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Biomedical subjects

R Goberna

Publications and source records attributed to R Goberna.

At least 73 records · Page 4Linked to original sources
PubMed1984

The effect of different kinds of refeeding on islet glucose phosphorylating activities.

The aim of the present work has been to study the regulating effect of different kinds of diet on the activities of enzymes that phosphorylates glucose into glucose 6 phosphate in the islets of Langerhans. The metabolism of glucose in the B cell is controlled by two different enzymes, hexokinase and glucokinase, whose activities are lowered during fasting; this coincides with lowered levels of blood glucose and blood insulin and with a blocking of the insulin-secretory response toward glucose. Refeeding with a high-carbohydrate diet restores glucokinase activity in islet extracts, blood insulin, and blood glucose. By contrast, refeeding with a low-carbohydrate diet restores hexokinase activity in islet extracts, restores poorly blood insulin, and is unable to unblock the insulin secretory response toward glucose. These results support the important role that glucokinase plays in the regulation of glycolytic flux and on insulin secretion in the B cell. Hexokinase could play a role in the regulation of the glycolytic flux when the B cell responds to other secretagogues.

Animals↗
PubMed1984

Glucose phosphorylating activities in the islets of Langerhans of the rat. Effect of fasting.

In the present study, the characterization of glucose phosphorylating activities in islet extracts and the effect of fasting on these activities have been studied. 4 mM glucose 6-phosphate strongly inhibits hexokinase activity, while glucokinase increased its activity. N-acetyl-glucosamine inhibits hexokinase activity (40% inhibition), while glucokinase increased its activity (19% increment). Both phosphorylating activities reaches a maximum when the Mg/ATP ratio is 1; increments in Mg/ATP ratio inhibits glucokinase activity while hexokinase activity is less dependent of Mg/ATP ratio. Fasting induces a progressive decrease of both phosphorylating activities in islet extracts. After 48 h fasting both activities are decreased 50%. After 96 h fasting, glucokinase activity disappeared completely, while hexokinase was 80% reduced. Our data suggest that hexokinase is less affected by fasting than glucokinase. In addition, other authors have reported that other glycolytic enzymes are little altered by fasting. Hence the fasting-induced-adaptation of glucokinase could account for the reduced rate of glycolysis and subsequently reduced insulin secretory response towards glucose.

Adenosine Triphosphate↗
PubMed1984

Effect of 2-bromostearate on glucose-phosphorylating activities and the dynamics of insulin secretion in islets of Langerhans during fasting.

Glucose-phosphorylating activity and insulin secretion were measured in homogenates of isolated rat islets and of perfused rat pancreas, respectively. Fasting for 96 h produced a significant decrease of both low-and high-Km glucose-phosphorylating activities and blocked the insulin secretory response to glucose. In the presence of glucose, 0.25 mM 2-bromostearate, a known inhibitor of fatty acid oxidation, partially restored the insulin response to glucose that was lost during fasting. This effect paralleled the restoration of glucose-phosphorylation activities (primarily the high-Km component) seen when islets isolated from 96-h-fasted rats were preincubated with 0.25 mM 2-bromostearate. It is concluded that fasting-induced adaptations of glucose-phosphorylating enzymes could account, at least in part, for the reduced insulin secretory response to glucose. 2-Bromostearate, an inhibitor of fatty acid oxidation, is able to restore both insulin secretory response and glucose-phosphorylating activities, suggesting possible interrelations among the correlated impairment in insulin secretion, islet glucose-phosphorylating activity, islet glucose metabolism, and the oxidation of fatty acids in the B-cell during fasting.

Animals↗
PubMed1983

Compartmentation of calcium in digitonin-disrupted guinea pig pancreatic acinar cells.

The treatment of guinea pig pancreatic acinar cells with digitonin leads to disruption of the plasma membrane, as judged by the liberation of cytosolic enzymes, without significant alteration of the mitochondrial membrane. The transport of calcium by the particulate residue was studied, and two different pools could be distinguished. One was supported by ATP or ADP, succinate providing the respiratory substrate, and was sensitive to the inhibitors, Ruthenium red and azide. The other pool needed the presence of ATP, ADP being ineffective, and also was unaffected by Ruthenium red or by azide, but was stimulated several-fold by oxalate. The Ruthenium red-sensitive calcium pool has characteristics resembling those of the transport of calcium by a mitochondrial fraction prepared from digitonin-treated acinar cells. In contrast, the Ruthenium red-insensitive calcium transport has characteristics resembling those of a microsomal fraction obtained from guinea pig pancreas. When the transport of calcium in digitonized cells was assayed at a calcium concentration range of 10(-8)-10(-4) M, preferential Ruthenium red-insensitive calcium transport could be observed at submicromolar calcium concentrations.

Animals↗
PubMed1982

Stimulating role of potassium ions and ouabain on glycogen synthesis in adipose tissue.

1. Exposure of fat-pads to increasing concentrations of K+ in the presence of insulin stimulates the incorporation of labelled glucose into glycogen. In the absence of hormone, only a slight incorporation of glucose into glycogen and slight glucose oxidation were detectable. 2. Ouabain alone, up to 100 microM, had no effect on synthesis of glycogen. Ouabain reinforced the effect of insulin on the conversion of glucose into glycogen in a Na+ medium and in a equimolar Na+-K+ medium, but not in a K+ medium. In addition, ouabain modified the optimal K+/Na+ ratio for glycogen synthesis. 3. The proportion of glycogen synthase in the active form was increased in a K+ medium, and a faster rate of conversion of synthase b into a was observed under these conditions. No difference was detected in the rate of inactivation of phosphorylase in a K+ or a Na+ medium. 4. Even though these results, taken together, are consistent with the proposed role of phosphorylase a in the regulation of synthase activation, the molecular mechanism of action of K+ in adipose tissue in increasing synthesis of glycogen cannot be explained simply by a faster inactivation of phosphorylase a. It is concluded that some undetermined effector(s) or signal could itself be a primary determinant for the greater activation of synthase observed in a K+ medium.

Adipose Tissue↗
PubMed1982

Biological properties of a calcium-modulator protein from Phycomyces blakesleeanus.

1. A modulator protein having properties similar to calmodulin-like proteins could be detected in Phycomyces the first day of growth. 2. The modulator protein from Phycomyces was partially purified to a specific activity of 9,000 Units/mg protein. 3. The modulator protein was similar to calmodulin with respect to: (a) the ability to stimulate a calcium-plus-calmodulin-dependent cAMP phosphodiesterase from bovine brain; (b) the inhibitory effect of trifluoperazine in this process; (c) the stimulation of the [Ca2+ + Mg2+)-ATPase from human erythrocyte membranes. 4. Maximum effect of modulator protein in the above mentioned reactions was achieved at micromolar calcium concentrations.

3',5'-Cyclic-AMP Phosphodiesterases↗
PubMed1981

Interaction of vasoactive intestinal peptide with a cell line (HeLa) derived from human carcinoma of the cervix: binding to specific sites and stimulation of adenylate cyclase.

The binding of vasoactive intestinal peptide (VIP) and its effect on cyclic AMP production were assessed in HeLa cells. The binding of [125I]VIP is a moderately rapid process, reversible, saturable, specific and dependent on temperature. Virtually no inactivation of the peptide is observed after 2 h of exposure to the cells. At 15 degrees C, the binding data obtained at steady state are compatible with the existence of two classes of binding sites: a first class with a Kd of 2.4 nM and low binding capacity (1.5 X 10(5) sites/cell) and a second class with a Kd of 100 nM and a high binding capacity (4.9 X 10(6) sites/cell). Secretin is eight times less potent than VIP in competing with 125I VIP but glucagon, insulin and somatostatin are inactive. VIP-induced stimulation of cyclic AMP production depends on time and temperature and is potentiated by a phosphodiesterase inhibitor. A concentration of VIP as low as 10(-10) M is able to stimulate adenylate cyclase. Half-maximal stimulation is observed at 10(-9) M and maximal stimulation (4 times above basal levels) at 10(-8) M VIP. Secretin is an agonist of VIP but exhibits a 1000 times lower potency with respect to adenylate cyclase activation. Glucagon, insulin and somatostatin do not show any effect. The presence of high-affinity binding sites and high sensitivity and specificity of adenylate cyclase for VIP in HeLa cells provide a good model to study the role of this peptide on cell proliferation and differentiation.

Adenylyl Cyclases↗
PubMed1981

Interaction of vasoactive intestinal peptide with human blood mononuclear cells.

The binding of vasoactive intestinal peptide (VIP) and the stimulation of adenylate cyclase were studied in mononuclear cells from human peripheral blood. The binding depended on time, temperature and pH, and was reversible, saturable and specific. Binding studies suggested the presence of 2 classes of binding site: a class with high affinity (Kd = 2.4 X 10(-10)M) and low capacity (8 fmoles/10(6) cells), and a class with low affinity (Kd = 8.0 X 10(-8)M) and high capacity (800 fmoles/10(6) cells) at 15 degrees C. Secretin displaced [125I]VIP from the cells with a 400-fold lower affinity than VIP, but glucagon, somatostatin and insulin did not show any effect. VIP was a potent and efficient stimulator of cyclic AMP production. The stimulation was observed at a concentration as low as 3 X 10(-11)M and depended on time, temperature and pH. Maximal cyclic AMP production (4-fold above basal levels) was observed with 10(-9) M at 15 degrees. Half-maximal response was obtained at 10(-10)M VIP. Secretin was an agonist of VIP but exhibited a 7000 times lower potency. Peripheral blood mononuclear cells constitute an easily accessible and suitable system for the study of VIP action in different physiological and pathophysiological conditions.

Adenylyl Cyclases↗
PubMed1981

Somatostatin action on insulin secretion induced by chicken and porcine vasoactive intestinal peptide in the perfused rat pancreas.

The isolated perfused rat pancreas with duodenal exclusion was used to study the stimulation of glucose-induced insulin release in response to chicken and porcine vasoactive intestinal peptide (VIP). The insulin response to 5.5 or 16.7 mM glucose was markedly enhanced by 750 pM porcine VIP and a concentration of 250 pM was still effective. At 250 pM, chicken VIp exhibited a slightly higher potency than porcine VIP at both glucose concentrations. The main difference between the two peptides was that the effect of porcine VIP disappeared immediately after the peptide suppression but tha of chicken VIP persisted for an additional period of 8-10 min. Somatostatin (10 ng/ml) blocked the stimulatory effect of both VIP molecules on glucose-induced insulin secretion. After suppression of VIP and somatostatin from the perfusion medium, insulin release increased to levels higher than those with glucose alone in the case of the avian peptide, but not in that porcine VIP. The data are consistent with previous results in the literature on stimulation of exocrine pancreas secretion and interaction with intestinal epithelium.

Animals↗
PubMed1981

Effects of oxaloacetate and beta-hydroxybutyrate on rat liver mitochondrial calcium flow during starvation.

Rat liver mitochondria accumulate calcium from the incubation medium both in the presence of ATP and of succinate with rotenone. Lowering free calcium concentration by means of EGTA leads to a reduction of maximum transport capacity to less than 25% of the total added calcium. Under these conditions addition of oxaloacetate inhibits calcium uptake, an effect which is blocked by equimolar concentrations of beta-hydroxybutyrate. Oxaloacetate and acetoacetate induce an efflux of previously accumulated mitochondrial calcium. These effects appear to be independent of free calcium concentration and of whether the mitochondria were obtained from fed rats or rats fasted for 16 h or 4 days.

Acetoacetates↗
PubMed1981

Effects of fasting and refeeding on insulin binding to liver plasma membranes and hepatocytes from normal rats.

The effect of acute and chronic starvation on insulin binding to liver cells and liver cell membranes was studied in normal rats. A short-term (1 day) fasting period did not induce significant changes in insulin binding to liver plasma membranes in spite of the hypoinsulinemia present in the rats, as compared with normo-insulinemic fed controls. Longer periods of fasting (2 and 4 days) elicited an increase in insulin binding to liver membranes that was due to an increase in the number of insulin receptors rather than to changes in affinity. All groups of membranes were similar with respect to the rates of association and dissociation of hormone, degree of negative cooperativity and extents of both hormone and receptor inactivation. The fasting-induced increase of insulin binding was not a consequence of the methodological procedure for membrane isolation since the same conclusions were obtained when intact hepatocytes were used. Refeeding of fasted animals resulted in a decrease of insulin binding together with an increase of insulinemia towards the fed control values. These results suggest that the number of insulin receptors is inversely related to the circulating levels of the hormone and support the hypothesis of insulin regulating its own receptor.

Animals↗
PubMed1981

Properties of vasoactive intestinal peptide-receptor interaction in rat liver membranes.

The properties of the specific receptors for vasoactive intestinal peptide (VIP) in rat liver plasma membranes have been studied by using 125I-VIP as a tracer. The binding of the peptide was a reversible, saturable and specific process, as well as time and temperature dependent. Peptide inactivation was also dependent on time and temperature and remained relatively low in the standard conditions used, as it happened in the inactivation of the binding sites. The binding data were compatible with the existence of two classes of VIP receptors: a high affinity (Kd = 4.2 x 10(-10) M) and low binding capacity (1.5 pmol VIP/mg protein) class and another one of low affinity (Kd = 1.7 x 10(-7) M) and high binding capacity (38.6 pmol VIP/mg protein). The specificity of the binding sites of VIP was established from the fact that binding of 125I-VIP was inhibited by native VIP and by 60-fold higher concentrations of secretin but not by the parent hormone glucagon, by insulin or somatostatin at concentrations as high as 10(-6) M.

Animals↗
PubMed1981

Guanyl nucleotide regulation of vasoactive intestinal peptide interaction with rat liver membranes.

This study shows the regulatory role of guanyl nucleotides on vasoactive intestinal peptide (VIP) interaction with rat liver plasma membranes. The binding of 125I-VIP to the membranes was partially inhibited by GTP and the GTP-analog Gpp(NH)p in a dose-dependent manner. This effect was mainly due to a dramatic increase of the dissociation rate of the complex tracer-membranes in the presence of guanyl nucleotides. The specificity of the binding inhibition was assessed from the lack of action of the other purine nucleotides tested. VIP stimulated cyclic AMP production in liver plasma membranes in the range 10(-10) -- 3 x 10(-7) M. Half-maximal stimulation was observed at 3 x 10(-10) M and maximal stimulation (4-fold basal value) at 10(-9) M VIP. Both GTP and Gpp(NH)p potentiated VIP-stimulated cyclic AMP production since the effect of nucleotide plus VIP was greater than the sum of the effects produced by the two agents separately. Therefore, guanyl nucleotides simultaneously inhibit the binding of VIP to its receptors and potentiate the stimulatory effect of VIP on adenylate cyclase activity in rat liver plasma membranes.

Adenylyl Cyclases↗
PubMed1981

Cyclic AMP-dependent protein kinase activity and lipolysis in adipose tissue. Effect of fasting, oligomycin and iodoacetamide.

The release of glycerol into the medium, the concentration of cAMP, and the cAMP-dependent protein-kinase activity were studied in adipocytes and in fat-pads obtained from epididymal adipose tissue of rats under different conditions of feeding. An increase in the tissue concentration of cAMP and in the protein-kinase activity was observed in vivo at 48 and 96 h of fasting. A diminished release of glycerol was found in adipocytes from rats fasted for 48 h, in the absence of glucose, and the maximum concentration of cAMP was inferior to that of fed rats. Oligomycin and iodoacetamide, in the presence of epinephrine and glucose, produce a diminution in the values of the parameters studied. No significant differences were observed, however, in the responses of tissue obtained from fed and fasting rats to these compounds. The present results confirm previous observations and show the dependence of the lipolytic process on carbohydrate metabolism.

Adipose Tissue↗
PubMed1980

Restoring effects of refeeding and dibutyril cyclic AMP on the increased glucagon secretion of rat pancreatic islets during starvation.

The glucagon release in the presence of glucose and the interaction of dibutyril cyclic AMP was studied in isolated pancreatic islets from fed and 96 h-fasted rats incubated for 30 min. In both states the increase of glucose concentration produced a similar inhibition of glucagon release and stimulation of cyclic AMP content. Higher glucagon secretion and lower cyclic AMP contents were observed in islets from fasted than in those from fed animals at both 2.75 and 16.7 mM glucose. Islets from rats starved for 96 h and refed for 48 h with normal diet or oral 20% glucose showed glucagon release patterns similar to those of controls. Addition of 2 mM dibutyril cyclic AMP to the incubation medium inhibited the fasting-induced increase of glucagon secretion at both glucose levels. These results show an inverse correlation between glucagon secretion and islet cyclic AMP content. However, the sensitivity of the islet to glucose-induced inhibition of glucagon secretion appears to be independent of the islet cyclic AMP levels.

Animals↗