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Biomedical subjects

J Sehlin

Publications and source records attributed to J Sehlin.

At least 109 records · Page 6Linked to original sources
PubMed1979

45Ca2+ uptake by dispersed pancreatic islet cells: effect of D-glucose and the calcium probe, chlorotetracycline.

Uptake of 45Ca2+ was studied in dispersed pancreatic islet cells from non-inbred ob/ob-mice. Like whole islets the dispersed cells responded to 20 mM D-glucose with a markedly increased 45Ca2+-labeling of both the lanthanum-nondisplaceable and the lanthanum-displaceable calcium pools. The pronounced effect of D-glucose could not be reproduced with 3-O-methyl-D-glucose, L-glucose, D-mannose, L-leucine, or D-leucine; however, 45Ca2+ uptake was greater in the presence of L-leucine as compared with D-leucine. 45Ca2+ uptake by dispersed cells or whole islets was stimulated severalfold by 100 microM or more chlorotetracycline. At the concentration of only 10 microM, chlorotetracycline had no effect on whole islets and partially inhibited 45Ca2+ uptake by the dispersed cells. The ability of D-glucose to stimulate 45Ca2+ uptake by islets or dispersed cells remained in the presence of 10 microM chlorotetracycline. Islet cell suspensions apparently represent a valid model for studying how Ca2+ interacts with the cells. However, when using chlorotetracycline as fluorescent Ca2+ probe, attention must be paid to it potential ionophoric activity. At only 10 microM, the drug seems to monitor a peripheral pool of Ca2+, some of which may reside in normal transport channels.

Animals↗
PubMed1978

Effect of Na+, K+ and Mg2+ on 45Ca+ uptake by pancreatic islets.

Microdissected pancreatic islets of noninbred ob/ob-mice were used to study ionic effects on the lanthanum-nondisplaceable 45Ca2+ uptake by islet cells. Omission of Mg2+ from the incubation medium had no effect, but the 45Ca2+ uptake was increased by omission of Na+ and decreased by omission of K+. Excess Mg2+ (1.2--15 mM) inhibited and excess K+ (4.7--25 mM) stimulated the 45Ca2+ uptake in a concentration-dependent manner. Stimulation of 45Ca2+ uptake in Na+-deficient islets was associated with an enhancement of the basal insulin release. Total abolishment of glucose-stimulated 45Ca2+ uptake in K+-deficient islets did not preclude a significant secretory reponse to glucose. It is concluded that the lanthanum-nondisplaceable 45Ca2+ uptake shows a partial correlation to insulin release.

Animals↗
PubMed1978

Possible toxic effects of normal and diabetic patient serum on pancreatic B-cells.

Serum from normal blood-donors and juvenile diabetic patients inhibited Rb+ accumulation and stimulated release of 51Cr and insulin in suspensions of dispersed pancreatic islet cells prepared from ob/ob mouse islets, which are rich in B-cells. The effects indicate the presence of a B-cytotoxic factor in human serum. Serum from mouse and fetal calf also inhibited the islet cell accumulation of Rb+. Toxicity was not suppressed by treating serum with protein A-Sepharose and did not correlate with islet cell binding of fluorescent antibodies to human immunoglobulin. Whereas all sera inhibited Rb+ accumulation, 3 of 6 diabetic patient sera, but no blood-donor serum, made the cells fluoresce on exposure to the fluorescent antibodies. Supporting a dependence on complement, toxicity remained after dialysis, but was destroyed by treating serum with zymosan-A or heating at 56 degrees for 30 min.

Animals↗
PubMed1978

Metabolism of cold-stored pancreatic islets.

A previous study showed that the ability of glucose to stimulate insulin release was retained in islets stored at 8 degrees C for one week provided that glucose was present in a high concentration in the storage medium. The metabolic properties of islets stored in the cold have now been further explored in an attempt to clarify the protective effect of glucose. During storage in the cold the islet formation of 3H2O from (5--3H) glucose and oxygen consumption were only a few per cent of that of fresh islets whereas the putake of 86Rb+ was 20--48%. Rewarming the cold-stored islets to 37 degrees C after one week of cold-storage restored the 86Rb+ uptake, the formation of 3H2O and 14CO2 from labelled glucose and oxygen consumption to 75, 80, 60 and 40% respectively of fresh islet levels. The results emphasize the usefulness of cold-storage for preservation of functionally intact isolated islets.

Animals↗
PubMed1978

86Rb+ fluxes and K+-stimulated nitrophenyl phosphatase activity in the pancreatic islets of genetically diabetic mice (C57BL/KsJ-db/db).

Fluxes of 86Rb+ and hydrolysis of p-nitrophenyl phosphate were measured in collagenase-isolated islets of diabetic C57BL/KsJ-db/db-mice and normal controls (C57BL/KsJ-+/+). Both types of islets accumulated Rb+ avidly, as originally reported for hand-dissected islets of non-inbred ob/ob-mice. KsJ-db/db-mouse islets showed enhanced accumulation of Rb+ and normal activity of K+-activated nitrophenyl phosphatase. D-glucose, 20 mmol/l, inhibited Rb+ efflux in normal islets but not in those from KsJ-db/db-mice. The glucose insensitivity of Rb+ efflux was observed in young animals, which exhibit glucose-induced insulin release, as well as in old animals, which do not secrete insulin in response to glucose. The anomalous regulation of Rb+ efflux already present in young animals may bear on the liability of KsJ-db/db-mouse B-cells to develop defective control of membrane potential, an abnormal metabolism of cyclic AMP, and a marked failure of insulin secretory capacity.

4-Nitrophenylphosphatase↗
PubMed1978

Interrelationship between chloride fluxes in pancreatic islets and insulin release.

The role of Cl- in the function of pancreatic beta-cells was studied by using islets of noninbred ob/ob mice. 36Cl- was rapidly taken up by islet cells; apparent isotope equilibrium was reached within 30 min. The apparent distribution ratio was 0.50--0.72 in N-2-hydroxyethylpiperazine-N'-2-ethanesulfonic acid (HEPES) medium and about 1.1 in Krebs-Henseleit medium. Uptake of 36Cl- was inhibited by 2,4-dinitrophenol, increased by ouabain, and not affected by omission of K+, Na+, or Ca2+. D-Glucose increased short-term uptake of 36Cl- but decreased the equilibrium content. Efflux of 36Cl- from prelabeled islets approached first-order kinetics with a half-life of about 5 min, was inhibited by 4-acetamido-4'-isothiocyanato-stilbene-2,2'-disulfonic acid or low temperature, was stimulated by D-glucose, D-mannose, or hydronium ion, and was unaffected by L-glucose or 3-O-methyl-D-glucose; D-manno-heptulose abolished the effect of D-glucose. Insulin secretion in response to D-glucose was reversibly inhibited in Cl- -deficient media. It is suggested that Cl- is nonpassively distributed across the beta-cell plasma membrane. D-Glucose-induced depolarization of beta-cells may partly be mediated by an increase of the Cl- permeability.

Animals↗
PubMed1978

Further studies on the relationship between insulin release and lanthanum-nondisplaceable 45Ca2+ uptake by pancreatic islets: effects of fructose and starvation.

Relationships between the release of insulin and the incorporation of 45Ca2+ into a lanthanum-nondisplaceable (intracellular) pool were studied in islets microdissected from the pancreatic glands of non-inbred ob/ob mice. In comparison with D-glucose, D-fructose was slowly oxidized and had only marginal effects on insulin release. However, fructose was as effective as glucose in stimulating the lanthanum-nondisplaceable 45Ca2+ uptake. The 45Ca2+ uptake was dose-dependent on the concentration of fructose in the range 0-20 mM; the same dose-dependence was obtained with glucose. Fasting the mice for 3 days caused a total block of the insulin secretory response to 20 mM glucose, but it produced an enhancement of the glucose-induced 45Ca2+ uptake. Both the inhibition of insulin release and the enhancement of 45Ca2+ uptake were counteracted by pretreating the isolated islets with 20-40 mM D-glucose; pretreatment with L-glucose or fructose could not counteract the effects of fasting. Although some functional relationship may exist between the lanthanum-nondisplaceable uptake of 45Ca2+ and the insulin secretory apparatus, it is concluded that the uptake of Ca2+ is not simply the result of stimulated insulin release.

Animals↗
PubMed1978

Effects of valinomycin on Rb+ fluxes, ATP content and insulin release in pancreatic islets.

Valinomycin, 0.5-500 nM, was tested for its effects on pancreatic islets microdissected fron non-inbred ob/ob-mice. Valinomycin decreased the islet accumulation Rb+ and the content of ATP in a dose-dependent manner; efflux of Rb+ from pre-loaded islets was not noticeably changed. Rb+ accumulation and ATP content correlated markedly; on the model of linear regression, less than 10% of the change Rb+ accumulation in valinomycin-treated islets was statistically attributable to factors other than ATP. Valinomycin did not cause a prompt inhibition of glucose-stimulated insulin release that could reflect hyperpolarization due to increased K+ permeability. The following conclusions are drawn: 1) The plasma membranes of beta-cells resemble those of neurons in having such a high ion permeability as to be relatively little influenced by valinomycin; 2) Islet accumulation of Rb+ is due to a vectorial catalyst in theplasma membrane rather than to uptake by mitochondria; 3) Rb+ accumulation in islets is ATP-dependent.

Adenosine Triphosphate↗
PubMed1977

Alloxan cytotoxicity in vitro. Inhibition of rubidium ion pumping in pancreatic beta-cells.

Exposing micro-dissected pancreatic islets of non-inbred ob/ob mice to 2-5 mM-alloxan for 10 min decreased the ability of the islets to accumulate Rb+. Rb+ accumulation in pieces of exocrine pancreas was unaffected by alloxan. When islets were treated with alloxan in the presence of 2-20 mM-D-glucose, the Rb+-accumulating ability was protected in a dose-dependent manner. The protective action of D-glucose was reproduced with 3-O-methyl-D-glucose but not with L-glucose or D-mannoheptulose; mannoheptulose prevented D-glucose from exerting its protective action. The inhibition of Rb+ accumulation was due to a decreased inward pumping, since alloxan did not affect Rb+ efflux from pre-loaded islets. The inhibitory effect of alloxan had a latency of about 1 min, as revealed by experiments with dispersed islet cells in suspension. Alloxan-treated islets showed only a marginal decrease in ATP and no change in glucose 6-phosphate concentration. Although alloxan slightly decreased the hydrolysis of ATP in a subcellular fraction enriched in plasma membranes, this effect could not be attributed to a ouabain-sensitive adenosine triphosphatase. The plasma membranes exhibited a K+-activated hydrolysis of p-nitrophenyl phosphate; this enzyme activity too was insensitive to alloxan. Glucose may protect the univalent-cation pump by preventing permeation of alloxan via a path coupled to the hexose-transport system. Inhibition of the pump may be fundamental to the induction of alloxan-diabetes.

Adenosine Triphosphate↗
PubMed1977

Effects of various modifiers of insulin release on the lanthanum-nondisplaceable 45Ca2+ uptake by isolated pancreatic islets.

The uptake of 45Ca2+ by a lanthanum-non-displaceable pool in pancreatic islets was studied; Raising the extracellular D-glucose concentration from 3 to 20 mM stimulated the 45Ca2+ uptake in hand-dissected islets of ob/bo-mice as well as in collagenase-isolated islets of ob/ob or normal mice. The effect was dose-dependent in the range of 0-20 mM D-glucose and was seen throughout a wide range of extracellular calcium concentrations (16 mumol-2.56 mmol of Ca2+ added per litre of medium). The 45Ca2+ uptake was also enhanced by other known insulin secretagogues (D-mannose, L-leucine, tolbutamide) and was uninfluenced by compounds lacking insulin-releasing capacity (3-O-methyl-D-glucose, L-glucose, D-galactose, D-leucine). The stimulatory effect of D-glucose was blocked by inhibitors of glucose-induced insulin release (D-mannoheptulose, diazoxide, L-adrenaline). The results support the view that the lanthanum-nondisplaceable calcium pool is related to the insulin-releasing mechanism, although the exact nature of this relationship is still unclear.

Animals↗
PubMed1977

Glucose-stimulated and La3+-nondisplaceable Ca2+ pool in pancreatic islets.

To study intracellular pools of calcium tissue specimens from noninbred ob/ob mice were labeled with 45Ca2+ and subsequently washed with La3+. D-glucose, 20 mM, enhanced the labeling of the La3+-nondisplaceable calcium in pancreatic islets but not in pieces of exocrine pancreas or liver. The disappearance of 45Ca2+ from labeled islets was accelerated by the ionophore, X-537A, but not by dibutyryl cyclic AMP, theophylline, or pentobarbital. On fractionation of 45Ca2+-labeled islets, the greatest radioactivity per unit of protein occured in a fraction rich in insulin secretory granules. The radioactivity of this fraction was higher after the islets had been loaded with 45Ca2+ in the presence of 20 mM glucose as compared to 3 mM glucose. It is concluded that the secretory granules make up a considerable part of the glucose-sensitive calcium store in the beta-cells.

Animals↗
PubMed1976

Calcium and secretion: distinction between two pools of glucose-sensitive calcium in pancreatic islets.

D-Glucose, but not L-glucose or 3-O-methyl-D-glucose, stimulates 45Ca2+ uptake by both lanthanum-displaceable and lanthanum-nondisplaceable pools in pancreatic islets. The nondisplaceable pool probably represents secretory granules, while the displaceable pool may be located in the beta-cell membrane. Kinetic studies with isotopically labeled islets suggest that only the displaceable pool participates in the short-term coupling of the glucose stimulus with secretion.

Animals↗
PubMed1976

The dynamics of insulin release from mouse pancreatic islet cells in suspension.

The overall dynamics of glucose-induced insulin release was strikingly similar in dispersed cells and intact islets perifused in parallel. Both preparations exhibited a latency of 1-2 min, after which period there was a brisk rise of insulin release followed by a sustained second phase. During the second phase, insulin release from dispersed cells attained a stable plateau rate, whereas the release from intact islets continued to rise. Epinephrine (1 muM) inhibited the release in both preparations, but the return to basal rate was faster in the dispersed cells than in the intact islets. The dispersed cells oxidized glucose at a constant rate for at least 60 min; the glucose oxidation was markedly sensitive to changes of the glucose concentration in the range of 3-20 mM.

Animals↗
PubMed1976

Ionic effects on the uptake of sulfonylurea (glibenclamide) by pancreatic islets.

The accumulation of glibenclamide was studied in the pancreatic islets of non-inbred ob/ob-mice. Microdissected islets were incubated in media of different ionic composition and the accumulation measured as the uptake of drug not accounted for by equilibration in the urea space. In 12 mM N-hydroxyethylpiperazine-N'-2-ethane sulphonic acid (HEPES) buffer the accumulation was only half of that in Krebs-Ringer bicarbonate buffer. Addition of 50 mM NaCl, Na2SO4, KCl, or LiCl to the HEPES buffer restored the accumulation to the level seen in Krebs-Ringer buffer. Studies with different concentrations of NaCl and Na2SO4 showed that the glibenclamide accumulation was sensitive to Na+ with little or no effect being attributable to the anions. CaCl2 or MgCl2 had much stronger effects than NaCl. It is concluded that cations participate in the binding of glibenclamide to beta-cells. The mechanism could be an increased ionic attraction between cell surfaces and hydrophilic regions of the drug, or a shielding of fixed surface anions allowing stronger hydrophobic interactions between drug and beta-cells.

Animals↗
PubMed1976

Interactions between the metabolism of L-leucine and D-glucose in the pancreatic beta-cells.

Beta-Cell-rich pancreatic islets microdissected from obese-hyperglycemic mice were used to study interactions between the metabolism of L-leucine and D-glucose. L-leucine reduced the islet content of aspartic acid whereas D-glucose, when added to L-leucine-incubated islets, increased the contents of aspartic acid and gamma-aminobutyric acid (GABA). D-glucose also increased the incorporation of L-leucine carbon into aspartic acid, GABA and glutamic acid suggesting stimulation of a malate shuttle mechanism. When expressed per mole of the individual amino acids, the incorporation of L-leucine carbon into GABA was 2.5-4 times higher than into glutamic acid indicating intracellular compartmentation of the latter amino acid. Both L-leucine and D-leucine stimulated 14CO2 production from 14C-labelled D-glucose. L-leucine did not affect 3H2O production from tritiated D-glucose. The present data do not indicate a role of other amino acids or D-glucose in L-leucine-stimulated insulin release.

Animals↗
PubMed1976

Effects of glucose on 45Ca2+ uptake by pancreatic islets as studied with the lanthanum method.

1. Fluxes of 45Ca2+ were studied in pancreatic islets from non-inbred ob/ob-mice. Because La3+ blocked the transmembrane fluxes of 45Ca2+ in islet cells, incubations aimed at measuring glucose-induced changes of the intracellular Ca2+ were ended by washing the islets with 2 mM-La3+ for 60 min. 2. Uptake of 45Ca2+ progressed for 2 hr; the intracellular concentration of exchangable Ca2+ was about 7 m-mole/kg dry wt., as estimated from the isotope distribution at apparent equilibrium in islets exposed to 3 mM D-glucose. Raising the D-glucose concentration to 20 mM enhanced the 45 Ca2+ uptake whether or not the islets had first been equilibrated with the isotope. The stimulatory effect of D-glucose was observed in Tris buffer containing no anions but Cl- as well as in polyanionic bicarbonate buffer. The effect could not be reproduced with equimolar L-glucose. 3. The rate of 45Ca2+ release was the same whether the islets had been pre-loaded in the presence of 3 or 20 mM D-glucose. Thus the 45Ca2+ that had been taken up in response to 20 mM D-glucose appeared to be released much more slowly than the bulk of intracellular 45Ca2+. The release of 45Ca2+ was not significantly influenced by D-glucose during the release period. Incubation for 30 min was require for half of the radioactivity to be released. 4. The rates of insulin secretion were about the same in uni-anionic Tris buffer as in polyanionic bicarbonate buffer. A marked insulin secretory response to 20 mM D-glucose was observed in either buffer. 5. It is concluded that 20 mM D-glucose causes a net uptake of Ca2+ from the extracellular fluid into the interior of the beta-cells. This uptake is probably not regulated at the level of the plasma membrane but more likely reflects an increased affinity of some intracellular phase or compartment for the ion. Because the observed uptake and release of intracellular 45Ca2+ are slow processes in comparison with the rapid effects of extracellular Ca2+ on insulin secretion, insulin secretion may also depend on a more superficial and La3+-displacable Ca2+ pool.

Animals↗
PubMed1976

On the possible role of thiol groups in the insulin-releasing action of mercurials, organic disulfides, alkylating agents, and sulfonylureas.

The thiol activity of pancreatic islets was spectrophotometrically assayed as the formation of 6-mercaptonicotinic acid from the organic disulfide, 6,6'-dithiodinicotinic acid. Islets containing more than 90% beta-cells were microdissected from non-inbred ob/ob-mice. Comparisons of intact with homogenized islets indicated that the organic disulfide penetrates relatively slowly into the beta-cells. When tested at concentrations know to enhance insulin release, p-chloromercuribenzene-sulfonic acid almost completely blocked the thiol activity of intact islets, whereas no significant effect was observed with iodoacetamide, D-glucose, or glibenclamide. Although glibenclamide had no demonstrable effect on the thiol activity of free L-cysteine, the binding of glibenclamide to serum albumin was decreased by blocking the albumin thiols with azobenzene-2-sulfenyl bromide. The uptake of glibenclamide by pancreatic islets was inhibited by cysteine or reduced glutathione. Cysteine, as well as 6,6'-dithiodinicotinic acid, also seemed to interact negatively with glibenat organic mercurials and disulfides stimulate insulin release by blocking thiol groups in the beta-cell plasma membranes. The thiol groups involved in iodoacetamide-induced secretion may escape detection by the assay employed, or target groups other than thiols may be involved. The data on glibenclamide are compatible with, but do not unequivocally support, the notion that thiol groups may play a role in sulfonylurea-induced insulin release.

4-Chloromercuribenzenesulfonate↗