Anoctamin 1 (Ano1) is required for glucose-induced membrane potential oscillations and insulin secretion by murine ß-cells.

Anions such as Cl(-) and HCO3 (-) are well known to play an important role in glucose-stimulated insulin secretion (GSIS). In this study, we demonstrate that glucose-induced Cl(-) efflux from ß-cells is mediated by the Ca(2+)-activated Cl(-) channel anoctamin 1 (Ano1). Ano1 expression in rat ß-cells is demonstrated by reverse transcriptase-polymerase chain reaction, western blotting, and immunohistochemistry. Typical Ano1 currents are observed in whole-cell and inside-out patches in the presence of intracellular Ca(++): at 1 µM, the Cl(-) current is outwardly rectifying, and at 2 µM, it becomes almost linear. The relative permeabilities of monovalent anions are NO3 (-) (1.83 ± 0.10) > Br(-) (1.42 ± 0.07) > Cl(-) (1.0). A linear single-channel current-voltage relationship shows a conductance of 8.37 pS. These currents are nearly abolished by blocking Ano1 antibodies or by the inhibitors 2-(5-ethyl-4-hydroxy-6-methylpyrimidin-2-ylthio)-N-(4-(4-methoxyphenyl)thiazol-2-yl)acetamide (T-AO1) and tannic acid (TA). These inhibitors induce a strong decrease of 16.7-mM glucose-stimulated action potential rate (at least 87 % on dispersed cells) and a partial membrane repolarization with T-AO1. They abolish or strongly inhibit the GSIS increment at 8.3 mM and at 16.7 mM glucose. Blocking Ano1 antibodies also abolish the 16.7-mM GSIS increment. Combined treatment with bumetanide and acetazolamide in low Cl(-) and HCO3 (-) media provokes a 65 % reduction in action potential (AP) amplitude and a 15-mV AP peak repolarization. Although the mechanism triggering Ano1 opening remains to be established, the present data demonstrate that Ano1 is required to sustain glucose-stimulated membrane potential oscillations and insulin secretion.

Expression and localization of glucose transporters in rodent submandibular salivary glands.

BACKGROUND/AIM: The submandibular gland is one of the three major salivary glands, producing a mixed secretion; this saliva is hypotonic compared to plasma. It also secretes glucose, but the mechanisms responsible for this process are poorly understood. Our study addressed the question whether glucose transporters are expressed and how are they localized within specific rodent submandibular cells, in order to estimate a possible implication in salivary glucose disposal. METHODS: Immunohistochemistry, RT-qPCR and Western blotting were performed to determine the presence/localization of glucose transporters in rodent submandibular glands. RESULTS: GLUT4 was identified in the submandibular salivary gland at both mRNA and protein level. The immunohistochemical analysis revealed its localization preponderantly in the ductal cells of the gland, near to the basolateral. SGLT1 and GLUT1 were highly expressed in submandibular tissues in both acinar and ductal cells, but not GLUT2. These results were confirmed by RT-qPCR. It was also documented that insulin stimulates the net uptake of D-glucose by ductal rings prepared from submandibulary salivary glands, the relative magnitude of such an enhancing action being comparable to that found in hemidiaphragms. CONCLUSION: At least three major glucose transporters are expressed in the rodent submandibular glands, of which GLUT4 is specifically localized near the basolateral side of ductal structures. This points-out its possible role in regulating glucose uptake from the bloodstream, most likely to sustain ductal cellular metabolism.

Uptake and metabolism of D-glucose in isolated acinar and ductal cells from rat submandibular glands.

The present study deals with the possible effects of selected environmental agents upon the uptake and metabolism of d-glucose in isolated acinar and ductal cells from the rat submandibular salivary gland. In acinar cells, the uptake of d-[U-(14) C]glucose and its non-metabolised analogue 3-O-[(14) C-methyl]-d-glucose was not affected significantly by phloridzin (0.1 mM) or substitution of extracellular NaCl (115 mM) by an equimolar amount of CsCl, whilst cytochalasin B (20 µM) decreased significantly such an uptake. In ductal cells, both phloridzin and cytochalasin B decreased the uptake of d-glucose and 3-O-methyl-d-glucose. Although the intracellular space was comparable in acinar and ductal cells, the catabolism of d-glucose (2.8 or 8.3 mM) was two to four times higher in ductal cells than in acinar cells. Phloridzin (0.1 mM), ouabain (1.0 mM) and cytochalasin B (20 µM) all impaired d-glucose catabolism in ductal cells. Such was also the case in ductal cells incubated in the absence of extracellular Ca(2+) or in media in which NaCl was substituted by CsCl. It is proposed that the ductal cells in the rat submandibular gland are equipped with several systems mediating the insulin-sensitive, cytochalasin B-sensitive and phloridzin-sensitive transport of d-glucose across the plasma membrane.

Comparison of GLUT1, GLUT2, GLUT4 and SGLT1 mRNA expression in the salivary glands and six other organs of control, streptozotocin-induced and Goto-Kakizaki diabetic rats.

BACKGROUND/AIMS: The expression and localization of several distinct glucose transporters (GLUT1, GLUT2, GLUT4, and SGLT1) was recently characterized in the parotid gland of normal rats by quantitative real-time PCR analysis, immunohistochemistry and Western blotting. The major aims of the present study was to compare the mRNA expression of these glucose transporters in both the parotid gland and submaxillary gland of control rats, streptozotocin-induced diabetic rats and hereditarily diabetic Goto-Kakizaki rats. METHODS: Quantitative real-time PCR analysis was performed in the parotid and submaxillary salivary glands and, for purpose of comparison, also in the heart, kidney, liver, lung, muscle and pancreas from control animals and either streptozotocin-treated or Goto-Kakizaki rats. RESULTS: The expression of GLUT4, but not GLUT1 or SGLT1, mRNA was decreased in the diabetic rats. The results also allow comparing both the mRNA expression level of the four glucose transporters in salivary glands and six other organs, and the diabetes-induced changes in such an expression in distinct locations. CONCLUSION: The mRNA expression of the insulin-dependent GLUT4 transporter was the sole to be significantly decreased in the salivary glands of diabetic animals. The possible consequence of such a decrease in terms of the control of salivary glucose concentration requires further investigation.

Glucose transport by acinar cells in rat parotid glands.

BACKGROUND/AIMS: Salivary glucose is often considered as being from glandular origin. Little information is available, however, on the possible role of glucose transporters in the secretion of the hexose by salivary glands. The major aim of the present study was to investigate the expression and localization of several distinct glucose transporters in acinar cells of rat parotid glands. METHODS: Quantitative real-time PCR analysis, immunohistochemistry and western blotting techniques were used to assess the presence of SGLT1, GLUT1, GLUT2 and GLUT4 in acinar cells of rat parotid glands. RESULTS: Quantitative real-time PCR documented the expression of SGLT1 and GLUT1 in parotid tissues, with a much lower level of GLUT4 mRNA and no expression of GLUT2 mRNA. Western blot analysis revealed the presence of SGLT1, GLUT1 and GLUT4 proteins, but not GLUT2 proteins in the parotid extract. Immunohistochemistry confirmed these findings. SGLT1 was specifically located at the baso-lateral membrane, co-localizing with Na(+)/K(+) ATPase. GLUT1 was found both at the baso-lateral and apical level. GLUT4 appeared to be also located at the baso-lateral level. However, too little GLUT4 was present to allow co-localization labeling. CONCLUSION: Based on these findings, a model is proposed for the transport of glucose into the acinar cells and thereafter into the acinar lumen.

Expression of TMEM16A and SLC4A4 in human pancreatic islets.

BACKGROUND/AIMS: Stimulation of insulin release by D-glucose is accompanied by Cl(-) and HCO(3)(-) efflux from pancreatic islet cells. The efflux of these anions may involve volume-regulated anion channels, including possibly TMEM16A, and the Na(+)-HCO(3)(-)-cotransporter SLC4A4. The present study was designed to explore the expression of both TMEM16A and SLC4A4 in human pancreatic islets. METHODS: Pancreases were obtained from human cadaveric donors. Immunodetection of TMEM16A and SLC4A4 was performed by immunohistochemistry on sections of fixed pancreas, while real-time PCR for the study of corresponding gene expression was performed on RNA extracted from both total pancreatic pieces and isolated pancreatic islets. RESULTS: RT-PCR yielded lower levels of SLC4A4 in isolated islets than in the total pancreas, whilst a mirror image prevailed for TMEM16A mRNA. Immunohistochemistry of human pancreas, however, indicated comparable immunostaining of SLC4A4 in insulin-producing cells and exocrine pancreatic cells, whilst that of TMEM16A appeared less pronounced in insulin-producing cells than in exocrine cells. CONCLUSION: The present findings support the view that, in humans like in rodent, the regulation of anion fluxes in insulin-producing cells may involve both SLC4A4 and TMEM16A.

A new role for aquaporin 7 in insulin secretion.

UNLABELLED: BACGROUNS/AIMS: Several insulinotropic agents were recently reported to cause ß-cell swelling. The possible participation of AQP7 to water transport was investigated in AQP7(+/+) or AQP7(-/-) mice. METHODS: Aquaporin expression, insulin secretion, cell swelling and electrical activity were investigated in pancreatic islets. RESULTS: RT-PCR revealed the expression of AQP5 and AQP8 mRNA. Double immunofluorescent labeling indicated their presence in ß-cells. Whilst basal insulin release from isolated pancreatic islets incubated at 2.8 mM D-glucose did not differ between AQP7(+/+) or AQP7(-/-) mice, the secretion of insulin evoked by the omission of 50 mM NaCl, the substitution of 50 mM NaCl by 100 mM glycerol or a rise in D-glucose concentration to 8.3 mM and 16.7 mM was severely impaired in the islets from AQP7(-/-) mice. Yet, exposure of ß-cells to either the hypotonic medium or a rise in D-glucose concentration caused a similar degree of swelling and comparable pattern of electrical activity in cells from AQP7(+/+) and AQP7(-/-) mice. Both the cell swelling and change in membrane potential were only impaired in AQP7(-/-) cells when exposed to 50 mM glycerol. CONCLUSION: It is proposed, therefore, that AQP7 may, directly or indirectly, play a role at a distal site in the exocytotic pathway.

(19)F-heptuloses as tools for the non-invasive imaging of GLUT2-expressing cells.

Suitable analogs of d-mannoheptulose are currently considered as possible tools for the non-invasive imaging of pancreatic islet insulin-producing cells. Here, we examined whether (19)F-heptuloses could be used for non-invasive imaging of GLUT2-expressing cells. After 20 min incubation, the uptake of (19)F-heptuloses (25 mM) by rat hepatocytes, as assessed by (19)F NMR spectroscopy, ranged from 0.50 (1-deoxy-1-fluoro-d-mannoheptulose and 3-deoxy-3-fluoro-d-mannoheptulose) to 0.25 (1,3-dideoxy-1,3-difluoro-d-mannoheptulose) and 0.13 (1-deoxy-1-fluoro-d-glucoheptulose, 3-deoxy-3-fluoro-d-glucoheptulose and 1,3-dideoxy-1,3-difluoro-d-glucoheptulose) µmol per 3×10(6)cells. (19)F MRI experiments also allowed the detection of 1-deoxy-1-fluoro-d-mannoheptulose in rat hepatocytes. All three (19)F-mannoheptuloses cited above, as well as 7-deoxy-7-fluoro-d-mannoheptulose and 1-deoxy-1-fluoro-d-glucoheptulose inhibited insulin release evoked in rat isolated pancreatic islets by 10mM d-glucose to the same extent as that observed with an equivalent concentration (10mM) of d-mannoheptulose, while 3-deoxy-3-fluoro-d-glucoheptulose and 1,3-dideoxy-1,3-difluoro-d-glucoheptulose (also 10mM) were less potent than d-mannoheptulose in inhibiting insulin release. The 1-deoxy-1-fluoro-d-mannoheptulose and 3-deoxy-3-fluoro-d-mannoheptulose only marginally affected INS-1 cell viability. These findings are compatible with the view that selected (19)F-heptuloses may represent suitable tools for the non-invasive imaging of hepatocytes and insulin-producing cells by (19)F MRI.

Expression of the electrogenic Na+-HCO3--cotransporter NBCe1 in tumoral insulin-producing BRIN-BD11 cells.

BACKGROUND/AIMS: The expression of the electrogenic Na+-HCO3--cotransporter NBCe1 was recently documented in rat pancreatic islet B-cells, it being speculated that such a protein participates in the extrusion of bicarbonate generated by the oxidative catabolism of nutrients from insulin-producing cells. Considering the prevalence of a Crabtree effect in tumoral insulin-producing cells, the possible presence of NBCe1 was now investigated in BRIN-BD11 cells, an insulin-producing cell line established by electrofusion of normal pancreatic B-cells with immortalized RINm5F cells. METHODS: The possible presence of NBCe1 in BRIN-BD11 cells was investigated by RT-PCR, western blot analysis and immunocytochemistry. The release of insulin and net uptake of 22Na+ were also measured in the BRIN-BD11 cells. RESULTS: RT-PCR, western blot analysis and immunocytochemistry documented the presence of NBCe1 in BRIN-BD11 cells. A reported inhibitor of NBCe1, i.e. tenidap, (50-100 microM), inhibited basal and hypotonicity-induced insulin release from the BRIN-BD11 cells, whilst increasing the net uptake of 22Na+ by the same cells. The latter effect was, in relative terms, more pronounced in the presence than absence of ouabain. CONCLUSION: BRIN-BD11 cells, like normal pancreatic islet B-cells, express NBCe1, with predominance of the B variant of this electrogenic Na+-HCO3--cotransporter.

Contrasting effects of glycerol and urea transport on rat pancreatic beta-cell function.

BACKGROUND/AIMS: Pancreatic beta-cell function is influenced by changes in cell volume. Such volume changes depend on water permeability of the plasma membrane, conferred in part by aquaporins. Islet cells express aquaporin 7 (AQP7), which is permeable to urea and glycerol in addition to water. We therefore investigated the effects of glycerol and urea on rat pancreatic beta-cell function. METHODS: Electrical activity and whole-cell current were studied using the perforated patch technique. Cell volume was measured by video-imaging and insulin release by radioimmunoassay. Aquaporin 7 expression was studied by RT-PCR, Western blot and double fluorescent immunolabelling. RESULTS: The isosmotic addition of glycerol and urea resulted in depolarization of the plasma membrane and electrical activity, accompanied by beta-cell swelling, activation of the volume-regulated anion channel (VRAC) and insulin release. However, the effects of glycerol, in contrast to urea, persisted throughout exposure to the osmolyte. Glycerol also caused beta-cell activation when added hyperosmotically. A non-metabolizable glycerol analogue had comparable effects to urea on beta-cells. The expression of AQP7 was demonstrated in rat beta-cells. CONCLUSION: Glycerol and urea can activate beta-cells via their rapid uptake across the beta-cell plasma membrane, possibly via AQP7. This results in cell swelling, VRAC activation, electrical activity and insulin release. Glycerol appears to exert an additional effect, possibly related to its intracellular metabolism.

Salivary glucose concentration and excretion in normal and diabetic subjects.

The present report aims mainly at a reevaluation of salivary glucose concentration and excretion in unstimulated and mechanically stimulated saliva in both normal and diabetic subjects. In normal subjects, a decrease in saliva glucose concentration, an increase in salivary flow, but an unchanged glucose excretion rate were recorded when comparing stimulated saliva to unstimulated saliva. In diabetic patients, an increase in salivary flow with unchanged salivary glucose concentration and glucose excretion rate were observed under the same experimental conditions. Salivary glucose concentration and excretion were much higher in diabetic patients than in control subjects, whether in unstimulated or stimulated saliva. No significant correlation between glycemia and either glucose concentration or glucose excretion rate was found in the diabetic patients, whether in unstimulated or stimulated saliva. In the latter patients, as compared to control subjects, the relative magnitude of the increase in saliva glucose concentration was comparable, however, to that of blood glucose concentration. The relationship between these two variables was also documented in normal subjects and diabetic patients undergoing an oral glucose tolerance test.

The metabolic syndrome of omega3-depleted rats. II. Body weight, adipose tissue mass and glycemic homeostasis.

Exposure of 7-week-old normal rats for 3-7 months to a diet deprived of long-chain polyunsaturated omega3 fatty acids was recently reported to induce changes in the fatty acid content and pattern of liver phospholipids and triglycerides similar to those otherwise found in second generation omega3-depleted rats. In the present study, the changes in body weight, parametrial adipose tissue mass, plasma glucose and insulin concentrations and insulin resistance index were investigated in the same control and omega3-depleted rats, which were then given access for 2 to 4-5 weeks to either a flaxseed oil-enriched diet (control and omega3-depleted rats) or a soybean oil-enriched diet (control rats). The body weight failed to differ between control and omega3-depleted rats. The latter rats, however, displayed increases in adipose tissue mass, plasma glucose and insulin concentrations, and insulin resistance index. In the control rats given access to the soybean or flaxseed oil-enriched diet, body weight and adipose tissue mass were little affected, but both the plasma glucose concentration and insulin resistance index decreased. In the omega3-depleted rats given access to the flaxseed oil-enriched diet, both body weight and adipose tissue mass underwent a rapid, pronounced and sustained increase, whilst the plasma glucose concentration and insulin resistance index decreased similarly to those in the control rats. The present design of omega3 fatty acid dietary deprivation thus reproduces the visceral obesity and insulin resistance otherwise observed in second-generation omega3-depleted rats. However, the supply of exogenous omega3 fatty acids to the omega3-depleted rats failed to oppose visceral obesity, possibly as a result of the orexigenic effects of these omega3 fatty acids.

The metabolic syndrome of omega3-depleted rats. I. Liver data.

Second-generation rats depleted in long-chain polyunsaturated omega3 fatty acids were recently proposed as a novel animal model for the metabolic syndrome. In the present study, a dietary deprivation of omega3 acids for 3-7 months was found sufficient to provoke in 6-week-old normal rats the same alteration of the fatty acid content and profile of liver phospholipids and triglycerides as that otherwise prevailing in the second-generation omega3-depleted rats, with emphasis on a severe decrease in their omega3 fatty acid content, alterations in the relative contribution of and ratio between selected long-chain polyunsaturated omega6 fatty acids, saturated and monodesaturated fatty acids and precursors of nervonic acid, and liver steatosis. When the omega3-depleted rats were exposed, after the first 7 months of the present experiments and for 2-4 weeks to a diet supplemented with 5% (w/w) flaxseed oil, most of these hepatic variables returned towards or beyond control values. In both the omega3-depleted rats and control animals, however, the eventual exposure to the flaxseed oil-enriched diet failed to suppress liver steatosis and, on the contrary, provoked a further increase in liver triglyceride content. It is proposed, therefore, that the present approach represents a simple and realistic animal model to study the consequences of omega3-depletion. Moreover, the results suggest that to oppose such consequences, e.g. liver steatosis, it may be necessary to combine the dietary supply of omega3 acids with a suitable control of food intake, in both qualitative and quantitative terms.

Rapid lipid enrichment in omega3 fatty acids: plasma data.

The bolus intravenous injection of a novel medium-chain triglyceride:fish oil emulsion to normal subjects was recently reported to enrich within 60 min the phospholipid content of leucocytes and platelets in long-chain polyunsaturated omega3 fatty acids. The present study, conducted in second generation omega3-depleted rats, aimed at investigating whether such a procedure may also increase within 60 min the phospholipid content of omega3 fatty acids in cells located outwards the bloodstream, in this case liver cells, and whether this coincides with correction of the perturbation in the liver triglyceride fatty acid content and profile otherwise prevailing in these rats. This first report deals mainly with the fatty acid pattern of plasma lipids in male omega3-depleted rats that were non-injected or injected with either the omega3-rich emulsion or a control medium-chain triglyceride:olive oil emulsion. The results provide information on the fate of the exogenous lipids present in the lipid emulsions and injected intravenously 60 min before sacrifice. Moreover, in the uninjected omega3-depleted rats the comparison between individual plasma and liver measurements indicated positive correlations in the fatty acid profile of phospholipids and triglycerides.

Rapid lipid enrichment in omega3 fatty acids: cause-to-effect relationships.

The bolus intravenous administration of a novel medium-chain triglyceride:fish oil emulsion to second generation rats depleted in long-chain polyunsaturated omega3 fatty acids was recently found to enrich within 60 min the content of both plasma and liver lipids in such omega3 fatty acids, this coinciding with correction of the perturbation in liver triglyceride fatty acid content and profile otherwise prevailing in these rats. The present report draws attention to cause-to-effect relationships between changes in liver phospholipid and triglyceride fatty acid content and/or pattern operative under these experimental conditions.

Rapid lipid enrichment in omega3 fatty acids: liver data.

The bolus intravenous injection of a novel medium-chain triglyceride:fish oil emulsion to normal subjects was recently reported to enrich within 60 min the phospholipid content of leucocytes and platelets in long-chain polyunsaturated omega3 fatty acids. The present study, conducted in second generation omega3-depleted rats, aims at investigating whether such a procedure may also increase within 60 min the phospholipid content of omega3 fatty acids in cells located outwards of the bloodstream, in this case liver cells, and whether this coincides with correction of the perturbation in the liver triglyceride fatty acid content and profile otherwise prevailing in these rats. The results indicate that such is indeed the case and further suggest a cause-to-effect relationship between the two events.

Correlation between liver and plasma fatty acid profile of phospholipids and triglycerides in rats.

Considering the changes in the fatty acid profile of liver lipids related to age, gender and nutritional status or occurring in pathological situations, this study aimed at investigating whether such changes could be judged from measurements conducted in plasma lipids. The fatty acid profile of both liver and plasma phospholipids and triglycerides was measured in 16 control animals and 26 rats depleted in long-chain polyunsaturated (n-3) fatty acids. Within each group of rats, significant correlations prevailed between the percentage of each fatty acid in liver versus plasma phospholipids or triglycerides. However, the plasma/liver ratio for the relative content of C20:5(n-3), C22:5(n-3) and C22:6(n-3) in triglycerides displayed abnormally high values in 2 control animals. The fatty acid profile of liver phospholipids and triglycerides can, as a rule, be judged from measurements made in the corresponding plasma lipids. For instance, measurements in plasma phospholipids could help to identify subjects deficient in (n-3) fatty acids and to assess the dietary correction of this defect.

Perturbation of 11-eicosenoate metabolism in female diabetic rats.

Considering the proposed preventive effect of nervonic acid on obesity- and diabetes-related coronary risk factors, the content of its precursors (oleic, 11-eicosenoic and 13-docosenoic acids) was measured in liver and plasma phospholipids and triglycerides, brain and spleen phospholipids, and adipose tissue lipids of fed or overnight fasted control and hereditarily diabetic Goto-Kakizaki female rats, as well as fed streptozotocin-induced diabetic female rats. In liver and brain phospholipids, the 11-eicosenoate/oleate ratio was significantly higher in diabetic rats than in control animals. Such was not the case in either spleen phospholipids or liver triglycerides and adipose tissue lipids. The increase in the liver phospholipid 11-eicosenoate/oleate ratio found in female diabetic rats represents a mirror image of the situation recently documented, in the same animal models of diabetes, in male rats. These contrasting findings may be relevant to the higher coronary heart disease risk prevailing in female, as compared to male, diabetic subjects.

Effects of plasma membrane Ca(2+)-ATPase overexpression upon D-glucose metabolism in insulin-producing BRIN-BD11 cells.

In order to investigate the possible link between PMCA (plasma-membrane Ca(2+)-ATPase) activity and D-glucose catabolism in insulin-producing cells, BRIN-BD11 cells were transfected with two isoforms of PMCA2. Transfection of insulin-producing BRIN-BD11 cells with PMCA2yb and PMCA2wb was documented by RT-PCR (reverse transcription-PCR), Western blot analysis, indirect immunofluorescence microscopy and (45)Ca(2+) uptake by microsomes. In the transfected cells, the overexpression of PMCA coincided with three major anomalies of D-glucose metabolism, namely a lower rate of D-[5-(3)H]glucose utilization prevailing at a low extracellular concentration of D-glucose (1.1 mM), a low ratio between D-[U-(14)C]oxidation and D-[5-(3)H]glucose utilization prevailing at a high extracellular glucose concentration (16.7 mM), and a high ratio between the net generation of (14)C-labelled acidic metabolites and amino acids and that of (3)H(2)O from D-[5-(3)H]glucose. These anomalies resulted in a decreased estimated rate of ATP generation (linked to the catabolism of the hexose) and a lowered ATP cell content, whether at low or high extracellular D-glucose concentrations. The net uptake of (45)Ca(2+) by intact cells was also decreased in the transfected cells, but to a greater extent than can apparently be attributed to the change in the ATP-generation rate. These findings document the relevance of PMCA activity to both D-glucose metabolism and Ca(2+) handling in insulin-producing cells, with emphasis on the key role of both cytosolic and mitochondrial Ca(2+) concentrations in the regulation of D-glucose catabolism. They also reveal that overexpression of PMCA leads, in insulin-producing cells, to an imbalance between ATP generation and consumption.

L-glutamine and palmitate catabolism in pancreatic islets from rats depleted in long-chain polyunsaturated omega 3 fatty acids.

The catabolism of D-glucose was recently found to be impaired in pancreatic islets from rats depleted in long-chain polyunsaturated omega3 fatty acids. The specificity of this alteration was now investigated by characterizing the oxidative fate of endogenous nutrients in islets preincubated with either L-[U-14C]glutamine or [U-14C]palmitate and then incubated variously in the absence of D-glucose, presence of the hexose or presence of metabolic poisons. Relative to their radioactive content after preincubation, the production of 14CO2 by islets prelabelled with [U-14C]glutamine was higher in omega3-depleted rats than control animals. The enhancing action of D-glucose upon such production was impaired, however, in the omega3-depleted rats. The net uptake of 14C-palmitate and absolute value for 14CO2 output were both increased in omega3-depleted rats, whilst the ratio between 14CO2 output and islet radioactive content was decreased in the same animals. The inhibition of 14CO2 production by metabolic poisons was comparable in all cases. These results are consistent with recent findings on such items as the availability of endogenous amino acids and uptake of unesterified fatty acids in extrapancreatic sites of the omega3-depleted rats. They also support the view that the alteration of D-glucose metabolism in the islets of the latter animals may be attributable, in part at least, to alteration of glucokinase kinetics by high intracellular acyl-CoA levels.