Glucose-induced insulin secretion is impaired and insulin-induced phosphorylation of the insulin receptor and insulin receptor substrate-1 are increased in protein-deficient rats

Malnutrition is related to diabetes in tropical countries. In experimental animals, protein deficiency may affect insulin secretion. However, the effect of malnutrition on insulin receptor phosphorylation and further intracellular signaling events is not known. Therefore, we decided to evaluate the rate of insulin secretion and the early molecular steps of insulin action in insulin-sensitive tissues of an animal model of protein deficiency. Pancreatic islets isolated from rats fed a standard (17%) or a low (6%) protein diet were studied for their secretory response to increasing concentrations of glucose in the culture medium. Basal as well as maximal rates of insulin secretion were significantly lower in the islets isolated from rats fed a low protein diet. Moreover, the dose-response curve to glucose was significantly shifted to the right in the islets from malnourished rats compared with islets from control rats. During an oral glucose tolerance test, there were significantly lower circulating concentrations of insulin in the serum of rats fed a low protein diet in spite of no difference in serum glucose concentration between the groups, suggesting an increased peripheral insulin sensitivity. Immunoblotting and immunoprecipitation were used to study the phosphorylation of the insulin receptor and the insulin receptor substrate-1 as well as the insulin receptor substrate-1-p85 subunit of phosphatidylinositol 3-kinase association in response to insulin. Values were greater in hind-limb muscle from rats fed a low protein diet compared with controls. No differences were detected in the total amount of protein corresponding to the insulin receptor or insulin receptor substrate-1 between muscle from rats fed the two diets. Therefore, we conclude that a decreased glucose-induced insulin secretion in pancreatic islets from protein-malnourished rats is responsible, at least in part, for an increased phosphorylation of the insulin receptor, insulin receptor substrate-1 and its association with phosphatidylinositol 3-kinase. These might represent some of the factors influencing the equilibrium in glucose concentrations observed in animal models of malnutrition and undernourished subjects.

Diet-induced obesity impairs AKT signalling in the retina and causes retinal degeneration

Retinopathy, a common complication of diabetes, is characterized by an unbalanced production of nitric oxide (NO), a process regulated by nitric oxide synthase (NOS). We hypothesized that retinopathy might stem from changes in the insulin receptor substrate (IRS)/PI3K/AKT pathway and/or expression of NOS isoforms. Thus, we analysed the morphology and apoptosis index in retinas of obese rats in whom insulin resistance had been induced by a high-fat diet (HFD). Immunoblotting analysis revealed that the retinal tissue of HFD rats had lower levels of AKT1, eNOS and nNOS protein than those of samples taken from control animals. Furthermore, immunohistochemical analyses indicated higher levels of iNOS and 4-hydroxynonenal and a larger number of apoptotic nuclei in HFD rats. Finally, both the inner and outer retinal layers of HFD rats were thinner than those in their control counterparts. When considered alongside previous results, these patterns suggest two major ways in which HFD might impact animals: direct activity of ingested fatty acids and/or via insulin-resistance-induced changes in intracellular pathways. We discuss these possibilities in further detail and advocate the use of this animal model for further understanding relationships between retinopathy, metabolic syndrome and type 2 diabetes. © 2012 John Wiley & Sons, Ltd.

Differential phosphorylation, desensitization, and internalization of α1A-adrenoceptors activated by norepinephrine and oxymetazoline

Loss of response on repetitive drug exposure (i.e., tachyphylaxis) is a particular problem for the vasoconstrictor effects of medications containing oxymetazoline (OXY), an α1-adrenoceptor (AR) agonist of the imidazoline class. One cause of tachyphylaxis is receptor desensitization, usually accompanied by phosphorylation and internalization. It is well established that a1A-ARs are less phosphorylated, desensitized, and internalized on exposure to the phenethylamines norepinephrine (NE), epinephrine, or phenylephrine (PE) than are the a1B and a1D subtypes. However, here we show in human embryonic kidney-293 cells that the low-efficacy agonist OXY induces G protein-coupled receptor kinase 2-dependent a1A-AR phosphorylation, followed by rapid desensitization and internalization (∼40% internalization after 5 minutes of stimulation), whereas phosphorylation of α1A-ARs exposed to NE depends to a large extent on protein kinase C activity and is not followed by desensitization, and the receptors undergo delayed internalization (∼35% after 60 minutes of stimulation). Native α1A-ARs from rat tail artery and vas deferens are also desensitized by OXY, but not by NE or PE, indicating that thisproperty of OXY is not limited to recombinant receptors expressed in cell systems. The results of the present study are clearly indicative of agonist-directed a1A-AR regulation. OXY shows functional selectivity relative to NE and PE at a1A-ARs, leading to significant receptor desensitization and internalization, which is important in view of the therapeutic vasoconstrictor effects of this drug and the varied biologic process regulated by α1A-ARs. Copyright © 2013 by The American Society for Pharmacology and Experimental Therapeutics.

Spatial memory in sedentary and trained diabetic rats: Molecular mechanisms

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Treadmill Training Increases SIRT-1 and PGC-1 alpha Protein Levels and AMPK Phosphorylation in Quadriceps of Middle-Aged Rats in an Intensity-Dependent Manner

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Decreased rate of protein synthesis, caspase-3 activity, and ubiquitin-proteasome proteolysis in soleus muscles from growing rats fed a low-protein, high-carbohydrate diet

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Myrcia bella Leaf Extract Presents Hypoglycemic Activity via PI3k/Akt Insulin Signaling Pathway

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Kinome profiling of osteoblasts on hydroxyapatite opens new avenues on biomaterial cell signaling

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Long-term obesity promotes alterations in diastolic function induced by reduction of phospholamban phosphorylation at serine-16 without affecting calcium handling

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Expressão proteica da via PI3K/AKT/mTOR em mastocitomas cutâneos caninos

Pós-graduação em Medicina Veterinária - FCAV

A protein kinase screen of Neurospora crassa mutant strains reveals that the SNF1 protein kinase promotes glycogen synthase phosphorylation

Glycogen functions as a carbohydrate reserve in a variety of organisms and its metabolism is highly regulated. The activities of glycogen synthase and glycogen phosphorylase, the rate-limiting enzymes of the synthesis and degradation processes, respectively, are regulated by allosteric modulation and reversible phosphorylation. To identify the protein kinases affecting glycogen metabolism in Neurospora crassa, we performed a screen of 84 serine/threonine kinase knockout strains. We identified multiple kinases that have already been described as controlling glycogen metabolism in different organisms, such as NcSNF1, NcPHO85, NcGSK3, NcPKA, PSK2 homologue and NcATG1. In addition, many hypothetical kinases have been implicated in the control of glycogen metabolism. Two kinases, NcIME-2 and NcNIMA, already functionally characterized but with no functions related to glycogen metabolism regulation, were also identified. Among the kinases identified, it is important to mention the role of NcSNF1. We showed in the present study that this kinase was implicated in glycogen synthase phosphorylation, as demonstrated by the higher levels of glycogen accumulated during growth, along with a higher glycogen synthase (GSN) ±glucose 6-phosphate activity ratio and a lesser set of phosphorylated GSN isoforms in strain Ncsnf1KO, when compared with the wild-type strain. The results led us to conclude that, in N. crassa, this kinase promotes phosphorylation of glycogen synthase either directly or indirectly, which is the opposite of what is described for Saccharomyces cerevisiae. The kinases also play a role in gene expression regulation, in that gdn, the gene encoding the debranching enzyme, was down-regulated by the proteins identified in the screen. Some kinases affected growth and development, suggesting a connection linking glycogen metabolism with cell growth and development.

Melatonin attenuates Her-2, p38 MAPK, p-AKT, and mTOR levels in ovarian carcinoma of ethanol-preferring rats

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Papel das espécies reativas de oxigênio na hiporreatividade de aortas à fenilefrina observada no final da prenhez de Ratas Espontaneamente Hipertensas (SHR)

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

The PI3K-Akt-eNOS pathway is involved in aortic hyporeactivity to Phenylephrine associated with late pregnancy in spontaneously hypertensive rats

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)