PubMed Health⌕ Search

Biomedical subjects

F M Matschinsky

Publications and source records attributed to F M Matschinsky.

At least 55 records · Page 3Linked to original sources
PubMed1994

Protein databases for compacted eight-cell and blastocyst-stage mouse embryos.

High-resolution two-dimensional sodium dodecyl sulfate-polyacrylamide (2D-SDS) gel electrophoresis combined with computerized analysis of gel images was used to construct and analyze protein data-bases for two stages of preimplantation mouse embryogenesis, the compacted eight-cell stage and the fully expanded blastocyst stage. These stages were chosen for their ease in identification of multiple synchronous embryos. Synchronous cohorts of 30-50 embryos were labelled with L-[35S]methionine for 2 hr. The embryos were then lysed in 30 microliters hot SDS sample buffer, and the lysates were stored at -80 degrees C until the gels were run. Five replicates were run for eight-cell embryos, and four for blastocyst-stage embryos. The samples were processed for 2D gel electrophoresis and fluorography; multiple exposures were made. Gel images were analyzed using the PDQUEST system, and databases were constructed. Analysis of the databases for both developmental stages showed high reproducibility of protein spots in multiple gel images. Of 1,674 total spots in eight-cell embryo standards, > 79% of spots had a percentage error (S.E.M./average) < 50%, and > 45% had a percentage error < 30%. Similarly, of 1,653 total spots in blastocyst-stage embryo standards, 74% of spots had a percentage error < 50%, and approximately 47% of spots had a percentage error < 30%. Forty-three spots (approximately 3% of the total spots) were found to be detected only in the eight-cell stage, while 75 spots were detected solely in the blastocyst stage. Sixty-nine proteins showed a greater than threefold increase in isotope incorporation from the eight-cell to the blastocyst stage, with a percentage error < 50% in both the eight-cell and the blastocyst stages. In contrast, 41 of the proteins showed a decrease during this period. Analysis of the protein databases described in this study has allowed us to document the overall quantitative changes in proteins from the compacted eight-cell stage to the blastocyst stage of mouse preimplantation development. These data-bases provide a valuable tool for further detailed quantitative analysis of specific proteins associated with developmental events. In addition they will permit analysis of the effects of environmental factors, such as growth factors, on early embryo development.

Animals↗
PubMed1994

Insulin family growth factors have specific effects on protein synthesis in preimplantation mouse embryos.

Previously constructed protein databases for two stages of preimplantation mouse embryogenesis, the compacted eight-cell stage and the fully expanded blastocyst stage, have been used to analyze the effects of insulin, IGF-I, and IGF-II on protein synthesis in these developmental stages. Proteins were labeled by placing, for 2 hr, synchronous cohorts of 35-50 embryos into human tubal fluid (HTF) medium containing L-[35S]-methionine (1 mCi/ml) in the presence or absence of one of the growth factors. The embryos were then washed with medium and lysed. Samples were processed for 2-D gel analysis. For each embryonic stage and each growth factor, four or five experimental replicates were done and the gel images were compared using the PDQUEST system. Using the computer-assisted analysis, we were able to identify proteins that showed a statistically significant (P < 0.05) change in synthesis. At the eight-cell stage of development insulin caused increased synthesis of two proteins and decreased synthesis in three proteins. Insulin-treated blastocyst stage embryos exhibited an increased synthesis in eight proteins and decreased synthesis for one protein. The effect of IGF-I at the eight-cell stage of development was mostly inhibitory; the synthesis of only one protein increased and the synthesis of five proteins showed a decrease. Similar results were obtained with blastocyst stage embryos; four proteins demonstrated an increase in synthesis while 14 proteins showed a decrease. Eight-cell stage embryos incubated with IGF-II had seven proteins with a decreased synthesis, although in blastocyst stage embryos, nine proteins showed increased synthesis.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1994

Mechanisms of action of nonglucose insulin secretagogues.

Insulin release induced by nonglucose secretagogues is initiated from beta-cell by a wide variety of stimuli through specific receptors or binding sites. Activation of receptors in turn generates or enhances the cytosol levels of cAMP, cADPR, IP3, DAG, and AA. These second messengers then activate protein kinases, change the ion currents cross the cell membrane, and mobilize intracellular Ca2+, thereby increasing phosphorylated proteins in the cytosol and augmenting [Ca2+]i. These events trigger exocytotic discharge of insulin. The crucial steps in receptor-mediated stimulation-secretion coupling and their relationship to glucose-stimulated insulin release is summarized in Figure 1. At the present stage of research, the general processes of secretagogue binding to receptors, of generating second messengers, of activating several types of protein kinase, and of altering the membrane potential as well as cytosol calcium levels has been intensively studied and qualitatively clarified. However, we know little about the exact nature of substrates of different protein kinases and their function in the insulin secretion process. With the help of molecular biology and protein chemistry, we expect that this gap will be filled in the near future.

Adenylyl Cyclases↗
PubMed1994

In situ glucose uptake and glucokinase activity of pancreatic islets in diabetic and obese rodents.

The present study evaluated the involvement of glucose transport and phosphorylation in glucose-stimulated insulin release from pancreatic islets. Using quantitative histochemical techniques, we investigated basal islet glucose content, islet glucose uptake in situ during acute extreme experimental hyperglycemia, and islet glucokinase activity in several animal models of diabetes and obesity. The basal islet glucose content in anaesthetized diabetic or obese rodents was either the same or higher than that in their relevant controls. The rate of glucose uptake of islet tissue in these animals after an i.v. glucose injection was different. The db+/db+ mouse and the obese Zucker rat exhibited significantly reduced islet glucose uptake rates. RIP-cHras transgenic mice, BHE/cdb rats and partially pancreatectomized rats showed normal islet glucose uptake rates. The activity of islet glucokinase was increased to a different degree related to the blood glucose level. All five animal models of diabetes or obesity exhibited either a delay or a reduction of insulin release in response to supra maximal glucose stimulation. Our results indicate that the impairment of glucose-induced insulin release in diabetes is not consistently associated with a reduction of islet glucose uptake nor a change of glucokinase activity.

Animals↗
PubMed1994

Effects of glucose on insulin secretion, glucokinase activity, and transgene expression in transgenic mouse islets containing an upstream glucokinase promoter-human growth hormone fusion gene.

We have analyzed in organ culture the effects of glucose on glucose-induced insulin secretion, glucokinase (GK) activity, and human growth hormone (hGH) expression in pancreatic islets from transgenic mice containing an upstream GK promoter-hGH fusion gene. Freshly isolated islets from these mice had a normal insulin secretory response to glucose but showed subtle defects after culture in low or high glucose for 4 days that may have been due to the accumulation of hGH in the culture media. Islets cultured from both normal and transgenic mice had approximately a fourfold induction of GK activity in response to an increased concentration of glucose in the culture media, whereas no such change in total islet hGH production was observed. Immunocytochemical localization of hGH in islets cultured in 3 mM glucose showed a pattern similar to that in freshly isolated islets. However, after culture in 30 mM glucose, hGH immunostaining became strikingly more heterogeneous. We conclude 1) that GK-hGH transgene expression does not appear to adversely affect glucose-stimulated insulin secretion in vivo or in freshly isolated islets, 2) that glucose does not induce transgene expression, thus providing additional evidence against an effect of glucose on GK gene transcription in the islet, and 3) that glucose stimulates the co-release of hGH with insulin, thereby enhancing the heterogeneous staining pattern seen among pancreatic beta-cells.

Animals↗
PubMed1993

Altered malate dehydrogenase activity in nucleus magnocellularis of the chicken following cochlea removal.

The metabolism of second order auditory neurons in nucleus magnocellularis (NM) in the chick brainstem can be profoundly altered when excitatory input from the cochlea is removed. Within hours of cochlea removal, NM neurons show an increase in histochemical staining for the Kreb's cycle enzyme succinate dehydrogenase (SDH), followed in several days by decreases in SDH staining. We examined the activity of another Kreb's cycle enzyme, malate dehydrogenase (MDH) using a histochemical stain and a sensitive quantitative biochemical assay for comparison. We found changes in MDH staining similar in time course and magnitude to those of SDH; within 4 h of cochlea removal, MDH activity increases in ipsilateral NM neurons. By 9 days after cochlea removal MDH activity decreases, although not to the same degree as seen with SDH. Biochemical measurements of MDH activity also showed an early increase in activity in ipsilateral NM, followed at 9 days survival by a decrease in activity. Biochemical measurements of the activity of other enzymes in NM may be useful in further defining the metabolic consequences of deafferentation.

Animals↗
PubMed1993

Sulfonylurea-binding sites and ATP-sensitive K+ channels in alpha-TC glucagonoma and beta-TC insulinoma cells.

alpha-Cells secrete glucagon in a fuel-dependent fashion. We tested the hypothesis that alpha-cells contain sulfonylurea- and ATP-sensitive K+ channels. We studied two clonal lines of alpha-TC cells (simian virus 40 T-antigen induced glucagonoma cells) and for reference purposes, similarly transformed beta-TC insulinoma cells. alpha-TC cells each contained approximately 3000 high-affinity binding sites for the sulfonylurea [3H]glyburide. Whole-cell ATP- and tolbutamide-sensitive K+ currents of alpha-TC and beta-TC cells, relative to cell surface area, were comparable. In cell-attached membrane patches of alpha-TC cells, two types of K+ channels were observed. They had slope conductances of approximately 63 and 33 pS when the electrode contained 151 mM K+. Tolbutamide and diazoxide decreased and enhanced, respectively, the open probability of these channels. The membrane of alpha-TC cells depolarized periodically. This electrical activity was inhibited by diazoxide. A physiological mixture of amino acids enhanced glucagon release, and high glucose partially inhibited this release. Tolbutamide also enhanced glucagon release, whereas diazoxide inhibited it. Thus, alpha-TC glucagonoma cells contain ATP-sensitive K+ channels that regulate glucagon release, yet allow inhibition of hormone release by glucose.

Adenosine Triphosphate↗
PubMed1992

Expression and function of GLUT-1 and GLUT-2 glucose transporter isoforms in cells of cultured rat pancreatic islets.

We have previously investigated glucose induction of glucokinase, glucose usage and insulin release in isolated cultured rat pancreatic islets (Liang, Y., Najafit, H., Smith, R. M., Zimmerman, E. C., Magnuson, M. A., Tal, M., and Mastchinsky, F. M. (1992) Diabetes (1992) 41, 792-806). Here we studied the expression and function of GLUT-1 and GLUT-2 glucose transporter isoforms, using the same system, i.e. isolated pancreatic rat islets immediately after isolation or cultured in the presence of 3 or 30 mM glucose for as long as 10 days. We found by immunofluorescence microscopy and Western and Northern blot analysis of islet extracts that GLUT-1 expression was induced in islet beta-cells in tissue culture both with low or high glucose present. The induction of GLUT-1 was specific to beta-cells but was not present in all beta-cells and was not detected in alpha-cells. GLUT-2 expression was also specific for beta-cells and was not observed in all beta-cells. Some beta-cells in culture coexpressed GLUT-1 and GLUT-2. The expression of the two glucose transporters was regulated in the opposite direction in response to glucose concentration in the culture medium. GLUT-1 was more effectively induced when glucose was low, and GLUT-2 expression was more pronounced when glucose was high in the culture media. Another difference between the two glucose transporters was that GLUT-2 expression was increased while GLUT-1 expression was decreased as culturing continued as long as 7 days. Thus, after 7 days of culture GLUT-2 expression in beta-cells was nearly the same at low and high glucose, whereas GLUT-1 was practically absent no matter what the glucose level was. In attempts to correlate GLUT-1 and GLUT-2 expression to beta-cell function glucose uptake and glucose-stimulated insulin release in fresh and cultured islets were measured. In freshly isolated islet glucose uptake was estimated to be 100-fold in excess of actual glucose use. Glucose uptake was reduced by 7-day culture to about one-third of that observed in freshly isolated islets no matter what the glucose concentration of the culture media. We conclude that in the present experimental system GLUT-1 and GLUT-2 expression and function are not closely associated with glucose usage rates or the secretory function of beta-cells.

Animals↗
PubMed1992

Identification of glucose response proteins in two biological models of beta-cell adaptation to chronic high glucose exposure.

High resolution 2-dimensional sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) combined with computerized analysis of gel images was used to search for proteins whose biosynthesis was induced or repressed in pancreatic islet cells chronically exposed to high glucose in an in situ and a tissue culture model of islet cell adaptation to excessive fuel load. The in situ model involved a 4-day intravenous infusion of either 50% glucose or 0.45% saline solution, followed by islet isolation, [35S]methionine labeling at 3 and 18 mM glucose for both groups, and protein analysis by 2-dimensional SDS-PAGE. The tissue culture model involved a 7-day culture of isolated rat islets in RPMI 1640 with 10% fetal calf serum containing either 3 or 30 mM glucose, followed by radiolabeling and 2-dimensional PAGE of proteins as in the in situ model. A small fraction of about 1.5% of the approximately 2000 identifiable proteins can be characterized as adaptive proteins. Of these altogether 58 proteins in the two models, 5 proteins were demonstrable in both models and two of these (proteins 1526 and 7622) are particularly noteworthy. Protein 1526 (Mr 57,000; pI 5.09) showed the same response pattern in both models and its expression was most enhanced when islets from chronically glucose-infused animals or those cultured for 7 days at 30 mM were radiolabeled at 18 mM glucose. Protein 7622 (Mr 68,000; pI 6.50) (also known as GSP-65; Collins, H.W., Buettger, C., and Matschinsky, F.M. (1990) Proc. Natl. Acad. Sci. U.S.A. 87, 5494-5498) showed a different labeling pattern in the two models: stimulation of [35S]methionine incorporation by 18 mM glucose both in control and experimental islets from the infusion study, but lack of such stimulation of radiolabeling in islets cultured for 7 days at 30 mM glucose in contrast to islets cultured at 3 mM. The experimental strategy and the methodology are evaluated and the significance of the results is discussed. Potentials of the approach and plans for future experiments are considered.

Animals↗
PubMed1992

Metabolic labelling and quantitation of proteins synthesized by single chick cochleas.

Molecular studies of the peripheral auditory system are made difficult by the small quantities of tissue available and by their relative inaccessibility. In addition, the cochlea and other hair cell-containing receptor organs are composed of both hair cells and supporting cells, as well as several other cell types. The identification of known proteins and the characterization of specific and novel protein molecules from these tissues require the use of sensitive techniques and a consideration of the complex histology. The chick cochlea was selected as an experimental system since the cochlea is relatively accessible in the bird, the receptor neuroepithelium contains a large number of hair cells compacted in a small area, and the physiology of the auditory periphery has been studied extensively. A general procedure is described for the metabolic radiolabelling of proteins from single cochleas followed by their solubilization, separation by high-resolution two-dimensional gel electrophoresis, and accurate quantitation. The method is highly reproducible and sensitive, and should prove useful in studies of proteins from the specialized cell types of the chick cochlea, including the identification of those whose rates of synthesis are modified in response to acoustic stimulation and sound damage or recovery.

Animals↗
PubMed1992

Concordant glucose induction of glucokinase, glucose usage, and glucose-stimulated insulin release in pancreatic islets maintained in organ culture.

Using cultured islets as the experimental system, this study established dosage-response and time-dependency curves of the inductive glucose effect on glucose-stimulated insulin release, glucose usage, and glucokinase activity. Glucose-stimulated insulin release in islets cultured for 1, 2, or 7 days was increased as a function of glucose concentration in the culture medium and as a function of time. Glucose usage in the cultured islets showed a close relationship with glucose concentration in the culture medium at both 2 and 7 days of culture. Glucokinase activity increased in islets cultured for 1, 2, or 7 days as a function of increasing glucose concentrations in the culture medium and as a function of time. The V(max) of glucokinase in islets cultured for 7 days in medium containing 30 mM glucose was twice the value of freshly isolated islets and was almost fivefold higher than that in islets cultured for 7 days in 3 mM glucose. The glucose induction of glucose-stimulated insulin release, of glucose usage, and of glucokinase activity were tightly correlated. The biochemical mechanisms of glucose induction of islet glucokinase were further studied. Immunoblotting with an antibody against C-terminal peptide of glucokinase showed that densities of a 52,000-kD protein band from tissue extracts of islets cultured for 7 days in 3, 12, and 30 mM glucose were 25, 44, and 270% compared with that of extract from freshly isolated islets (100%). RNA blot analysis of glucokinase mRNA demonstrated virtually the same levels in fresh islets and islets after 7 days of culture in 3 or 30 mM glucose. The adaptive response of glucokinase to glucose appears therefore to be occurring at a translational or posttranslational site in cultured islets. These data greatly strengthen the concept that glucose is the regulator that induces the activity of glucokinase, which in turn determines the rate change of glucose usage as well as glucose-stimulated insulin release from beta-cells. Thus, the hypothesis that glucokinase is the glucose sensor of beta-cells is strengthened further.

Animals↗
PubMed1992

Human retinal pigment epithelial cells cultured in hyperglycemic media accumulate increased amounts of glycosaminoglycan precursors.

Confluent human retinal pigmented epithelial cells were cultured on microcarrier beads in the presence of 5.6 or 26 mmol/l glucose with or without the aldose-reductase inhibitor Sorbinil (200 microM) for 2 wk. At the end of the incubation period, perchloric acid extracts were prepared and analyzed by 31P nuclear magnetic resonance spectroscopy. As assessed by this method, the phosphorylated metabolites of cells incubated with 5.6 or 26 mmol/l glucose differed significantly in the concentrations of a number of uridine diphosphate (UDP)-conjugated monosaccharides, which were elevated two- to threefold in cells incubated in 26 mmol/l glucose over control samples. The affected metabolites were identified (through a series of spiking experiments) to be UDP-N-acetylglucosamine, UDP-N-acetylgalactosamine, and UDP-glucuronic acid. Coincubation of the cells with Sorbinil 200 microM in the presence of 26 mmol/l glucose had no effect on this accumulation. Under normal circumstances, these molecules selectively and sequentially are incorporated into the polysaccharide chains of glycosaminoglycans (GAGs), whose presence and distribution in the basement membranes is affected adversely by diabetes mellitus. These data suggest that the availability of the monosaccharide precursor is not the rate-limiting step for GAG synthesis in the presence of pathologic glucose concentrations. Thus, the lost GAG content in the basement membranes of diabetic patients may be caused by changes elsewhere in the biosynthesis and/or catabolism of the polysaccharide-linked protein molecules.

Aldehyde Reductase↗
PubMed1991

The role of ATP and free ADP in metabolic coupling during fuel-stimulated insulin release from islet beta-cells in the isolated perfused rat pancreas.

The isolated perfused rat pancreas was used to test the hypothesis that total cellular ATP or the ratio of ATP/free ADP plays the primary role in coupling intermediary metabolism to the biophysical events that are the basis of glucose-stimulated insulin release. The pancreas was preperfused for 20 min with 4.0 mM of a physiological mixture of 20 amino acids plus 4.2 mM glucose, and insulin release was then stimulated for 150 s by suddenly increasing the glucose to 8.3 mM. The pancreas was sampled at 24, 48, 72, and 150 s after the switch. The content of total ATP, ADP, AMP, Pi, phosphocreatine, and creatine were measured in beta-cell enriched cores of pancreatic islets microdissected from freeze-dried pancreas cryostat sections. Metabolites were measured by quantitative histochemical enzymatic cycling techniques. Modeling studies were carried out to assess the impact of biochemical analytical results on the membrane potential of the beta-cells. The level of free ADP was calculated using the creatine kinase equilibrium reaction and an intracellular pH of 7.2. First phase insulin release was stimulated at least 10-fold with the maximum reached 45 s after adding high glucose. The biochemical analytical data demonstrate that the total cellular level of the putative coupling factor ATP and of the ratios ATP/free ADP and ATP/free ADP x Pi are not significantly influenced by a glucose level change that causes a more than 10-fold surge of insulin release. The strength and limitations of the present experimental strategy and the implications of the results for our understanding of metabolic coupling in glucose-stimulated insulin release are discussed.

Adenosine Diphosphate↗
PubMed1991

Effects of alternate RNA splicing on glucokinase isoform activities in the pancreatic islet, liver, and pituitary.

Different glucokinase isoforms are produced by tissue-specific alternative RNA splicing in the liver and pancreatic islet, the only tissues in which glucokinase activity has been detected. To determine whether differences in protein structure brought about by alternative RNA splicing have an effect on glucose phosphorylating activity, we expressed cDNAs encoding four different hepatic and islet glucokinase isoforms and determined the Km and Vmax of each. When the glucokinase B1 and L1 isoforms were expressed in eukaryotic cells, both high Km glucose phosphorylating activity and immunoreactive protein were detected. However, when the glucokinase B2 and L2 isoforms were expressed, both of which differ by deletion of 17 amino acids in a region between the putative glucose and ATP-binding domains, no high Km glucose phosphorylating activity and much less immunoreactive protein were detected. When the glucokinase B1 and B2 isoforms were expressed in Escherichia coli as fusion proteins with glutathione S-transferase, affinity-purified B1 fusion protein was able to phosphorylate glucose whereas the B2 fusion protein was not, thus indicating that the lack of glucose phosphorylating activity from both the B2 and L2 isoforms is due to lack of intrinsic activity in addition to accumulation of less protein. The Km values of the B1 and L1 isoforms, which differ from each other by 15 amino acids at the NH2 terminus, were similar, but the Vmax of the B1 isoform was 2.8-fold higher than that of the L1 isoform. Mutagenesis of the first two potential initiation codons in the glucokinase B1 cDNA from ATG to GTC (methionine to valine) indicated that the first ATG was crucial for activity and is, therefore, the likely translation initiation codon. Messenger RNAs encoding both the B2 and L2 isoforms of glucokinase were detected in islet and liver by polymerase chain reaction amplification of total cDNA, indicating that mRNAs utilizing this weak alternate splice acceptor site in the fourth exon are normally present in both the liver and islet but as minor components. A regulatory role for weak alternate splice acceptor and donor sites in the glucokinase gene was suggested by examining the expression of the gene in the pituitary and in AtT-20 cells. Interestingly, although glucokinase mRNAs of appropriate sizes were detected in both the AtT-20 cells and rat pituitaries, neither exhibited any detectable high Km glucose phosphorylating activity.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed1991

Hormone secretion by isolated perfused pancreas of aging Fischer 344 rats.

This is the first study investigating hormone secretion by the isolated perfused pancreas of the aged Fischer 344 rat. Nutrient-induced release of insulin, glucagon, and somatostatin in overnight-fasted rats aged 2, 10, 18, 24, and 30 mo was studied. After an equilibration period, the pancreas was perfused for 20 min with buffer A (containing 4.2 mM glucose plus a 3.5 mM mixture of amino acids) then for 20 min with buffer B (containing 8.3 mM glucose and 7.0 mM amino acids). When stimulated by buffer B, the amount of insulin secreted increased (P less than 0.05) from immaturity (2 mo) to adulthood (10-18 mo) because of growth of the organ. From adulthood to very old age an equal amount of insulin was released in all groups during the last 19 min of perfusion with buffer B. Fasting blood glucose levels remained constant throughout life, whereas pancreatic insulin stores and plasma insulin levels rose, reaching peak values at 18 mo. The alpha-cell appeared to be deemphasized relative to the beta-cell during development but not thereafter, as indicated by the findings that from immaturity to adulthood pancreatic glucagon stores expanded less than insulin stores and that glucagon release significantly decreased. Only minor changes in somatostatin release from the delta-cells were observed after the rat reached adulthood. We conclude that the endocrine secretory response of the pancreas is well maintained throughout life in the Fischer 344 rat.

Aging↗
PubMed1991

Measurement of extracellular space in the rabbit AV node.

We used quantitative histochemistry to measure the size of the extracellular space (ESC) in various regions of the rabbit heart. When inulin, sucrose, and sorbitol were used as ECS markers, the ECS of the AV-nodal tissue was found to be, respectively, 2.4, 2.2, and 2.5 times larger than that of left ventricular muscle. Glucose was also measured over a 50-fold serum concentration range as an extracellular marker for AV-nodal tissue, left ventricular muscle, and Purkinje fibers. Measurements with glucose also revealed that the ECS of the AV node was 2.5-2.8 times larger than that of ventricular muscle. In contrast, the ECS of the AV node was the same as that of Purkinje fibers when glucose was used as an extracellular marker. ATP content, measured as an intracellular marker, was similar in both AV-nodal and contractile tissue. Collectively, the data obtained with all extracellular markers indicate that the ECS of the AV-nodal region is approximately 2.5 times larger than that of adjacent contractile tissue. Differences in the size of the ECS in various regions of the heart probably have functional significance and should be considered appropriately during the interpretation of data obtained by biochemical and densitometric approaches.

Acetylcholinesterase↗
PubMed1991

Content of CoA-esters in perifused rat islets stimulated by glucose and other fuels.

Fluorometry and high-performance liquid chromatography were used to measure the content of free CoA and the esters of acetate, malonate, succinate, and long-chain fatty acids in isolated perifused rat pancreatic islets exposed to 25 mM glucose or a mixture of fuels (25 mM glucose plus 10 mM glutamine, 10 mM lactate, and 1 mM pyruvate) to assess the role of intermediates of lipid metabolism as candidate metabolic coupling factors in the mechanism of fuel-induced insulin secretion. Insulin secretion was stimulated in a biphasic manner with the fuel mixture, showing twice the potency compared with high glucose alone. Islets perifused for 3 min with high glucose alone or the fuel mixture compared with 2.5 mM glucose showed a significant increase in malonyl-CoA and succinyl-CoA and a decrease in acetyl-CoA. Free CoA and long-chain acyl-CoA levels were unaltered. Perifused islets stimulated with 25 mM glucose for 30 min showed a significant increase in succinyl-CoA and long-chain acyl-CoA and decrease in acetyl-CoA, whereas malonyl-CoA was not affected. However, when islets were stimulated by the fuel mixture for 30 min, malonyl-CoA was maintained at a high level, and the change in succinyl-CoA and long-chain acyl-CoA was similar to that observed in islets stimulated with 25 mM glucose alone. The acetyl-CoA concentration in the islets stimulated with the fuel mixture decreased slightly. These results confirm the viability of the hypothesis that malonyl-CoA and long-chain acyl-CoA serve as metabolic coupling factors in signal transduction when islets are stimulated by high glucose or glucose combined with other fuels.

Acyl Coenzyme A↗