , Melatonina/metabolismo , Norepinefrina , Glândula Pineal/metabolismo , Receptores Adrenérgicos beta/metabolismo , Glutamato de SódioRESUMO
Octopamine (OA), a biogenic monoamine, is known to mediate several immune responses. This study analyzed the effects of OA on immunological regulation in the tiger shrimp Penaeus monodon. The immune parameters including total haemocyte count, differential haemocyte count, phenoloxidase activity, respiratory bursts, superoxide dismutase activity, and phagocytic activity and clearance efficiency in response to the pathogen, Photobacterium damselae, were determined when shrimp were individually injected with saline or OA at 100 or 1000â¯pmol shrimp-1. In addition, the intracellular second messengers in haemocyte such as Ca2+ and adenosine 3',5'-cyclic monophosphate (cAMP) were examined in shrimp receiving saline or OA at 1 or 10â¯nmol shrimp-1. Results showed that all of the immune parameters significantly increased at 2-4â¯h in OA-injected shrimp except hyaline cells in 100â¯pmol shrimp-1-injected shrimp at 4â¯h, but phenoloxidase activity per granulocyte significantly decreased at 2-4â¯h. However, these had returned to saline control levels after receiving OA for 8â¯h except differential haemocyte count and phenoloxidase activity per granulocyte for 16â¯h. An injection of OA also significantly increased the survival rate of shrimp challenged with Pho. damselae. Shrimp receiving OA at 1 and 10â¯nmol shrimp-1 significantly increased the intracellular Ca2+ concentration ([Ca2+]i) at 30-60â¯min and 30â¯min, and cAMP concentration [cAMP]i) at 5-15â¯min and 15â¯min, respectively. However, [Ca2+]i at 50-60â¯min, and [cAMP]i at 30-60â¯min returned to saline control when the shrimp received OA at 10â¯nmol shrimp-1, and at 1 and 10â¯nmol shrimp-1, respectively. These results suggest that OA administration by injection at ≤1000â¯pmol shrimp-1 mediates transient upregulation of immunity together with the increased resistance of P. monodon to Pho. damselae, which are modulated through intracellular Ca2+ and cAMP second messenger pathways.
Assuntos
Regulação da Expressão Gênica/imunologia , Imunidade Inata/efeitos dos fármacos , Octopamina/metabolismo , Penaeidae/genética , Penaeidae/imunologia , Transdução de Sinais/imunologia , Adjuvantes Imunológicos/farmacologia , Agonistas alfa-Adrenérgicos/administração & dosagem , Agonistas alfa-Adrenérgicos/metabolismo , Animais , Cálcio/metabolismo , AMP Cíclico/metabolismo , Perfilação da Expressão Gênica , Octopamina/administração & dosagem , Photobacterium/fisiologia , Transdução de Sinais/efeitos dos fármacos , Regulação para Cima/imunologiaRESUMO
Type I Diabetes (T1D) is associated with reduced leptin levels and increased stress axis activity marked by elevations in norepinephrine (NE) levels in the paraventricular nucleus (PVN) of the hypothalamus. We hypothesized that leptin suppresses stress axis activity in T1D through central and peripheral mechanisms. In the first experiment, adult male Sprague Dawley rats were implanted with a cannula in the PVN and randomly divided into a non-diabetic group treated with vehicle (nâ¯=â¯6) and a diabetic group treated with streptozotocin (nâ¯=â¯13). Food intake and water intake was measured for 14â¯days. On the last day, a subset of diabetic rats were treated with 500⯵g of leptin i.p. Rats were subjected to push-pull perfusion of the PVN and hourly blood sampling for 5â¯h. In the next experiment, diabetic rats were treated either with an alpha-2 adrenergic agonist, clonidine (CLON), or a beta adrenergic agonist isoproterenol (ISO), to reverse the effects of leptin. Rats were subjected to push pull perfusion and hourly blood sampling. In experiment 1, T1D increased food intake, water intake, NE release in the PVN and circulating CS levels. Leptin treatment decreased NE release modestly but produced a robust reduction in corticosterone (CS) levels. In experiment 2, CLON but not ISO was able to reverse the effect of leptin on NE levels in the PVN, however, both agonists were capable of blocking leptin's effects on circulating CS. These results suggest that in diabetic rats, the sensitivity of the hypothalamus to beta adrenergic agonists is altered, while the adrenals remain sensitive to both alpha and beta adrenergic agonists.
Assuntos
Leptina/metabolismo , Norepinefrina/metabolismo , Núcleo Hipotalâmico Paraventricular/fisiopatologia , Agonistas Adrenérgicos/metabolismo , Agonistas alfa-Adrenérgicos/metabolismo , Animais , Clonidina/farmacologia , Corticosterona/análise , Corticosterona/sangue , Diabetes Mellitus Experimental/fisiopatologia , Sistema Hipotálamo-Hipofisário/efeitos dos fármacos , Isoproterenol/farmacologia , Leptina/fisiologia , Masculino , Norepinefrina/fisiologia , Sistema Hipófise-Suprarrenal/efeitos dos fármacos , Ratos , Ratos Sprague-DawleyRESUMO
NEW FINDINGS: What is the central question of this study? We sought to determine whether human skeletal muscle feed arteries (SFMAs) express TRPV1 channels and what role they play in modulating vascular function. What is the main finding and its importance? Human SMFAs do express functional TRPV1 channels that modulate vascular function, specifically opposing α-adrenergic receptor-mediated vasocontraction and potentiating vasorelaxation, in an endothelium-dependent manner, as evidenced by the α1 -receptor-mediated responses. Thus, the vasodilatory role of TRPV1 channels, and their ligand capsaicin, could be a potential therapeutic target for improving vascular function. Additionally, given the 'sympatholytic' effect of TRPV1 activation and known endogenous activators (anandamide, reactive oxygen species, H+ , etc.), TRPV1 channels might contribute to functional sympatholysis during exercise. To examine the role of the transient receptor potential vanilloid type 1 (TRPV1 ) ion channel in the vascular function of human skeletal muscle feed arteries (SMFAs) and whether activation of this heat-sensitive receptor could be involved in modulating vascular function, SMFAs from 16 humans (63 ± 5 years old, range 41-89 years) were studied using wire myography with capsaicin (TRPV1 agonist) and without (control). Specifically, phenylephrine (α1 -adrenergic receptor agonist), dexmedetomidine (α2 -adrenergic receptor agonist), ACh and sodium nitroprusside concentration-response curves were established to assess the role of TRPV1 channels in α-receptor-mediated vasocontraction as well as endothelium-dependent and -independent vasorelaxation, respectively. Compared with control conditions, capsaicin significantly attenuated maximal vasocontraction in response to phenylephrine [control, 52 ± 8% length-tensionmax (LTmax ) and capsaicin, 21 ± 5%LTmax ] and dexmedetomidine (control, 29 ± 12%LTmax and capsaicin, 2 ± 3%LTmax ), while robustly enhancing maximal vasorelaxation with ACh (control, 78 ± 8% vasorelaxation and capsaicin, 108 ± 13% vasorelaxation) and less clearly enhancing the sodium nitroprusside response. Denudation of the endothelium greatly attenuated the maximal ACh-induced vasorelaxation equally in the control and capsaicin conditions (â¼17% vasorelaxation) and abolished the attenuating effect of capsaicin on the maximal phenylephrine response (denuded + capsaicin, 61 ± 20%LTmax ). Immunohistochemistry identified a relatively high density of TRPV1 channels in the endothelium compared with the smooth muscle of the SMFAs, but because of the far greater volume of smooth muscle, total TRPV1 protein content was not significantly attenuated by denudation. Thus, SMFAs ubiquitously express functional TRPV1 channels, which alter vascular function, in terms of α1 -receptors, in a predominantly endothelium-dependent manner, conceivably contributing to the functional sympatholysis and unveiling a therapeutic target.
Assuntos
Artérias/metabolismo , Músculo Esquelético/metabolismo , Músculo Liso Vascular/metabolismo , Canais de Cátion TRPV/metabolismo , Agonistas alfa-Adrenérgicos/metabolismo , Adulto , Idoso , Idoso de 80 Anos ou mais , Artérias/efeitos dos fármacos , Capsaicina/farmacologia , Endotélio Vascular/efeitos dos fármacos , Endotélio Vascular/metabolismo , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Músculo Esquelético/irrigação sanguínea , Músculo Esquelético/efeitos dos fármacos , Músculo Liso Vascular/irrigação sanguínea , Músculo Liso Vascular/efeitos dos fármacos , Nitroprussiato/farmacologia , Fenilefrina/farmacologia , Receptores Adrenérgicos alfa/metabolismo , Vasoconstrição/efeitos dos fármacos , Vasodilatação/efeitos dos fármacosRESUMO
Most vertebrates increase cardiac output during activity by elevating heart rate with relatively stable stroke volume. However, several studies have demonstrated 'intrinsic autoregulation' of cardiac output where artificially increased heart rate is associated with decreased stroke volume, leaving cardiac output unchanged. We explored the capacity of noradrenaline to overcome autoregulation in the anaconda heart. Electrically pacing in situ perfused hearts from the intrinsic heart rate to the maximum attainable resulted in a proportional decrease in stroke volume. However, noradrenaline, which increased heart rate to the same frequency as pacing, maintained stroke volume and thus increased cardiac output. In atrial and ventricular preparations, noradrenaline significantly increased the force of contraction and contraction kinetics. Thus, the increased contractility associated with adrenergic stimulation ameliorates filling limitations at high heart rates. Although heart rate appears the primary regulated variable during activity, this may only be achieved with compensatory amendments in myocardial contractility provided by adrenergic stimulation.
Assuntos
Boidae/fisiologia , Coração/fisiologia , Norepinefrina/metabolismo , Agonistas alfa-Adrenérgicos/metabolismo , Animais , Débito Cardíaco , Homeostase , Masculino , Contração Miocárdica , Volume SistólicoRESUMO
The host stress hormone norepinephrine (NE), also called noradrenaline, is reported to augment bacterial growth and pathogenicity, but few studies have focused on the effect of NE on the activity of antimicrobials. The aim of this study was to clarify whether NE affects antimicrobial activity against multidrug-resistant Acinetobacter baumannii (MDR-AB). Time-kill studies of tigecycline (TIG) and colistin (COL) against MDR-AB as well as assays for factors contributing to antibiotic resistance were performed using MDR-AB clinical strains both in the presence and absence of 10 µM NE. In addition, expression of three efflux pump genes (adeB, adeJ and adeG) in the presence and absence of NE was analysed by quantitative reverse transcription PCR. Viable bacterial cell counts in TIG-supplemented medium containing NE were significantly increased compared with those in medium without NE. In contrast, NE had little influence on viable bacterial cell counts in the presence of COL. NE-supplemented medium resulted in an ca. 2 log increase in growth and in bacterial cell numbers adhering on polyurethane, silicone and polyvinylchloride surfaces. Amounts of biofilm in the presence of NE were ca. 3-fold higher than without NE. Expression of the adeG gene was upregulated 4-6-fold in the presence of NE. In conclusion, NE augmented factors contributing to antibiotic resistance and markedly reduced the in vitro antibacterial activity of TIG against MDR-AB. These findings suggest that NE treatment may contribute to the failure of TIG therapy in patients with MDR-AB infections.
Assuntos
Acinetobacter baumannii/efeitos dos fármacos , Agonistas alfa-Adrenérgicos/metabolismo , Antibacterianos/farmacologia , Antagonismo de Drogas , Minociclina/análogos & derivados , Norepinefrina/metabolismo , Acinetobacter baumannii/fisiologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Aderência Bacteriana , Carga Bacteriana , Biofilmes/crescimento & desenvolvimento , Colistina/metabolismo , Meios de Cultura/química , Feminino , Perfilação da Expressão Gênica , Genes Bacterianos , Humanos , Masculino , Viabilidade Microbiana/efeitos dos fármacos , Pessoa de Meia-Idade , Minociclina/farmacologia , Reação em Cadeia da Polimerase em Tempo Real , Tigeciclina , Fatores de TempoRESUMO
Epidemiological and experimental evidence has shown that psychological stress can propel cancer progression. However, its role in anti-angiogenic therapy is not well understood. We previously found that exogenous norepinephrine attenuated the effect of sunitinib, a multi-targeted anti-angiogenic agent, in a mouse melanoma model. Here, we further evaluated the effects of chronic stress on sunitinib therapy in colorectal cancer models. We found that chronic restraint stress markedly weakened the efficacy of sunitinib, primarily through promoting the expression of vascular endothelial growth factor (VEGF) and interleukin-8 (IL-8) to stimulate tumor angiogenesis in vivo. This effect could be sufficiently mimicked by exogenous norepinephrine and blocked by the ß-antagonist propranolol. In vitro, norepinephrine up-regulated expression of VEGF and IL-8 in sunitinib-treated cancer cells mainly through the ß-adrenoceptor-cAMP-PKA signaling pathway. Norepinephrine also abrogated sunitinib-induced inhibition of cancer cell migration, but had no effect on direct anti-proliferative activity of sunitinib on cancer cells. These findings suggest that psychological stress might attenuate anti-angiogenic therapy primarily through activating beta-adrenergic signaling to promote tumor angiogenesis. It is also suggested that ß-blockers might improve anti-angiogenic outcome under psychological stress.
Assuntos
Agonistas alfa-Adrenérgicos/metabolismo , Inibidores da Angiogênese/farmacologia , Neoplasias Colorretais/tratamento farmacológico , Neoplasias Colorretais/metabolismo , Indóis/farmacologia , Norepinefrina/metabolismo , Pirróis/farmacologia , Estresse Psicológico , Animais , Linhagem Celular Tumoral , Doença Crônica , Modelos Animais de Doenças , Feminino , Humanos , Interleucina-8/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Receptores Adrenérgicos beta/metabolismo , Transdução de Sinais , Sunitinibe , Fator A de Crescimento do Endotélio Vascular/metabolismoRESUMO
A number of hypotheses have been put forth regarding the etiology of schizophrenia, including the dopamine hypothesis, NMDA receptor hypofunction hypothesis, and others. A lesser known theory is that elevated noradrenergic signaling plays a causative role in the disease. This paper briefly re-examines the merits of this hypothesis, including as it relates to some recently published studies. Several lines of evidence are investigated, including: endogenous level studies of norepinephrine (NE); modulation of the disease by noradrenergic drugs; association of the disease with bipolar disorder and hypertension, since these latter two conditions may involve elevated NE transmission; and effects of psychological stress on the disease, since stress can produce elevated release of NE. For many of these lines of evidence, their relationship with prepulse inhibition of startle is examined. A number of these studies support the hypothesis, and several suggest that elevated NE signaling plays a particularly prominent role in the paranoid subtype of schizophrenia. If the hypothesis is correct for some persons, conventional pharmaceutical treatment options, such as use of atypical antipsychotics (which may themselves modulate noradrenergic signaling), may be improved if selective NE transmission modulating agents are added to or even substituted for these conventional drugs.
Assuntos
Agonistas alfa-Adrenérgicos/metabolismo , Norepinefrina/metabolismo , Esquizofrenia/etiologia , Transdução de Sinais , Agonistas alfa-Adrenérgicos/efeitos adversos , Antipsicóticos/efeitos adversos , Transtorno Bipolar/complicações , Humanos , Hipertensão/complicações , Norepinefrina/fisiologia , Esquizofrenia/fisiopatologia , Estresse Psicológico/complicações , Estresse Psicológico/metabolismoRESUMO
Microglia, the resident immune cells of the central nervous system (CNS), monitor the brain for disturbances of tissue homeostasis by constantly moving their fine processes. Microglia respond to tissue damage through activation of ATP/ADP receptors followed by directional process extension to the damaged area. A common feature of several neurodegenerative diseases is the loss of norepinephrine, which might contribute to the associated neuroinflammation. We carried out a high resolution analysis of the effects of norepinephrine (NE) on microglial process dynamics in acute brain slices from mice that exhibit microglia-specific enhanced green fluorescent protein expression. Bath application of NE to the slices resulted in significant process retraction in microglia. Analysis of adrenergic receptor expression with quantitative PCR indicated that resting microglia primarily express ß2 receptors but switch expression to α2A receptors under proinflammatory conditions modeled by LPS treatment. Despite the differential receptor expression, NE caused process retraction in both resting and LPS-activated microglia cultured in the gelatinous substrate Matrigel in vitro. The use of subtype-selective receptor agonists and antagonists confirmed the involvement of ß2 receptors in mediating microglial process dynamics in resting cells and α2A receptors in activated cells. Co-application of NE with ATP to resting microglia blocked the ATP-induced process extension and migration in isolated microglia, and ß2 receptor antagonists prolonged ATP effects in brain slice tissues, suggesting the presence of cross-talk between adrenergic and purinergic signaling in microglia. These data show that the neurotransmitter NE can modulate microglial motility, which could affect microglial functions in pathogenic situations of either elevated or reduced NE levels.
Assuntos
Trifosfato de Adenosina/farmacologia , Agonistas alfa-Adrenérgicos/farmacologia , Movimento Celular/efeitos dos fármacos , Microglia/metabolismo , Norepinefrina/farmacologia , Receptores Adrenérgicos alfa 2/metabolismo , Receptores Adrenérgicos beta 2/metabolismo , Agonistas alfa-Adrenérgicos/metabolismo , Animais , Movimento Celular/fisiologia , Células Cultivadas , Camundongos , Camundongos Transgênicos , Microglia/citologia , Norepinefrina/metabolismo , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/fisiologiaRESUMO
The sympathetic nervous system relies on distinct populations of neurons that use noradrenaline or acetylcholine as neurotransmitter. We show that fating of the sympathetic lineage at early stages results in hybrid precursors from which, genetic cell-lineage tracing reveals, all types progressively emerge by principal mechanisms of maintenance, repression and induction of phenotypes. The homeobox transcription factor HMX1 represses Tlx3 and Ret, induces TrkA and maintains tyrosine hydroxylase (Th) expression in precursors, thus driving segregation of the noradrenergic sympathetic fate. Cholinergic sympathetic neurons develop through cross-regulatory interactions between TRKC and RET in precursors, which lead to Hmx1 repression and sustained Tlx3 expression, thereby resulting in failure of TrkA induction and loss of maintenance of Th expression. Our results provide direct evidence for a model in which diversification of noradrenergic and cholinergic sympathetic neurons is based on a principle of cross-repressive functions in which the specific cell fates are directed by an active suppression of the expression of transcription factors and receptors that direct the alternative fate.
Assuntos
Diferenciação Celular , Neurônios Colinérgicos/citologia , Proteínas de Homeodomínio/metabolismo , Receptores de Fatores de Crescimento/metabolismo , Fatores de Transcrição/metabolismo , Tirosina 3-Mono-Oxigenase/genética , Acetilcolina/metabolismo , Agonistas alfa-Adrenérgicos/metabolismo , Animais , Agonistas Colinérgicos/metabolismo , Neurônios Colinérgicos/fisiologia , Cromossomos Artificiais Bacterianos , Embrião de Mamíferos , Feminino , Regulação da Expressão Gênica no Desenvolvimento , Biblioteca Gênica , Proteínas de Homeodomínio/genética , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Norepinefrina/metabolismo , Fenótipo , Proteínas Proto-Oncogênicas c-ret/genética , Proteínas Proto-Oncogênicas c-ret/metabolismo , Receptor trkC/genética , Receptor trkC/metabolismo , Receptores de Fatores de Crescimento/genética , Sistema Nervoso Simpático/citologia , Sistema Nervoso Simpático/fisiologia , Fatores de Transcrição/genética , Tirosina 3-Mono-Oxigenase/metabolismoRESUMO
The properties of the hyperpolarization-activated cation current (I(h)) were investigated in rat periglomerular dopaminergic neurons using patch-clamp recordings in thin slices. A reliable identification of single dopaminergic neurons was made possible by use of a transgenic line of mice expressing eGFP under the tyrosine hydroxylase promoter. At 37 °C and minimizing the disturbance of the intracellular milieu with perforated patches, this current shows a midpoint of activation around -82.7 mV, with a significant level of opening already at rest, thereby giving a substantial contribution to the resting potential, and ultimately playing a relevant function in the control of the cell excitability. The blockage of I(h) has a profound influence on the spontaneous firing of these neurons, which result as strongly depressed. However the effect is not due to a direct role of the current in the pacemaker process, but to the I(h) influence on the resting membrane potential. I(h) kinetics is sensitive to the intracellular levels of cAMP, whose increase promotes a shift of the activation curve towards more positive potentials. The direct application of DA and 5-HT neurotransmitters, physiologically released onto bulbar dopaminergic neurons and known to act on metabotropic receptors coupled to the cAMP pathway, do not modifythe I(h) amplitude. On the contrary, noradrenaline almost halves the I(h) amplitude. Our data indicate that the HCN channels do not participate directly to the pacemaker activity of periglomerular dopaminergic neurons, but influence their resting membrane potential by controlling the excitability profile of these cells, and possibly affecting the processing of sensory information taking place at the entry of the bulbar circuitry.
Assuntos
Canais de Cátion Regulados por Nucleotídeos Cíclicos/fisiologia , Neurônios Dopaminérgicos/fisiologia , Bulbo Olfatório/fisiologia , Canais de Potássio/fisiologia , 1-Metil-3-Isobutilxantina/farmacologia , Agonistas alfa-Adrenérgicos/metabolismo , Agonistas alfa-Adrenérgicos/farmacologia , Animais , Benzazepinas/farmacologia , Colforsina/farmacologia , AMP Cíclico/metabolismo , AMP Cíclico/farmacologia , Canais de Cátion Regulados por Nucleotídeos Cíclicos/metabolismo , Dopamina/metabolismo , Dopamina/farmacologia , Neurônios Dopaminérgicos/metabolismo , Proteínas de Fluorescência Verde/genética , Proteínas de Fluorescência Verde/metabolismo , Canais Disparados por Nucleotídeos Cíclicos Ativados por Hiperpolarização , Ivabradina , Cinética , Potenciais da Membrana/efeitos dos fármacos , Potenciais da Membrana/fisiologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Norepinefrina/metabolismo , Norepinefrina/farmacologia , Bulbo Olfatório/citologia , Bulbo Olfatório/metabolismo , Técnicas de Patch-Clamp , Canais de Potássio/metabolismo , Pirimidinas/farmacologia , Ratos , Tirosina 3-Mono-Oxigenase/genética , Tirosina 3-Mono-Oxigenase/metabolismoRESUMO
ERK1/2 (extracellular-signal-regulated kinase 1/2) and their substrates RSKs (p90 ribosomal S6 kinases) phosphorylate different transcription factors, contributing differentially to transcriptomic profiles. In cardiomyocytes ERK1/2 are required for >70% of the transcriptomic response to endothelin-1. In the present study we investigated the role of RSKs in the transcriptomic responses to the G(q)-protein-coupled receptor agonists endothelin-1, phenylephrine (a generic α(1)-adrenergic receptor agonist) and A61603 (α(1A)-adrenergic receptor selective). Phospho-ERK1/2 and phospho-RSKs appeared in cardiomyocyte nuclei within 2-3 min of stimulation (endothelin-1>A61603≈phenylephrine). All agonists increased nuclear RSK2, but only endothelin-1 increased the nuclear RSK1 content. PD184352 (inhibits ERK1/2 activation) and BI-D1870 (inhibits RSKs) were used to dissect the contribution of RSKs to the endothelin-1-responsive transcriptome. Of the 213 RNAs up-regulated after 1 h, 51% required RSKs for their up-regulation, whereas 29% required ERK1/2 but not RSKs. The transcriptomic response to phenylephrine overlapped with, but was not identical with, endothelin-1. As with endothelin-1, PD184352 inhibited the up-regulation of most phenylephrine-responsive transcripts, but the greater variation in the effects of BI-D1870 suggests that differential RSK signalling influences global gene expression. A61603 induced similar changes in RNA expression in cardiomyocytes as phenylephrine, indicating that the signal was mediated largely through α(1A)-adrenergic receptors. A61603 also increased expression of immediate early genes in perfused adult rat hearts and, as in cardiomyocytes, up-regulation of the majority of genes was inhibited by PD184352. PD184352 or BI-D1870 prevented the increased surface area induced by endothelin-1 in cardiomyocytes. Thus RSKs play a significant role in regulating cardiomyocyte gene expression and hypertrophy in response to G(q)-protein-coupled receptor stimulation.
Assuntos
Agonistas alfa-Adrenérgicos/farmacologia , Regulação da Expressão Gênica , Miócitos Cardíacos/metabolismo , Receptor de Endotelina A/agonistas , Receptores Adrenérgicos alfa 1/metabolismo , Proteínas Quinases S6 Ribossômicas 90-kDa/metabolismo , Agonistas alfa-Adrenérgicos/metabolismo , Animais , Benzamidas/farmacologia , Núcleo Celular/metabolismo , Masculino , Proteína Quinase 3 Ativada por Mitógeno/genética , Proteína Quinase 3 Ativada por Mitógeno/metabolismo , Miócitos Cardíacos/citologia , RNA/metabolismo , Ratos , Ratos Sprague-Dawley , Receptor de Endotelina A/metabolismo , Proteínas Quinases S6 Ribossômicas 90-kDa/genética , Transdução de SinaisRESUMO
BACKGROUND: Cocaine has long been known to increase blood pressure, but the degree and mechanism of vasoconstricting action remain poorly understood. Here we examine the interaction between cocaine and alpha-adrenoceptor agonists, with the action of reuptake inhibition minimized. METHODS: Cocaine was administered to isolated rings of rat thoracic aorta, alone and in combination with three different adrenoceptor agonists: phenylephrine, methoxamine, and norepinephrine. Synergy analysis begins with the predicted additive effect of the combination of two agonists, based upon dose equivalence theory. This case where one agonist (cocaine) has no effect when administered alone requires only a t-test to demonstrate that a departure from additivity has occurred. RESULTS: At doses where cocaine alone produced no vasoconstriction, it potentiated the vasoconstriction produced by all three alpha agonists, a clear indication of synergism between cocaine and these agents. Higher doses of cocaine in combination with alpha adrenoceptor agents gave an inverted-U shaped (hormetic) dose-effect curve, i.e., dose-related relaxation at higher doses. The hormetic dose-effect relation was analyzed using computational methodology based on dose equivalence to derive the unknown second component of action that causes relaxation. CONCLUSIONS: Cocaine exhibits both vasoconstricting and vasorelaxant effects. This relaxing component, possibly related to activation of myosin light chain phosphatase, was quantified as a dose-effect curve. Most important is the synergism between cocaine and alpha-adrenoceptor stimulation which cannot be explained as an action due to reuptake inhibition, and has not been previously described.
Assuntos
Inibidores da Captação Adrenérgica/farmacologia , Agonistas alfa-Adrenérgicos/farmacologia , Aorta Torácica/efeitos dos fármacos , Cocaína/farmacologia , Biologia Computacional/métodos , Vasoconstrição/efeitos dos fármacos , Vasodilatação/efeitos dos fármacos , Inibidores da Captação Adrenérgica/metabolismo , Agonistas alfa-Adrenérgicos/metabolismo , Animais , Aorta Torácica/metabolismo , Cocaína/metabolismo , Sinergismo Farmacológico , Masculino , Ratos , Ratos Sprague-Dawley , Vasoconstrição/fisiologia , Vasodilatação/fisiologiaRESUMO
The fluorescence and ultraviolet spectroscopy were explored to study the interaction between Oxymetazoline hydrochloride (OMZH) and mucin under imitated physiological condition. The results demonstrated that the fluorescence quenching mechanism between OMZH and mucin is a combined quenching process. The binding constants (K(a)), binding sites (n) and the corresponding thermodynamic parameters (ΔG, ΔH, and ΔS) of the interaction system were calculated at different temperatures. The hydrogen bonds and van der Waals forces play a major role in the interaction between OMZH and mucin. According to Förster non-radiation energy transfer theory, the binding distance between OMZH and mucin was calculated.
Assuntos
Agonistas alfa-Adrenérgicos/metabolismo , Mucinas/metabolismo , Descongestionantes Nasais/metabolismo , Oximetazolina/metabolismo , Agonistas alfa-Adrenérgicos/química , Sítios de Ligação , Mucinas/química , Descongestionantes Nasais/química , Oximetazolina/química , Ligação Proteica , Espectrometria de Fluorescência , Espectrofotometria Ultravioleta , TermodinâmicaRESUMO
The fluorescence and ultraviolet spectroscopy were explored to study the interaction between Naphazoline hydrochloride (Naphcon) and bovine serum albumin (BSA) at three different temperatures (292, 301, and 310 K) under imitated physiological conditions. The quenching mechanism of BSA by Naphacon was interpreted using the Stern-Volmer mechanism, and a combined quenching (dynamic and static quenching). The binding constants, binding sites and the corresponding thermodynamic parameters (ΔG, ΔH, and ΔS) of the interaction system were calculated at different temperatures. According to Förster non-radiation energy transfer theory, the binding distance between BSA and Naphcon was found to be 4.71 nm. Synchronous fluorescence spectroscopy showed the conformation of BSA changed in the presence of Naphacon. In addition, the effect of some common metal ions (Mg(2+), Ca(2+), Ni(2+), Cu(2+), and Fe(2+)) on the binding constant between Naphcon and BSA was examined.
Assuntos
Agonistas alfa-Adrenérgicos/metabolismo , Nafazolina/metabolismo , Soroalbumina Bovina/metabolismo , Agonistas alfa-Adrenérgicos/química , Animais , Sítios de Ligação , Bovinos , Nafazolina/química , Ligação Proteica , Soroalbumina Bovina/química , Espectrometria de Fluorescência , Espectrofotometria Ultravioleta , TermodinâmicaRESUMO
Female sperm storage is common among organisms with internal fertilization. It is important for extended fertility and, in cases of multiple mating, for sperm competition. The physiological mechanisms by which females store and manage stored sperm are poorly understood. Here, we report that the biogenic amines tyramine (TA) and octopamine (OA) in Drosophila melanogaster females play essential roles in sperm storage. D. melanogaster females store sperm in two types of organs, a single seminal receptacle and a pair of spermathecae. We examined sperm storage parameters in females mutant in enzymes required for the biochemical synthesis of tyrosine to TA and TA to OA, respectively. Postmating uterine conformational changes, which are associated with sperm entry and accumulation into storage, were unaffected by the absence of either TA or OA. However, sperm release from storage requires both TA and OA; sperm were retained in storage in both types of mutant females at significantly higher levels than in control flies. Absence of OA inhibited sperm depletion only from the seminal receptacle, whereas absence of both OA and TA perturbed sperm depletion from both storage organ types. We find innervation of the seminal receptacle and spermathecae by octopaminergic-tyraminergic neurons. These findings identify a distinct role for TA and OA in reproduction, regulating the release of sperm from storage, and suggest a mechanism by which Drosophila females actively regulate the release of stored sperm.
Assuntos
Neurotransmissores/metabolismo , Octopamina/metabolismo , Espermatozoides/metabolismo , Tiramina/metabolismo , Inibidores da Captação Adrenérgica/metabolismo , Agonistas alfa-Adrenérgicos/metabolismo , Animais , Cruzamentos Genéticos , Proteínas de Drosophila/fisiologia , Drosophila melanogaster , Feminino , Fertilização , Masculino , Modelos Biológicos , Mutação , Neurônios/metabolismo , ReproduçãoRESUMO
Our light and electron microscopy observations have revealed that the chromatic unit for the caudal fin in the blue variant of the Siamese fighting fish consists exclusively of dermal chromatophores comprised of compact and overlapping light-reflecting motile iridophores underlined by a layer of light absorbing melanophores. The 2 subtypes that make up about 70% of the skin tissue are located just below the basal layer of the considerably thin epidermis. The administration of K-rich saline or norepinephrine induced prompt, but gradual and reversible, changes in the color of the skin from blue to a brown-yellowish color. The induced color change is attributable either to the neurotransmitter releasing effects of the K-rich saline or to the direct effects of norepinephrine on the postsynaptic alpha adrenergic receptors. Both of these agents induced aggregation of the melanosomes within the melanophores and apparently shifted the wavelength of the light reflected by the iridophores towards the shorter (blue) end of the spectrum. Based on the distribution and architectural arrangement of the iridophores and melanophores as well as their physiological responses, we conclude that the generation of the blue coloration in this fish predominantly occurs through motile iridophores via a multilayered thin-film interference phenomenon of the non-ideal type. The presence of the underlying melanophores provides a black sheet of melanin that enhances the chroma and purity of the color.
Assuntos
Cromatóforos/citologia , Cromatóforos/metabolismo , Perciformes/fisiologia , Pele/citologia , Agonistas alfa-Adrenérgicos/metabolismo , Animais , Cromatóforos/efeitos dos fármacos , Cor , Norepinefrina/metabolismo , Pele/efeitos dos fármacosRESUMO
AIMS: We investigated how modification of levels of the sex hormones 17ß-estradiol and testosterone affects vascular contraction and nongenomic vascular effects of 17ß-estradiol. METHODS: Male and female rats were treated with vehicle, 17ß-estradiol (25 µg/kg/day) or testosterone (1 mg/kg/day) for 14 consecutive days after sham gonadectomy or gonadectomy was performed. Isometric tensions were then measured from mesenteric arteries of each group of rats. RESULTS: Contraction to phenylephrine was increased in mesenteric arteries from rats with or without gonadectomy treated with testosterone for 14 days compared to their intact controls. Contraction to phenylephrine was reduced in mesenteric arteries of rats with or without gonadectomy treated with 17ß-estradiol for 14 days compared to their intact controls. Incubation of mesenteric arteries with 17ß-estradiol (1 nmol/l) for 30 min reduced contraction to phenylephrine in mesenteric arteries of rats that were treated with testosterone for 14 days. This acute incubation of 17ß-estradiol had no effect on arteries from rats that were treated with 17ß-estradiol for 14 days. The acute effect of 17ß-estradiol (1 nmol/l) is preserved in arteries without endothelium. CONCLUSION: Our results suggest that 14 days' testosterone treatment enhances while 14 days' 17ß-estradiol treatment suppresses contraction as well as the nongenomic effects of 17ß-estradiol in the vascular smooth muscles.
Assuntos
Estradiol/farmacologia , Estrogênios/farmacologia , Hormônios Esteroides Gonadais/sangue , Artérias Mesentéricas/fisiologia , Agonistas alfa-Adrenérgicos/metabolismo , Agonistas alfa-Adrenérgicos/farmacologia , Animais , Peso Corporal , Modelos Animais de Doenças , Relação Dose-Resposta a Droga , Esquema de Medicação , Avaliação Pré-Clínica de Medicamentos , Endotélio Vascular/fisiologia , Estradiol/sangue , Estrogênios/sangue , Feminino , Hormônios Esteroides Gonadais/farmacologia , Humanos , Masculino , Músculo Liso Vascular/fisiologia , Fenilefrina/metabolismo , Fenilefrina/farmacologia , Distribuição Aleatória , Ratos , Ratos Sprague-Dawley , Fatores Sexuais , Testosterona/sangue , Testosterona/farmacologia , Vasoconstrição/efeitos dos fármacos , Vasoconstrição/fisiologia , Vasodilatação/efeitos dos fármacos , Vasodilatação/fisiologiaRESUMO
Activation of coronary vascular α-adrenoceptors results in vasoconstriction which competes with metabolic vasodilation during sympathetic activation. Epicardial conduit vessel constriction is largely mediated by α(1)-adrenoceptors; the constriction of the resistive microcirculation largely by α(2)-adrenoceptors, but also by α(1)-adrenoceptors. There is no firm evidence that α-adrenergic coronary vasoconstriction exerts a beneficial effect on transmural blood flow distribution. In fact, α-blockade in anesthetized and conscious dogs improves blood flow to all transmural layers, during normoperfusion and hypoperfusion. Also, in patients with coronary artery disease, blockade of α(1)- and α(2)-adrenoceptors improves coronary blood flow, myocardial function and metabolism.
Assuntos
Circulação Coronária/fisiologia , Receptores Adrenérgicos alfa/metabolismo , Vasoconstrição/fisiologia , Agonistas alfa-Adrenérgicos/metabolismo , Antagonistas Adrenérgicos alfa/metabolismo , Animais , Doença da Artéria Coronariana , Doença das Coronárias/fisiopatologia , Vasos Coronários/metabolismo , Cães , Coração/fisiopatologia , Humanos , Isquemia , Vasodilatação/fisiologiaRESUMO
UNLABELLED: Phosphodiesterase-4 (PDE4) plays a critical role in the regulation of ß-adrenergic receptor-stimulated cyclic adenosine monophosphate cell signaling in the heart. (R)-rolipram, a PDE4-selective inhibitor, has been studied previously as a radiotracer for the quantification of PDE4 levels. The aim of this study was to characterize (R)-(11)C-rolipram binding in the rat myocardium in vivo, using small-animal PET. METHODS: Male Sprague-Dawley rats (n = 30) were administered (R)-(11)C-rolipram and imaged for 60 min to evaluate tracer binding and reproducibility, quantified using Logan slope analysis of the distribution volume. Dynamic (13)N-ammonia imaging was performed to quantify myocardial blood flow and assist in cardiac regional analysis. Saturation studies evaluated the sensitivity of (R)-(11)C-rolipram to PDE4 blocking by unlabeled cold (R)-rolipram (0.0001-1.0 mg/kg), for estimation of the median effective dose (ED(50)) in the heart. (R)-(11)C-rolipram response to enhanced norepinephrine stimulation of the ß-adrenergic receptor with desipramine (20 mg/kg, intravenous) was also studied. Intrarat variability studies (n = 5) were conducted with test-retest imaging at 16 ± 7 d. RESULTS: A reduction of Logan slope was observed with increasing cold mass coadministered with the tracer, with an ED(50) of 0.0019 mg/kg (95% confidence interval, 0.0014-0.0052) estimated from the saturation studies. This ED(50) predicted less than 10% enzyme occupancy at 0.0002 mg of cold (R)-rolipram per kilogram (mass/body weight). Low-occupancy imaging at 0.00018 ± 0.00002 mg/kg produced a mean Logan slope of 5.5 ± 0.85 mL/cm(3). Enzyme saturation of more than 90%, compared with low-occupancy conditions, occurred at more than 0.02 mg/kg, with a complete blocking dose (>1 mg of (R)-rolipram per kilogram) resulting in a Logan slope of 3.3 ± 0.1 mL/cm(3), representing a 40% reduction. Compared with baseline, a Logan slope of 6.8 ± 0.7 mL/cm(3) in desipramine-challenged animals was observed, representing a 30% increase due to acute norepinephrine stimulation, despite a reduction in myocardial blood flow. Intrarat and intraoperator variability was less than 5% between repeated measures. CONCLUSION: (R)-(11)C-rolipram shows the ability to monitor increases and decreases in PDE4 availability in the rat myocardium, with good reproducibility.