Doctor of Philosophy / Department of Animal Sciences and Industry / Bradley J. Johnson / Zilpaterol (ZIL) is a β-adrenergic receptor (β-AR) agonist that has been recently approved for use in feedlot cattle to improve production efficiencies and animal performance. One of the mechanisms through which this occurs is increased skeletal muscle growth. Therefore, two experiments were conducted to determine the effects of ZIL both in vivo and in vitro. In the first experiment, ZIL addition to bovine satellite cells resulted in a tendency to increase IGF-I mRNA and increased myosin heavy chain IIA (MHC) mRNA with 0.001 [micro symbol]M and decreased MHC mRNA with 0.01 and 10 [micro symbol]M. There were no effects of ZIL on protein synthesis or degradation. In myoblast cultures, there was a decrease in all three β-AR mRNA, and this was also reported in western blot analysis with a reduction in β2-AR expression due to ZIL treatment. In myotubes, there was an increase in β2-AR protein expression. In the second and third experiment, ZIL improved performance and carcass characteristics of feedlot steers and heifers. Additionally, ZIL decreased MHC IIA mRNA in semimembranosus muscle tissue collected from both steers and heifers. An additional part of the third study was conducted to determine the effects of melengestrol acetate (MGA) on bovine satellite cell and semimembranosus muscle gene expression. There were no effects of MGA on the expression of genes analyzed from semimembranosus muscle tissue collected. However, the addition of MGA to cultured bovine satellite cells resulted in increased β1 and β2-AR mRNA. These experiments aid in our understanding of the mechanism of action of MGA in heifers, as well as the effects of ZIL on both steers and heifers. Furthermore, they increase our knowledge and understanding of the mechanism of action of ZIL, as well as other β-agonists used to promote growth and efficiency in feedlot animals.

Master of Science / Department of Animal Sciences and Industry / Bradley J. Johnson / Zilpaterol hydrochloride (ZH) is a growth promotant that is approved for use in finishing cattle to improve growth performance and increase lean tissue accumulation. Little is known about the combined effects of ZH with anabolic steroid hormone implants. There is also little published data on the effect these growth promotants have on genes that play a role in skeletal muscle synthesis and degradation. Therefore, two separate studies were conducted to address these issues. The first study evaluated the effects of ZH and the steroid implant Revalor-S (RS) on animal performance and skeletal muscle gene expression in feedlot steers. Four treatments were used to analyze the effects of RS implanted 58 days before ZH, which was fed for 30 days with a 3 day withdrawal. It was determined that ZH and RS additively contribute to improved live and carcass performance; however these compounds had different effects on the abundance of the receptors for ZH as well as the abundance of myosin heavy chain (MHC) mRNA in skeletal muscle of feedlot steers. It was also determined that ZH can cause a transition in the abundance of MHC mRNA isoforms in skeletal muscle that are available for the translation of larger, faster, more glycolytic fiber types of MHC. The second study evaluated the effects of two types of anabolic steroid hormones on myosin heavy chain gene expression. Four treatments were used to measure the effects of trenbolone acetate (TBA) and estradiol (E[subscript]2) on performance and the amount of MHC mRNA in skeletal muscle of finishing steers. It was determined that anabolic steroid implants improve live animal performance, however there was no alteration in the abundance of MHC mRNA in skeletal muscle of feedlot steer for 28 days after implantation; however there was an increase in intermediate fiber type IIA of MHC mRNA in skeletal muscle with increasing days on feed. From these studies we concluded that ZH and anabolic steroids do have an effect on growth performance; however they may differ in the distinct mechanism of action utilized to enhance lean tissue deposition in feedlot steers.

Placental insufficiency-induced intrauterine growth restriction (IUGR) increases risk of mortality and morbidity in newborn infants and domestic animals. IUGR fetuses are typically exposed to prolonged hypoxemia, hypoglycemia, and hypercatecholaminemia, which results in perinatal pancreatic β-cell dysfunction. Recent evidence indicates that chronic exposure to norepinephrine in utero suppresses insulin secretion through α2-adrenergic receptors (ARs), but if the adrenergic actions are blocked compensatory hyper insulin secretion response is observed in the IUGR sheep fetus. In the current studies, we demonstrate that chronic NE exposure alone can produce the compensatory enhancement of β-cell responsiveness following termination of a chronic NE infusion. In the fetus NE was continuously infused at 1-4 μg/min for seven days starting at 131 days of gestational age (term = 145 days). During treatment, NE infused fetuses had higher (P < 0.05) plasma NE concentrations and lower (P < 0.01) insulin concentrations than vehicle infused control fetuses. Glucose stimulated insulin secretion (GSIS), which measures β-cell function, prior to NE treatment was not different between treatments. However, insulin concentrations during hyperglycemic steady state period of GSIS studies and area under the curve of glucose-potentiated arginine-induced insulin secretion were higher (P < 0.01) than control values and this augmentation was confirmed at 3 hours, 24 hours, and five days in NE-infused fetuses after discontinuing the infusion. Pancreatic islets isolated within 10 hours post NE infusion had lower (P < 0.05) mRNA expression of α1D (58%), α2A (43%), α2C (42%), α1 (67%) adrenergic receptors (ARs), and uncoupling protein 2 (40%) compared to islets from controls. Isolated islets from NE-infused fetuses 5 days after NE treatment had lower (P < 0.05) inhibitory responsiveness from NE and a greater (P < 0.05) maximal insulin release with glucose simulation in static incubations compared to controls. These findings show that following chronic NE exposure insulin secretion responsiveness was augmented and was coupled with desensitized adrenergic signaling. Moreover, this compensatory β-cell enhancement persists for days indicating chronic NE exposure permanently alters β-cell responsiveness.

No description available.

No description available.

Diabetes Mellitus (DM) is a metabolic disorder in which the body fails to achieve glucose homeostasis, due to either insulin resistance or reduced insulin secretion or both. This inadequate glucose control leads to hyperglycemia which, if left unchecked, leads to secondary complications like nephropathy, neuropathy, retinal degeneration and other serious conditions. In non-disease state, normal glucose level in the blood is maintained by pancreatic β-cells, which secrete insulin. However, during diabetes development, there is loss of β-cell mass and function; resulting in decreased insulin secretion which is the ultimate cause of hyperglycemia. The ability to non-invasively monitor changes in the β-cell mass during the development or treatment of diabetes would be a significant advance in diabetes management. However, a primary limitation for analysis of β-cell mass and developing dysfunction is the lack of specificity of β-cell targeting agents. Our novel approach for achieving the required specificity for a usable β-cell targeted contrast agent is to target a set of receptors on the cell surface that, as a combination, are unique to that cell. Through genetic screening, Glucagon Like Peptide-1 Receptor (GLP-1R) and α2Adrenergic Receptor (α2AR) were chosen as a potential molecular barcode for β-cells since their combination expression is relatively unique to the β-cells. GLP-1R and α2AR are both G-protein couple receptors (GPCRs) that, apart from being a β-cell specific combination, play an important role in regulating fundamental downstream signaling pathways in β-cells. To target these receptors effectively, we synthesized a multivalent ligand composed of Yohimbine (Yhb), an α2 adrenergic receptor (α2AR) antagonist, linked to an active Glucagon-like Peptide 1 analog (GLP-1₇₋₃₆). In this manuscript, I describe the synthesis and characterization of binding selectivity and signaling ability of GLP-1/Yhb at the cellular level. Using high throughput binding assays, we observed high affinity binding of GLP-1/Yhb to βTC3 cells, a β-cell mimetic line expressing both receptors, at a Kd of ~3 nM. Using microscopy, we observed significant Cy5-tagged GLP-1/Yhb binding and rapid internalization in cells expressing the complementary receptor pair at low concentrations, as low as 1 nM and 5 nM. When one of the receptors was made inaccessible due to presence of saturating quantities of a single unlabeled monomer, GLP-1/Yhb-Cy5 failed to bind to the cells at low concentrations (<10 nM). Similarly, in cells where either GLP-1R or α2AR were knocked down (using shRNA), binding of GLP-1/Yhb was significantly reduced (≤half of cells with both receptors), indicating strong selectivity of the ligand to cells expressing the combination of receptors. We also observed that GLP-1/Yhb construct modulates downstream signaling inβ TC3 cells resulting in enhanced Glucose Stimulated Insulin Secretion (GSIS). In presence of stimulatory glucose, GLP-1/Yhb significantly potentiated GSIS with a half-maximal effective dose of 2.6 nM. Compared to GLP-1₇₋₃₆ alone or GLP-1₇₋₃₆ and Yhb monomers added together, only GLP-1/Yhb could significantly potentiate GSIS at 1 nM, demonstrating that GLP-1/Yhb could translate high affinity binding to increased efficacy for GSIS potentiation. Unlike for insulin secretion, high affinity divalent binding did not translate to increased cAMP production at low concentrations, with significant increases above baseline seen only at 10 nM and higher. Nevertheless, these data show that GLP-1/Yhb binds selectively to β-cells and affects signaling, demonstrating its potential for targeted β-cell imaging and therapy. Overall, our work indicates that synthetic heterobivalent ligands, such as GLP-1/Yhb can be developed to increase cellular specificity and sensitivity making them a strong candidate for both noninvasive imaging and targeted therapy.

Opioid/adrenergic interactions were studied to evaluate two hypotheses: (1) naloxone potentiates the effect of epinephrine on cardiac contractility by increasing circulating epinephrine concentrations; and (2) endogenous and exogenous opioids alter left cardiac nerve stimulationinduced norepinephrine release and cardiac function. A canine isolated heart-lung preparation was used for the first study. Plasma epinephrine was determined and myocardial epinephrine uptake was calculated during intravenous epinephrine infusion. Naloxone (4 mg) was given and the epinephrine infusion was repeated. Naloxone increased cardiac contractility, coronary blood flow, and the coronary sinus epinephrine concentration. When coronary blood flow was subsequently held constant (100% above resting), naloxone increased only contractility. This result indicated that the previously observed increase in coronary sinus epinephrine was flow dependent. Corticosterone (an uptake II blocker) was employed as a positive control. Corticosterone increased the contractile response to epinephrine, but unlike naloxone, corticosterone was accompanied by a clear decrease in myocardial epinephrine uptake. The stereospecificity of the response to naloxone was investigated and (+) naloxone equaled or exceeded (-) naloxone in potentiating the inotropic effect of epinephrine. In the second study, the left cardiac nerve was isolated and electrically stimulated in intact dogs. Norepinephrine overflow gradually declined during successive control stimulations. Pretreatment with naloxone (100 Mg/kg) prevented or delayed the decline. An intracoronary dynorphin 1-9 infusion (2 nmol/min/kg for 20 minutes) reduced both norepinephrine overflow and cardiac performance, and both effects were prevented by pretreatment with naloxone (100 /xg/kg) . To summarize, naloxone potentiated the inotropic effect of infused epinephrine without altering circulating epinephrine concentrations or myocardial epinephrine uptake. This effect of naloxone was not stereospecific and probably not mediated through a traditional opiate receptor. Endogenous and exogenous opioids inhibited the left cardiac nerve stimulation-induced norepinephrine overflow, suggesting that opiate receptors may regulate cardiac excitability by modulating norepinephrine release.

Le diabète prédispose à des complications affectant divers organes comme le systémecardiovasculaire. La cardiopathie ischémique chez les patients diabétiques pourrait être liée àl'accumulation de produits avancés de glycation (AGEs). Dans les coeurs de rats ischémiques,l'expression des récepteurs aux AGEs et de ses ligands est considérablement augmentée etimpliquée dans les lésions de l'ischémie / reperfusion (I / R), même en absence de diabète. Ila été récemment rapporté que le myocarde humain diabétique ne peut pas être protégé par lepréconditionnement. Dans ce contexte, notre hypothèse était que le préconditionnement β1-adrénergique pourrait être modifié en présence d'AGE. En utilisant un modèle de coeur isolénon travaillant de rat, cette étude a pour but d’étudier les effets des AGEs sur lacardioprotection induite par la stimulation des récepteurs β1-adrénergiques (β1-ARs) par lexamotérol (xa). Les effets bénéfiques induits par le xa pendant la reperfusion ont étésupprimés par l'administration de l’albumine glyquée (Alb-Gly) pendant la perfusion du xa,tandis que l’albumine (Alb) n’a pas modifié cette protection. Ces résultats suggèrent que lesAGEs suppriment la cardioprotection résultant de l'activation de la voie β1-AR ce quicontribue à des dommages cardio-vasculaires chez les patients diabétiques.D’ autre part, le pyridoxal 5’-phosphate (PLP), un métabolite naturel de la pyridoxine qui estun antagoniste des récepteurs purinergiques, empêche la surcharge cellulaire en calcium etpeut réduire les dommages d'ischémie-reperfusion. Plusieurs travaux ont mis en evidence unediminution des taux du PLP chez les patients souffrant d'un infarctus du myocarde par rapportà un groupe témoin sain. Plus récemment, il a été signalé qu’un taux bas de PLP confère unrisque indépendant de maladie coronaire. Cette corrélation de la réduction du PLP chez lespatients souffrant d’infarctus est soutenue par les effets préventifs de la vitamine B6 sur lesmaladies coronaires et le diabète. Plusieurs travaux ont montré que le PLP et la pyridoxaminepréviennent la progression de la néphropathie induite par la STZ chez les rats diabétiques eninhibant la formation des AGEs. En utilisant un modèle de coeur isolé non-travaillant de rat,cette étude a examiné les mécanismes de préconditionnement pharmacologique (PC) induitpar la stimulation des récepteurs P2Y par le pyridoxal-5'-phosphate (PLP). La suppression del'effet cardioprotecteur du PLP par le MRS2578, antagoniste des récepteurs P2Y6 et parl’U73122 qui bloque la phospholipase C est en accord avec l’implication du récepteur P2Y6dans le préconditionnement. La suppression de l'effet cardioprotecteur du PLP par l'AMPαS,antagoniste des récepteurs P2Y11, l’H89 inhibiteur de la PKA et par l’U73122 démontrel’implication aussi des récepteurs P2Y11 dans ce préconditionnement. L’exposition préischémique à des concentrations nanomolaires de PLP protège contre les lésions de l’ I / R.P2Y11 et P2Y6 représentent ainsi les récepteurs candidats les plus probables pour le PCcardiaque induit par le PLP. / Diabetes predisposes to complications affecting various organs such as cardiovascular system.Ischemic heart disease in diabetic patients might be linked to the accumulation ofadvanced-glycation end products (AGEs). In ischemic rat hearts, expression of receptor forAGEs and its ligands is significantly enhanced and involved in cardiac ischemia/reperfusion(I/R) injury even in the absence of diabetes. It has recently been reported that diabetic humanmyocardium cannot be protected by preconditioning. In this context, our hypothesis was thatβ1-adrenergic preconditioning might be altered in the presence of AGEs. Using an isolatednon-working rat heart model, this study investigated the effect of AGEs on cardioprotectioninduced by transient β1-adrenoceptor (β1-AR) stimulation with xamoterol (xa). The beneficialeffects induced by xa during reperfusion were suppressed by the administration of glycatedalbumin (Gly-Alb) during xa infusion, whereas albumin (Alb) did not hamper xa inducedprotection. These results suggest that AGEs suppress the cardioprotection resulting from theactivation of β1-ARs and thus might contribute to cardiovascular damages seen in diabeticpatients.Pyridoxal 5'-phosphate (PLP), a natural metabolite of pyridoxine which is an antagonist ofpurinergic receptors prevents cellular calcium overload and may reduce ischemia-reperfusioninjury. Low plasma pyridoxal-5′-phosphate levels have been observed in patients sufferingfrom myocardial infarction, when compared with a healthy control group. More recently, lowPLP level has been reported to confer an independent risk for coronary artery disease. Thefact that this correlation of PLP reduction in infarct patients has functional importance issupported by the preventive effects of vitamin B6 on coronary heart disease and diabetes. PLPprevented progression of nephropathy in STZ-induced diabetic rats by inhibiting formation ofAGEs. Using an isolated non-working rat heart model, this study investigated the mechanismsof pharmacological pre-conditioning (PC) induced by P2Y receptor stimulation withpyridoxal-5’-phosphate (PLP). The suppression of the cardioprotective effects of PLP byMRS2578 and U73122 is in agreement with the P2Y6 receptor as a receptor for PLP-inducedPC.The suppression of the cardioprotective effects of PLP by AMPαS, the PKA inhibitor (H89),and (U73122) is in agreement with the P2Y11 receptor as a receptor for PLP-induced PC.Pre-ischemic exposure to nanomolar concentrations of PLP is protective against I/R. P2Y11and P2Y6 represent the most likely candidate receptors for PLP-induced cardiac PC.

Introdução: A influência de variantes genéticas dos genes da visfatina e dos recep- tores ?2-adrenérgicos em resposta ao treinamento físico é ainda inconclusiva. Obje- tivos: Verificar os efeitos do treinamento com danças afro-brasileiras por 13 sema- nas sobre parâmetros de saúde de crianças com sobrepeso ou obesidade e a in- fluência das variantes genéticas acima descritas na magnitude de resposta a este treinamento. Materiais e métodos: 30 crianças (9 ± 1,1 anos) realizaram um trei- namento que consistia em 5 minutos de aquecimento a 60% da FCmáx, quatro mo- mentos dez minutos de 70% a 80% da FCmáx intercalados com cinco momentos de dois minutos de recuperação ativa a 60% da FCmáx, 3x/sem, 60 min por sessão. Antes e após o treinamento realizou-se avaliação da composição corporal, índice de massa corporal (IMC), z- score do IMC, circunferência da cintura (CC), relação cintu- ra/estatura (RCE), pressão arterial sistólica (PAS) e diastólica (PAD), capacidade aeróbia e analise da variabilidade da frequência cardíaca (VFC). Análises sanguí- neas foram realizadas para genotipagem, avaliação do perfil lipídico, glicemia, insu- lina e HOMA-IR. A análise estatística foi realizada utilizando modelo de regressão de efeitos mistos. Resultados: Após o treinamento com dança afro-brasileira houve redução significativa (p<0,05) no z-score do IMC (-6,6%), na relação cintura/estatura (-4,8%) e consumo calórico (-15,3%). Considerando os genótipos dos polimorfismos estudados houve uma diminuição significativa no valor do z-score do IMC (-10%) para o genótipo AG do gene do receptor ?2-adrenérgico Arg16Gly, mas nenhuma alteração significativa para os genótipos relacionados aos genes da visfatina e Gln27Glu do receptor ?2-adrenérgico. Conclusão: O treinamento com dança afro- brasileira foi uma estratégia com efeito positivo sobre o score-z do IMC e relação cintura/estatura em crianças com sobrepeso e obesidade. E o genótipo AG do poli- morfismo do receptor ?2-adrenérgico Arg16Gly apresentou melhor resposta ao trei- namento no z-score do IMC, o que pode indicar uma influência genética à resposta ao treinamento. / Introduction: The influences of NAMPT and ?2-adrenergic receptors polymorphisms in response to dance training remain unclear. Objectives: To verify the effects of dance training on health parameters of overweight or obese children and to verify the influence of the genetic variants previously mentioned in response to 13 weeks of training with African-Brazilian dance. Methods: Thirty children (9 ± 1.1 years) per- formed a training that consisted of 10 minutes at 60% of HRmax, four moments from 70% to 80 % of HRmax interspersed with five minutes of active recovery at 60% of HRmax, in total of 60 minutes of training session. Before and after the training body composition, body mass index (BMI), waist circumference (WC), systolic and diastolic blood pressure (DBP), physical fitness and heart rate variability (HRV) were evaluat- ed. Blood analyzes were performed for genotyping and evaluation of the lipid profile, glucose, insulin and HOMA-IR. Statistical analysis was performed using a general linear mixed effects model. Results: The African-Brazilian dance training resulted in a significant reduction (p<0.05) in BMI z-score (-6.6%), waist-to-height ratio (-4.8%) and caloric intake (-15.3%). In the analysis of each polymorphism, it was possible to observe a significant decrease in the z-score BMI (-10%) for the AG genotype of the Arg16Gly polymorphism in the ?2-adrenergic receptor gene, but there is no signifi- cant difference for the visfatin and Gln27Glu of the ?2-adrenergic receptor polymor- phisms. Conclusion: Afro-Brazilian dance training was a strategy with a positive ef- fect on BMI-z score and waist-to-height ratio in overweight and obese children. In addition, the AG genotype of the ?2-adrenergic receptor Arg16Gly polymorphism presented a better response to training on the BMI z-score, which can suggest a ge- netic influence on the training response.

A hipertensão acomete aproximadamente 1 bilhão de pessoas em todo o mundo, no entanto, a sua etiologia exata é em muitos casos indefinida e o mecanismo de ação molecular não foi elucidado completamente. O óxido nítrico ( ) é liberado de forma contínua nos vasos de resistência, contribuindo para o controle fisiológico da pressão arterial, sendo que a redução em sua biodisponibilidade está correlacionada com o desenvolvimento da hipertensão. Durante muito tempo, acreditava-se que o nitrato e o nitrito eram apenas metabólitos inertes da via do e somente em 1994 foi demonstrado que o nitrito, derivado do nitrato, era um substrato para a produção do no estômago, de modo que a administração de nitrito de sódio exerce efeitos anti-hipertensivos. De fato, há diversas evidências demonstrando os efeitos do nitrito sobre a pressão arterial, porém, não há um consenso sobre o seu mecanismo de ação. Trabalhos recentes sugerem que os efeitos antihipertensivo do nitrito se baseiam na geração de S-nitrosotióis no estômago e na capacidade destas moléculas em nitrosar proteínas da parede vascular, alterando a sua função. Há diversas proteínas da parede vascular com função crítica no controle do tônus vascular que podem ser alvos para a nitrosação. Neste trabalho, optamos por avaliar o processo de nitrosação na sinalização do receptor -adrenérgico em ratos hipertensos e normotensos tratados com nitrito de sódio ou veículo pela via oral por 7 dias. Também foi realizada a incubação de anéis de aorta com nitrito de sódio ou S-nitrosoglutationa na presença ou ausência de ascorbato. O tratamento com nitrito de sódio oral por 7 dias foi responsável por reduzir a pressão arterial sistólica dos animais hipertensos e aumentar as quantidades de nitrito e espécies nitrosadas no plasma. Também foi observado uma redução na reatividade aórtica à fenilefrina em animais tratados com nitrito, sendo que este efeito é abolido na presença de ascorbato. A incubação de anéis de aorta com nitrito de sódio não demonstrou causar efeitos sobre a reatividade vascular, porém, o pré-tratamento com S-nitrosoglutationa seguido da sua remoção foi responsável por reduzir a reatividade aórtica à fenilefrina e aumentar a quantidade de proteínas nitrosadas na aorta. Estes efeitos também foram abolidos pela incubação com ascorbato. Estes resultados nos sugerem que os S-nitrosotióis, e não o nitrito, exercem efeito anti-hipertensivo através da redução da reatividade vascular na aorta, modulando a sinalização do receptor -adrenérgico. Estes efeitos aparentam se basear na nitrosação de proteínas, visto que a incubação com ascorbato é responsável por abolir estes efeitos e há um aumento na quantidade de proteínas nitrosadas na aorta após a pré-incubação com S-nitrosoglutationa / Hypertension affects approximately 1 billion people worldwide, however, its exact etiology is in many cases indefinite and the mechanism of molecular action has not been elucidated completely. Nitric oxide ( ) is released continuously in resistance vessels, contributing to the physiological control of blood pressure, and the reduction in its bioavailability is correlated with the development of hypertension. For a long time, it was believed that nitrate and nitrite were only inert metabolites of the pathway, and it was only in 1994 that nitrite derived from nitrate was shown to be a substrate for the production of in the stomach, so that administration of sodium nitrite exerts antihypertensive effects. In fact, there is a lot of evidence demonstrating the effects of nitrite on blood pressure, but there is no consensus on its mechanism of action. Recent work has suggested that the antihypertensive effects of nitrite are based on the generation of S-nitrosothiols in the stomach and on the ability of these molecules to nitrosate proteins in the vascular wall, altering their function. There are several vascular wall proteins with critical function in vascular tone control that may be targets for nitrosation. In this work, we chose to evaluate the process of nitrosation in -adrenergic receptor signaling in hypertensive and normotensive rats treated with sodium nitrite or vehicle orally for 7 days. Aortic rings were also incubated with sodium nitrite or S-nitrosoglutathione in the presence or absence of ascorbate. Treatment with oral sodium nitrite for 7 days was responsible for reducing the systolic blood pressure of hypertensive animals and increasing the amounts of nitrite and nitrosated species in the plasma. A reduction in aortic reactivity to phenylephrine has also been observed in nitrite treated animals, and this effect is abolished in the presence of ascorbate. The incubation of aortic rings with sodium nitrite was not shown to have an effect on vascular reactivity, however, pre-treatment with S-nitrosoglutathione followed by its removal was responsible for reducing aortic reactivity to phenylephrine and increasing the amount of nitrosated proteins in the aorta. These effects were also abolished by incubation with ascorbate. These results suggest that S-nitrosothiols, not nitrite, exert an antihypertensive effect by reducing vascular reactivity in the aorta, modulating -adrenergic receptor signaling. These effects appear to be based on protein nitrosation, since incubation with ascorbate is responsible for abolishing these effects and there is an increase in the amount of nitrosated proteins in the aorta after preincubation with S-nitrosoglutathione