Expression of the kynurenine pathway enzymes in the pancreatic islet cells. Activation by cytokines and glucolipotoxicity.

The tryptophan/kynurenine pathway (TKP) is the main route of tryptophan degradation and generates several neuroactive and immunomodulatory metabolites. Experimental and clinical data have clearly established that besides fat, muscle and liver, pancreatic islet tissue itself is a site of inflammation during obesity and type 2 diabetes. Therefore it is conceivable that pancreatic islet exposure to increased levels of cytokines may induce upregulation of islet kynurenine metabolism in a way resembling that seen in the brain in many neurodegenerative disorders. Using normal rat islets and the INS-1 ß-cell line, we have demonstrated for the first time that: 1/only some TKP genes are constitutively expressed, both in ß-cells as well as non ß-cells; 2/ the regulatory enzyme indoleamine 2,3-dioxygenase (IDO1) is not constitutively expressed; 3/ IDO1 and kynurenine 3-monoxygenase (KMO) expression are potently activated by proinflammatory cytokines (IFN-γ, IL-1ß) and glucolipotoxicity respectively, rather in ß-cells than in non ß-cells; 4/ Islet kynurenine/kynurenic acid production ratio is enhanced following IFN-γ and glucolipotoxicity; 5/ acute exposure to KYN potentiates glucose-induced insulin secretion by normal islets; and 6/ oxidative stress or glucocorticoid modulates TKP genes only marginally. Pancreatic islets may represent a new target tissue for inflammation and glucolipotoxicity to activate the TKP. Since inflammation is now recognized as a crucial mechanism in the development of the metabolic syndrome and more specifically at the islet level, it is needed to evaluate the potential induction of the TKP in the endocrine pancreas during obesity and/or diabetes and its relationship to the islet cell functional alterations.

A euglycaemic/non-diabetic perinatal environment does not alleviate early beta cell maldevelopment and type 2 diabetes risk in the GK/Par rat model.

AIMS/HYPOTHESIS: We used the GK/Par rat, a spontaneous model of type 2 diabetes with early defective beta cell neogenesis, to determine whether the development of GK/Par offspring in a non-diabetic intrauterine/postnatal environment would prevent the alteration of fetal beta cell mass (BCM) and ultimately decrease the risk of diabetes later in adult life. METHODS: We used an embryo-transfer approach, with fertilised GK/Par ovocytes (oGK) being transferred into pregnant Wistar (W) or GK/Par females (pW and pGK). Offspring were phenotyped at fetal age E18.5 and at 10 weeks of age, for BCM, expression of genes of pancreatic progenitor cell regulators (Igf2, Igf1r, Sox9, Pdx1 and Ngn3), glucose tolerance and insulin secretion. RESULTS: (1) Alterations in neogenesis markers/regulators and BCM were as severe in the oGK/pW E18.5 fetuses as in the oGK/pGK group. (2) Adult offspring from oGK transfers into GK (oGK/pGK/sGK) had the expected diabetic phenotype compared with unmanipulated GK rats. (3) Adult offspring from oGK reared in pW mothers and milked by GK foster mothers had reduced BCM, basal hyperglycaemia, glucose intolerance and low insulin, to an extent similar to that of oGK/pGK/sGK offspring. (4) In adult offspring from oGK transferred into pW mothers and milked by their W mothers (oGK/pW/sW), the phenotype was similar to that in oGK/pGK/sGK or oGK/pW/sGK offspring. CONCLUSIONS/INTERPRETATION: These data support the conclusion that early BCM alteration and subsequent diabetes risk in the GK/Par model are not removed despite normalisation of the prenatal and postnatal environments, either isolated or combined.

Chromosomal mapping of genetic loci associated with non-insulin dependent diabetes in the GK rat.

Goto-Kakizaki (GK) rats are a well characterized model for non-insulin dependent diabetes mellitus (NIDDM). We have used a combination of physiological and genetic studies to identify quantitative trait loci (QTLs) responsible for the control of glucose homeostasis and insulin secretion in a F2 cohort bred from spontaneously diabetic GK rats. The genetic dissection of NIDDM allowed us to map up to six independently segregating loci predisposing to hyperglycaemia, glucose intolerance or altered insulin secretion, and a seventh locus implicated in body weight. QTLs implicated in glucose tolerance and adiposity map to the same region of rat chromosome 1, and may indicate the influence of a single locus. Our study demonstrates that distinct combinations of genetic loci are responsible for different physiological characteristics associated with the diabetic phenotype in the GK rat, and it constitutes an important step for directing the search for the genetic factors involved in human NIDDM.

IL-1 antagonism reduces hyperglycemia and tissue inflammation in the type 2 diabetic GK rat.

Recent studies suggest an inflammatory process, characterized by local cytokine/chemokine production and immune cell infiltration, regulates islet dysfunction and insulin resistance in type 2 diabetes. However, the factor initiating this inflammatory response is not known. Here, we characterized tissue inflammation in the type 2 diabetic GK rat with a focus on the pancreatic islet and investigated a role for IL-1. GK rat islets, previously characterized by increased macrophage infiltration, displayed increased expression of several inflammatory markers including IL-1beta. In the periphery, increased expression of IL-1beta was observed primarily in the liver. Specific blockade of IL-1 activity by the IL-1 receptor antagonist (IL-1Ra) reduced the release of inflammatory cytokines/chemokines from GK islets in vitro and from mouse islets exposed to metabolic stress. Islets from mice deficient in IL-1beta or MyD88 challenged with glucose and palmitate in vitro also produced significantly less IL-6 and chemokines. In vivo, treatment of GK rats with IL-1Ra decreased hyperglycemia, reduced the proinsulin/insulin ratio, and improved insulin sensitivity. In addition, islet-derived proinflammatory cytokines/chemokines (IL-1beta, IL-6, TNFalpha, KC, MCP-1, and MIP-1alpha) and islet CD68(+), MHC II(+), and CD53(+) immune cell infiltration were reduced by IL-1Ra treatment. Treated GK rats also exhibited fewer markers of inflammation in the liver. We conclude that elevated islet IL-1beta activity in the GK rat promotes cytokine and chemokine expression, leading to the recruitment of innate immune cells. Rather than being directly cytotoxic, IL-1beta may drive tissue inflammation that impacts on both beta cell functional mass and insulin sensitivity in type 2 diabetes.

The GK rat beta-cell: a prototype for the diseased human beta-cell in type 2 diabetes?

Increasing evidence indicates that decreased functional beta-cell mass is the hallmark of type 2 diabetes (T2D) mellitus. Nowadays, the debate focuses on the possible mechanisms responsible for abnormal islet microenvironment, decreased beta-cell number, impaired beta-cell function, and their multifactorial aetiologies. This review is aimed to illustrate to what extend the Goto-Kakizaki rat, one of the best characterized animal models of spontaneous T2D, has proved be a valuable tool offering sufficient commonalities to study these aspects. We propose that the defective beta-cell mass and function in the GK model reflect the complex interactions of multiple pathogenic players: (i) several independent loci containing genes responsible for some diabetic traits (but not decreased beta-cell mass); (ii) gestational metabolic impairment inducing an epigenetic programming of the pancreas (decreased beta-cell neogenesis and/or proliferation) which is transmitted to the next generation; and (iii) loss of beta-cell differentiation due to chronic exposure to hyperglycemia/hyperlipidemia, inflammatory mediators, oxidative stress and to perturbed islet microarchitecture.

Impaired pancreatic beta cell function in the fetal GK rat. Impact of diabetic inheritance.

The Goto-Kakisaki (GK) rat is a genetic model of non-insulin-dependent diabetes. At 21.5 d of age we found that GK fetuses had an increased plasma glucose concentration, a decreased plasma insulin level, and a reduced pancreatic beta cell mass. To investigate the beta cell function during fetal life we used a hyperglycemic clamp protocol applied to the mothers, which allowed us to obtain a steady-state hyperglycemia in the corresponding fetuses. At variance, with Wistar (W) fetuses, plasma insulin concentration in GK fetuses did not rise in response to hyperglycemia. In contrast, GK fetal pancreas released insulin in response to glucose in vitro to the same extent as W fetal pancreas. Such a discrepancy between the in vivo and in vitro results suggests that the lack of pancreatic reactivity to glucose as seen in vivo is extrinsic to the fetal GK beta cell. Finally, the importance of gestational hyperglycemia was investigated by performing crosses between GK and W rats. Fetuses issued from crosses between W mother and GK father or GK mother and W father had a beta cell mass close to normal values and were still able to increase their plasma insulin levels in response to hyperglycemia in vivo. Our data suggest that hyperglycemia in utero does not influence the severity of the decrease of the beta cell mass or the lack of the insulin secretory response to glucose in the fetal GK rat. Moreover they indicate that conjunction of GK genes originating from both parents is necessary in order for these defects to be fully expressed.

GLP-1 based therapy for type 2 diabetes.

Type 2 diabetes mellitus is a major and growing health problem throughout the world. Current treatment approaches include diet, exercise, and a variety of pharmacological agents including insulin, biguanides, sulfonylureas and thiazolidinediones. New therapies are still needed to control metabolic abnormalities, and also to preserve beta-cell mass and to prevent loss of beta-cell function. Glucagon-like peptide 1 (GLP-1) is a drug candidate which potentially fulfils these conditions. GLP-1 is an incretin hormone secreted by intestinal L-cells in response to meal ingestion is a novel pharmacological target with multiple antihyperglycemic actions. GLP-1 glucoregulatory actions include glucose-dependent enhancement of insulin secretion, inhibition of glucagon secretion, slowing of gastric emptying and reduction of food intake. GLP-1 is rapidly inactivated by amino peptidase, dipeptidyl peptidase IV (DPP-IV) and the utility of DPP-IV inhibitors are also under investigation. There is a recent upsurge in the development of GLP-1 mimetics and DPP-IV inhibitors as potential therapy for type 2 diabetes. However, both the strategies are having their own advantages and limitations. The present review summarizes the concepts of GLP-1 based therapy for type 2 diabetes and the current preclinical and clinical development in GLP-1 mimetics and DPP-IV inhibitors. Further, the potential advantages and the limitations of both the strategies are discussed.

Insulin treatment improves the spontaneous remission of neonatal streptozotocin diabetes in the rat.

Neonatal rats injected with streptozotocin (STZ, 100 mg/kg) at birth exhibit an acute diabetes that is characterized by a spontaneous and incomplete remission. The short- and long-term effect of exogenous insulin on the course of this neonatal diabetes has been studied. Insulin treatment (20 mU/g body wt./day, for 4 days) diminished the percentage of glycosuric animals on day 5 after birth (10%) as compared with the percentage in the non-insulin-treated diabetics (STZ) (67%). On the 14th day, the body weight and the pancreatic insulin content of insulin-treated animals (STZ + I) were significantly higher than the corresponding values in the STZ animals. Glucose tolerance tests performed sequentially indicated that from 21 days to 7 mo, the plasma insulin response in the (STZ + I) females was clearly increased as compared with that observed in the STZ group. However, it did not reach the insulin response of the controls except in the 21-day-old females and, as a function of age, it declined progressively at variance with the normal age-related pattern. These findings indicate that insulin treatment (sufficient to reduce daily glycosuria) applied during the overtly diabetic period markedly improved the recovery of the insulin stores in the pancreas. Moreover, the long-term effect of the treatment was a long lasting if not permanent improvement of the in vivo insulin response to glucose.

Early development of beta-cells is impaired in the GK rat model of type 2 diabetes.

The Goto-Kakisaki (GK) rat is a genetic model of type 2 diabetes obtained by selective inbreeding of mildly glucose-intolerant Wistar rats. Previous studies have shown that at birth, the beta-cell mass of the GK rat is severely reduced compared with that of the Wistar rat. Therefore, beta-cell deficit could be the primary defect leading to type 2 diabetes in this model. To identify the abnormality at the origin of the beta-cell mass deficit, we compared the fetal development of GK and Wistar rats. Our study reveals that during early development (embryonic day 12-14 [E12-14]), GK fetuses present a delayed global growth that progressively recovers: at birth, no size or weight difference persists. However, from E18 onward, the weight and DNA content of the pancreas and liver are reduced by 30% in the GK fetuses. Cell proliferation is reduced in the GK pancreas from E16 to E20. Whereas apoptotic cells are scarce in the Wistar fetal pancreas, a wave of apoptosis from E16 to E18 was detected in the GK pancreas. Analysis of pancreas differentiation revealed that from E12 to E14, there are no significant differences in the number of alpha- and beta-cells between the GK and Wistar pancreas. However, by E16, the average number of beta-cells in the GK pancreas represents only 50% that of the Wistar pancreas, and this difference persists until birth. The number of alpha-cells was reduced by 25% from E18 to E21. To determine whether the defect in GK pancreas development depends on intrinsic pancreatic factors or on endocrine extrapancreatic factors, we performed in vitro cultures of E12 pancreatic rudiments. The cultures show that in vitro, the growth and endocrine differentiation of the GK and Wistar pancreatic rudiments are identical. Thus, impaired development of the GK pancreas probably results from insufficiency of extrapancreatic factor(s) necessary for the growth and survival of fetal pancreatic cells.

Dynamics of glucose-induced insulin release during the spontaneous remission of streptozocin diabetes induced in the newborn rat.

Neonatal rats injected with streptozocin (STZ, 100 mg/kg) at birth exhibited an acute diabetes followed by a spontaneous remission. We have previously shown that this recovery from neonatal diabetes is due to B-cell regeneration and reaccumulation of pancreatic insulin stores starting from 3 to 5 days after birth. The B-cell population during this period is heterogeneous with both surviving B-cells that have escaped the toxic effect of STZ and newly formed B-cells. To evaluate to what extent this B-cell population is functionally normal, we have measured in vitro the dynamics of glucose-induced insulin secretion from pancreatic fragments of rats treated with STZ at birth. The insulin responses were tested at intervals after STZ treatment, from day 1 to day 21 using perifusion of pancreatic fragments, and on day 21 and at 5 mo using perfusion of isolated pancreas. While the glucose-induced insulin release was completely obtunded (2% of the normal response) on day 1 after STZ, it could be demonstrated after day 3. Moreover, it increased as a function of age (6% and 36% of the normal responses on day 5 and day 14, respectively). This restoration of the insulin response to glucose closely paralleled the recovery of pancreatic insulin stores (6% and 51% of normal values, respectively, on day 5 and day 14). In sharp contrast with the lack of glucose response in vitro in the adult, glucose-induced insulin release was still detected on day 21 regardless of the in vitro system used (perifusion or perfusion). Furthermore, on day 21 the B-cells of the STZ rats exhibited a tendency toward enhanced insulin response to arginine, which is a prominent feature in adult rats previously treated with STZ at birth.(ABSTRACT TRUNCATED AT 250 WORDS)

In vivo hepatic and peripheral insulin sensitivity in rats with non-insulin-dependent diabetes induced by streptozocin. Assessment with the insulin-glucose clamp technique.

Non-insulin-dependent diabetes mellitus (NIDDM) was obtained in adult female rats by neonatal administration of streptozocin (STZ). At 2 mo of age, the basal plasma glucose values in the postabsorptive state were elevated, the glucose disappearance rate measured after intravenous (i.v.) glucose load was significantly lower in the diabetic than in control rats, and in vivo glucose-induced insulin release was drastically reduced. To quantify and characterize the in vivo insulin sensitivity in rats with NIDDM, we have used the insulin-glucose clamp technique. The effects of different concentrations of insulin on glucose production, glucose utilization, and glucose clearance (measured by using 3-3H-glucose) were studied in anesthetized diabetic or control rats while in the postabsorptive state. An inherent condition to set up a valid experimental design was to take into consideration, in the diabetics, the influence of the high blood glucose concentration on glucose uptake and glucose production, since the blood glucose concentration by itself affects these two parameters by a mass action effect independent of insulin. The issue was addressed by evaluating glucose production and utilization in three experimental groups: diabetics clamped at their basal blood glucose level (170 mg/dl), controls clamped at their basal blood glucose level (110 mg/dl), and controls clamped at high blood glucose level (170 mg/dl). In the basal state, glucose production was significantly higher in the diabetics than in controls. When plasma insulin was clamped at submaximal levels (300 microU/ml), the suppression of glucose production was significantly more important in the diabetics than in the two control groups.(ABSTRACT TRUNCATED AT 250 WORDS)

Relation of insulin deficiency to impaired insulin action in NIDDM adult rats given streptozocin as neonates.

We assessed the impact of chronic insulin deficiency on basal and insulin-stimulated glucose utilization by the whole-body mass in vivo in female albino Wistar rats. This assessment was based on a comparison of results in rats given streptozocin (STZ) on day of birth (n0-STZ), when 2 days old (n2-STZ), or when 5 days old (n5-STZ). At 10 wk of age, the n2-STZ rats exhibited characteristics similar to those obtained in the n0-STZ rats: normal growth, modest elevation of basal plasma glucose (8.23 +/- 0.24 mM), glucose intolerance, depleted pancreatic insulin stores (approximately 50% of normal value), and lack of insulin release in response to glucose in vivo. In contrast, the n5-STZ rats exhibited frank basal hyperglycemia (glucose 11.9 +/- 1.1 mM) and glucose intolerance, increased glycosylated hemoglobins, strong reduction of the pancreatic insulin stores (10% of normal value), decreased basal plasma insulin levels (50% of normal value), and lack of insulin release in response to glucose in vivo. Changes in the sensitivity of the neonatal beta-cell to STZ and the regeneration capacity of the beta-cells during the 1st postnatal wk were liable factors for the contrast. In vivo insulin action was assessed with the euglycemic-hyperinsulinemic clamp technique in 10-wk-old anesthetized animals. In the n2-STZ rats compared with controls 1) endogenous glucose production was significantly higher despite a normal plasma insulin level in the basal state, 2) endogenous glucose production rate was similarly suppressed by hyperinsulinemia, and 3) glucose utilization by the whole-body mass was similarly increased by hyperglycemia.(ABSTRACT TRUNCATED AT 250 WORDS)

Insulin treatment improves glucose-induced insulin release in rats with NIDDM induced by streptozocin.

Insulin-deficient diabetes in humans, as well as in the neonatal streptozocin-induced rat model of non-insulin-dependent diabetes mellitus (NIDDM), are associated with islet beta-cell insensitivity to glucose. We hypothesized that the chronic hyperglycemia-hypoinsulinemia pattern causes this impairment of the glucose influence on insulin secretion. This study was designed to determine whether the glucose defect could be counteracted by normalizing the diabetic state in rats with NIDDM after insulin therapy. Mixte lente insulin (5 U X kg-1 X day-1) was given daily at 1700 h over 24 h or 5 consecutive days. Insulin secretion was studied the morning after the last insulin injection with the isolated perfused pancreas preparation. Fed basal plasma glucose levels decreased in diabetic rats from 183 +/- 8 to 136 +/- 10 mg/dl after the 1-day insulin treatment and to 135 +/- 5 mg/dl after the 5-day insulin treatment (vs. 116 +/- 3 mg/dl in control rats). Pancreatic insulin stores were not affected by insulin therapy. Although the 1-day insulin treatment did not modify the lack of glucose response in the diabetic rats, the 5-day insulin treatment improved their glucose-induced insulin secretion. Moreover, insulin therapy improved the priming effect of glucose on a second stimulation with glucose. The return of this glucose effect was hardly detectable after the 1-day insulin therapy but was clearly present after the 5-day treatment.(ABSTRACT TRUNCATED AT 250 WORDS)

Insulin administration enhances growth of the beta-cell mass in streptozotocin-treated newborn rats.

We have previously reported that the damage caused by streptozotocin (STZ) administration to the beta-cells in newborn rats was followed by spontaneous recovery from neonatal diabetes. Our present data indicate that STZ administration on the day of birth (day 1) reduced the total beta-cell mass on day 4 to only 10% of the normal value and that after such damage, 27% of the corresponding normal beta-cell mass was spontaneously regained on day 7. During days 4-7, the contribution of the predicted beta-cell growth (due to the replication of preexisting differentiated beta-cells) to the total beta-cell growth represented only 56%. Therefore, recruitment of new beta-cells from a precursor pool indeed represents a significant mechanism for beta-cell regeneration after STZ during this period of life. Here, we report for the first time that 1) insulin therapy from days 2 to 4 did not significantly influence the occurrence of beta-cell damage after STZ administration (total beta-cell mass on day 4 was reduced to 12% of the normal value) and 2) insulin therapy from days 2 to 6 did improve the otherwise spontaneous beta-cell regeneration, since on day 7 total beta-cell mass was 44% of the corresponding normal beta-cell mass. During days 4-7, the contribution of the predicted beta-cell growth to the total beta-cell growth represented only 32% in the insulin-treated STZ group. Finally the insulin-favored regeneration of the beta-cells reflects both an increased replication from differentiated beta-cells and an increased neogenesis from precursor/stem cells, with this last pathway being preferentially activated.

Benfluorex normalizes hyperglycemia and reverses hepatic insulin resistance in STZ-induced diabetic rats.

We have examined the effect of chronic (20 days) oral administration of benfluorex (35 mg/kg) in a rat model of NIDDM, induced by injection of STZ 5 days after birth and characterized by frank hyperglycemia, hypoinsulinemia, and hepatic and peripheral insulin resistance. We assessed the following: 1) basal blood glucose and insulin levels, 2) glucose tolerance and glucose-induced insulin release in vivo and in vitro, and 3) basal and insulin-stimulated in vivo glucose production and glucose utilization, using the insulin-clamp technique in conjunction with isotopic measurement of glucose turnover. The in vivo insulin response of several individual tissues also was evaluated under the steady-state conditions of the clamp, using the uptake of the glucose analogue 2-deoxy-D-glucose as a relative index of glucose metabolism. In the benfluorex-treated diabetic rats, postabsorptive basal plasma glucose levels were decreased (8.1 +/- 0.2 mM compared with 10.5 +/- 0.5 mM in the pair-fed untreated diabetic rats and 6.1 +/- 0.2 mM in the benfluorex-treated nondiabetic rats), whereas the basal and glucose-stimulated intravenous glucose tolerance test plasma insulin levels were not improved. Such a lack of improvement in the glucose-induced insulin release after benfluorex treatment was confirmed under in vitro conditions (perfused pancreas). In the pair-fed untreated diabetic rats, the basal glucose production and overall glucose utilization were significantly increased, and during hyperinsulinemia both liver and peripheral tissues revealed insulin resistance. In the benfluorex-treated diabetic rats, the basal glucose production and basal overall glucose utilization were normalized. After hyperinsulinemia, glucose production was normally suppressed, whereas overall glucose utilization was not significantly improved.(ABSTRACT TRUNCATED AT 250 WORDS)

Experimental chemical diabetes and pregnancy in the rat. Evolution of glucose tolerance and insulin response.

The effect of pregnancy on the course of experimental chemical diabetes (CD) has been studied in the rat. Glucose tolerance tests (0.5 g/kg i.v.) have been performed serially in the virgin state (2 mo), late pregnancy (20.5 day of gestation), and 1 and 2 mo after delivery, in control and in CD female rats. During gestation in the controls basal plasma glucose is decreased, and plasma glucose levels after glucose load, and also lower than levels found in the virgin state. Glucose tolerance is not significantly affected. Nevertheless, glucose-induced insulin secretion in pregnant animals is increased compared with the virgin state. Glucose tolerance remains unchanged 1 and 2 mo postpartum, but insulin response to glucose becomes significantly lower than in the virgin state. In the pregnant CD rats basal plasma glucose is decreased, but plasma glucose levels after glucose load are similar to values found in the virgin state, thus suggesting decreased glucose tolerance. Glucose-induced insulin secretion is increased compared with the virgin state. Glucose tolerance remains deteriorated 1 and 2 mo postpartum, but insulin secretion is no longer significantly different. These findings indicate that in CD female rats glucose tolerance is and remains deteriorated by pregnancy, while in normal female rats it is and remains unchanged. Thus, despite increased insulin response to glucose during late gestation in the CD rats, the diabetogenicity of pregnancy is confirmed with this experimental model.

Glucose insensitivity and amino-acid hypersensitivity of insulin release in rats with non-insulin-dependent diabetes. A study with the perfused pancreas.

Non-insulin-dependent diabetes (NIDDM) was obtained in adult rats following a neonatal streptozotocin injection. Rats with NIDDM exhibited slightly lowered plasma insulin, slightly elevated basal plasma glucose values (less than 200 mg/dl), and low pancreatic insulin stores (50% of the controls). Insulin secretion was studied in this model using the isolated perfused pancreas technique. Insulin response to glucose stimulation over the range 5.5-22 mM was lacking, thus indicating complete loss of B-cell sensitivity to glucose. Even in presence of theophylline, the B-cells remained insensitive to glucose. In contrast, glyceraldehyde elicited an insulin release as important as that obtained in the control pancreata. This could possibly suggest that the B-cell dysfunction in rats with NIDDM involves a block in glucose metabolism in the early steps of glycolysis prior to the triose-phosphate. Mannose stimulated insulin secretion less in the diabetics than in the controls. The insulin secretion obtained in response to isoproterenol indicated that the ability of the adenylcyclase to generate cAMP in the B-cells of the diabetics was not decreased. The insulinotropic actions of acetylcholine and tolbutamide were normal and increased, respectively, as compared with the controls. In the absence of glucose, the B-cells of the diabetics were unexpectedly hypersensitive to arginine and leucine. The alpha-ketoisocaproate effect in the diabetics was not significantly different from that obtained in the controls. The possibility that enhancement of insulin response to leucine in the diabetics might be related to a more active conversion of leucine to ketoisocaproate along the first steps of intraislet leucine metabolism is proposed.

Glucagon-like peptide-1 and exendin-4 stimulate beta-cell neogenesis in streptozotocin-treated newborn rats resulting in persistently improved glucose homeostasis at adult age.

In neonatal Wistar rats injected with streptozotocin (STZ) at birth (n0-STZ model), a recognized model of beta-cell regeneration, we investigated the capacity of early treatment with glucagon-like peptide 1 (GLP-1) or exendin-4 to promote beta-cell regeneration and thereby improve islet function in the long term, when animals become adults. To this end, n0-STZ rats were submitted to GLP-1 or exendin-4 from postnatal day 2 to day 6 only, and their beta-cell mass and pancreatic functions were tested on day 7 and at 2 months. On day 7, both treatments increased body weight, decreased basal plasma glucose, decreased insulinemia, and increased pancreatic insulin content in n0-STZ rats. At the same age, the beta-cell mass, measured by immunocytochemistry and morphometry methods, was strongly increased in n0-STZ/GLP-1 and n0-STZ/Ex rats compared with n0-STZ rats, representing 51 and 71%, respectively, of the beta-cell mass in Wistar rats, whereas n0-STZ beta-cell mass represented only 21% of the Wistar control value. Despite such early improved beta-cell mass, which is maintained at adult age, the basal and glucose-stimulated insulin secretion (in vivo after intravenous glucose load or in vitro using perfused pancreas) were not improved in the 2-month-old n0-STZ rats previously treated with GLP-1 or exendin-4 compared with untreated n0-STZ rats. However, both treated groups significantly exhibited a decreased basal plasma glucose level and an increased plasma glucose clearance rate compared with the 2-month-old untreated n0-STZ group at adult age. These findings in the n0-STZ model indicate for the first time that GLP-1 or exendin-4 applied during the neonatal diabetic period exert both short- and long-term beneficial effects on beta-cell mass recovery and glucose homeostasis. However, the increase in beta-cell mass, which is still present in the adult n0-STZ rats previously treated, contrasts with the poor beta-cell responsiveness to glucose. Further studies are needed to understand the dissociation between beta-cell regeneration and the lack of improvement in beta-cell function.

Beta-cell insensitivity to glucose in the GK rat, a spontaneous nonobese model for type II diabetes.

In early 1988, a colony of GK rats was started in Paris with progenitors issued from F35 of the original colony reported by Goto and Kakisaki. When studied longitudinally up to 8 mo, GK rats showed as early as 1 mo (weaning) significantly higher basal plasma glucose (9 mM) and insulin levels (doubled), altered glucose tolerance (intravenous glucose), and a very poor insulin secretory response to glucose in vivo compared with Wistar controls. Males and females were similarly affected. Studies of in vitro pancreatic function were carried out with the isolated perfused pancreas preparation. Compared with nondiabetic Wistar rats, GK rats at 2 mo showed a significantly increased basal insulin release, no insulin response to 16 mM glucose, and hyperresponse to 19 mM arginine. Pancreatic insulin stores were only 50% of that in Wistar rats. Perfusion of GK pancreases for 50 or 90 min with buffer containing no glucose partially improved the insulin response to 16 mM glucose and markedly diminished the response to 19 mM arginine, whereas the responses by Wistar pancreases were unchanged. These findings are similar to those reported in rats with non-insulin-dependent diabetes induced by neonatal streptozocin administration and support the concept that chronic elevation in plasma glucose may be responsible, at least in part, for the beta-cell desensitization to glucose in this model. The GK rat seems to be a valuable model for identifying the etiology of beta-cell desensitization to glucose.

Insulin production and glucose metabolism in isolated pancreatic islets of rats with NIDDM.

Rats with non-insulin-dependent diabetes mellitus (NIDDM) induced by neonatal injection of streptozocin are known to have a deficient insulin response to glucose. To evaluate to what extent this glucose insensitivity can be attributed to a perturbation of the islet glucose metabolism, we estimated the rates of glucose phosphorylation, glucose utilization, oxygen consumption, and glucose oxidation in islets isolated from normal and NIDDM rats and compared these values with rates of islet insulin biosynthesis and release in vitro. The data confirm that islets from rats with NIDDM display a deficient response to glucose of both insulin biosynthesis and release that is still present after an overnight culture of the islets at 5.5 mM glucose. Furthermore, they show that islets of these rats have 1) normal low- and high-Km glucose-phosphorylating activities and no major alteration of the glucose utilization rate, 2) decreased insulin release in response to glyceraldehyde, 3) decreased rates of basal respiration and glucose oxidation and a markedly reduced stimulation by glucose of both islet oxygen consumption and glucose oxidation, and 4) decreased glucose-stimulated net 45Ca uptake. We conclude that the relative unresponsiveness to glucose of islets from NIDDM rats is associated with, and perhaps due to, a deficient islet glucose metabolism. This defect is not due to gross alterations in the glycolytic pathway but probably reflects alteration in the islet mitochondria function.