Mediation of Endogenous ß-Endorphin in the Plasma Glucose-Lowering Action of Herbal Products Observed in Type 1-Like Diabetic Rats.

Recently, there have been advances in the development of new substances effective in managing diabetic disorders. Opioid receptors couple multiple systems to result in various biological effects, although opioids are best known for analgesia. In the present review, we used our recent data to describe the advance in plasma glucose-lowering action of herbal products, especially the mediation of ß-endorphin in glucose homeostasis of insulin-deficient diabetes. In type 1-like streptozotocin-induced diabetic rats, we identified many products purified from herbs that show a dose-dependent plasma glucose-lowering action. Increase in ß-endorphin secretion from the adrenal gland may activate peripheral opioid µ-receptors (MOR) to enhance the expression of muscle glucose transporters and/or to reduce hepatic gluconeogenesis at the gene level, thereby leading to improved glucose utilization in peripheral tissues for amelioration of severe hyperglycemia. It has also been observed that stimulation of α(1)-adrenoceptors (α(1)-ARs) in the adrenal gland by some herbal products is responsible for the increase in ß-endorphin secretion via a phospholipase C-protein kinase dependent pathway. However, an increase in ß-endorphin secretion from the adrenal gland by herbal products can function via another receptor. New insights into the mediation of endogenous ß-endorphin activation of peripheral MOR by herbal products for regulation of glucose homeostasis without the presence of insulin have been established. Therefore, an increase in ß-endorphin secretion and/or direct stimulation of peripheral MOR via an insulin-independent action might serve as the potential target for development of a therapeutic agent or promising adjuvant in intensive plasma glucose control.

Abelmoschus moschatus (Malvaceae), an aromatic plant, suitable for medical or food uses to improve insulin sensitivity.

Abelmoschus moschatus (Malvaceae) is an aromatic and medicinal plant, distributed in many parts of Asia, including south Taiwan. The present study was undertaken to clarify whether the herb is effective in improving insulin resistance. Insulin resistance in rats was induced by a diet containing 60% fructose for 6 weeks. The degree of insulin resistance was measured by homeostasis model assessment of basal insulin resistance (HOMA-IR). Insulin sensitivity was calculated using the composite whole body insulin sensitivity index (ISIcomp) during the oral glucose tolerance test. Insulin receptor-related signaling mediators in soleus muscles of rats were evaluated by immunoprecipitation or immunoblotting. The extract of A. moschatus had a higher level of polyphenolic flavonoids. A. moschatus extract (200 mg/kg per day) displayed the characteristics of rosiglitazone (4 mg/kg per day) in reducing the higher HOMA-IR index as well as elevating ISIcomp in fructose chow-fed rats after a 2-week treatment. Treatment with moschatus extract for 2 weeks increased post-receptor insulin signaling mediated by enhancements in insulin receptor substrate-1-associated phosphatidylinositol 3-kinase step and glucose transporter subtype 4 translocation in insulin-resistant soleus muscles. A. moschatus is therefore proposed as potentially useful adjuvant therapy for patients with insulin resistance and/or the subjects wishing to increase insulin sensitivity.

Increase of beta-endorphin secretion by syringin, an active principle of Eleutherococcus senticosus, to produce antihyperglycemic action in type 1-like diabetic rats.

We employed streptozotocin-induced diabetic rats (STZ-diabetic rats) as type 1 diabetes-like animal models to investigate the mechanism(s) of antihyperglycemic action produced by syringin, an active principle purified from the rhizome and root part S of ELEUTHEROCOCCUS SENTICOSUS (Araliaceae). Bolus intravenous (i. v.) injection of syringin dose-dependently decreased the plasma glucose of STZ-diabetic rats in 30 minutes in a way parallel to the increase of plasma beta-endorphin-like immunoreactivity (BER). Syringin enhanced BER release from the isolated adrenal medulla of STZ-diabetic rats in a concentration-dependent manner from 0.001 to 10 micromol/l. Bilateral adrenalectomy in STZ-diabetic rats eliminated the activities of syringin (1 mg/kg, i. v.) including the plasma glucose-lowering effect and the plasma BER-elevating effect. Also, syringin failed to lower plasma glucose in the presence of micro-opioid receptor antagonists and/or in the micro-opioid receptor knockout diabetic mice. In conclusion, the obtained results suggest that syringin can enhance the secretion of beta-endorphin from adrenal medulla to stimulate peripheral micro-opioid receptors resulting in a decrease of plasma glucose in diabetic rats lacking insulin.

Ginsenoside Rh2 is one of the active principles of Panax ginseng root to improve insulin sensitivity in fructose-rich chow-fed rats.

Ginsenoside Rh2, one of the ginsenosides contained in the Panax ginseng root, was employed to screen the effect on insulin resistance of rats induced by a diet containing 60% fructose. Single intravenous injection of ginsenoside Rh2 decreased the plasma glucose concentrations in 60 minutes in a dose-dependent manner from 0.1 mg/kg to 1 mg/kg in rats with insulin resistance induced by fructose-rich chow. Repeated intravenous injection of ginsenoside Rh2 (1 mg/kg per injection, 3 times daily) into rats which received fructose-rich chow for 3 consecutive days decreased the value of glucose-insulin index, the product of the areas under the curve of glucose and insulin during the intraperitoneal (i.p.) glucose tolerance test. This means that ginsenoside Rh2 has an ability to improve insulin action on glucose disposal. The plasma glucose lowering action of tolbutamide, induced by the secretion of endogenous insulin, is widely used to characterize the formation of insulin resistance. Time for the loss of plasma glucose lowering response to tolbutamide (10 mg/kg, i.p.) in rats during insulin resistance induction by fructose-rich chow was also markedly delayed by the repeated treatment of ginsenoside Rh2, as compared to the vehicle-treated control. Thus, the repeated treatment of ginsenoside Rh2 delayed the development of insulin resistance in high fructose feeding rats. Increase of insulin sensitivity by ginsenoside Rh2 was further identified using the plasma glucose lowering action of exogenous insulin in streptozotocin-induced diabetic rats (STZ-diabetic rats). Repeated injection of ginsenoside Rh2 at the same dosing (1 mg/kg, 3 times daily) into STZ-diabetic rats for 10 days made an increase of the responses to exogenous insulin. Taken together, it can be concluded that ginsenoside Rh2 has an ability to improve insulin sensitivity and it seems suitable to use ginsenoside Rh2 as an adjuvant for diabetic patients and/or the subjects wishing to increase insulin sensitivity.

Role of tumor suppressor PTEN in tumor necrosis factor alpha-induced inhibition of insulin signaling in murine skeletal muscle C2C12 cells.

In an attempt to clarify the role of phosphatase and tensin homologue deleted on chromosome 10 (PTEN) in muscle insulin resistance, we investigated the effect of PTEN on phosphoinositide 3 (PI3)-kinase/Akt related insulin signaling pathway in skeletal muscle-like C2C12 cells damaged by tumor necrosis factor-alpha (TNFalpha). C2C12 cells cultured with TNFalpha (10 ng/ml) for 1 h displayed a marked decrease of insulin-stimulated 2-[14C]-deoxy-D-glucose (2-DG) uptake in parallel with an elevation of PTEN mRNA and protein levels. However, pretreatment of PTEN antisense oligonucleotide (AS) (1 micromol/l for 3 days) for specific inhibition of PTEN expression in C2C12 cells abolished the TNFalpha-induced changes in 2-DG uptake. Similar pretreatment with PTEN AS, but not with sense oligonucleotide (1 micromol/l for 3 days), eliminated the ability of TNFalpha to impair insulin-stimulated signals including p85 regulatory subunit of PI3-kinase expression and the degree of Akt serine phosphorylation as well as protein expression in glucose transporter subtype 4. Data taken from cultured C2C12 cells emphasize the negative regulatory of muscle PI3-kinase/Akt signaling pathways as the major substrate of PTEN but also support the concept that PTEN contributes to the development of insulin resistance in skeletal muscle.

Mechanism for blockade of angiotensin subtype 1 receptors to lower plasma glucose in streptozotocin-induced diabetic rats.

AIMS: We investigated the mechanism(s) by which valsartan, a selective antagonist of angiotensin subtype 1 (AT(1)) receptor, decreased plasma glucose in streptozotocin (STZ)-induced diabetic rats. METHODS: The plasma glucose concentration was assessed by the glucose oxidase method. The concentration of beta-endorphin in plasma or medium incubating adrenal medulla was measured using an enzyme-linked immunosorbent assay. The mRNA levels of the subtype 4 form of glucose transporter (GLUT4) in soleus muscle and phosphoenolpyruvate carboxykinase (PEPCK) in the liver were detected by Northern blotting analysis, while the protein levels of GLUT4 in isolated soleus muscle and hepatic PEPCK were investigated using Western blotting analysis. RESULTS: A single intravenous injection of valsartan dose-dependently increased plasma beta-endorphin-like immunoreactivity (BER) in parallel with the lowering of plasma glucose concentration in STZ-induced diabetic rats. Naloxone and naloxonazine inhibited the plasma glucose-lowering action of valsartan at doses sufficient to block opioid micro-receptors. In contrast to its action in wild-type diabetic mice, valsartan failed to modify plasma glucose in opioid micro-receptor knockout diabetic mice. Bilateral adrenalectomy in STZ-induced diabetic rats eliminated both the plasma glucose-lowering action and the plasma BER-elevating action of valsartan. In the isolated adrenal medulla of STZ-induced diabetic rats, angiotensin II (Ang II) or valsartan did not affect spontaneous BER secretion. Activation of cholinergic receptors by 1.0 micromol/l acetylcholine (ACh) enhanced BER secretion from the isolated adrenal medulla of STZ-induced diabetic rats, but not in the presence of 1.0 nmol/l Ang II, while valsartan reversed this inhibition by Ang II in a concentration-dependent manner. Treatment of STZ-induced diabetic rats with valsartan (0.2 mg/kg) three times daily for 3 days resulted in an increase in gene expression of GLUT4 in soleus muscle and impeded the reduction of elevated mRNA or protein level of hepatic PEPCK. Both of these effects were blocked by opioid micro-receptor antagonist. CONCLUSIONS: The results suggest that blockade of AT(1) receptor by valsartan may enhance the adrenal beta-endorphin secretion induced by ACh, activating the opioid micro-receptors to increase glucose utilization and/or to decrease hepatic gluconeogenesis, resulting in the reduction of plasma glucose in STZ-induced diabetic rats.

Plasma glucose-lowering action of Hon-Chi in streptozotocin-induced diabetic rats.

Hon-Chi was used for anti-hyperglycemic activity screening in streptozotocin-induced diabetic rats (STZ-diabetic rats) in an attempt to develop new substances for handling diabetes. Mandarin Hon-Chi is red yeast rice fermented with Monascus pilous and Monascus purpureus. Single oral administration of Hon-Chi decreased plasma glucose in STZ-diabetic rats in a dose-dependent manner from 50 mg/kg to 350 mg/kg. Similar treatment with Hon-Chi also lowered the plasma glucose in normal rats as effectively as that produced in STZ-diabetic rats. In addition, oral administration of Hon-Chi at the highest dose (350 mg/kg) attenuated the elevation of plasma glucose induced by an intravenous glucose challenge test in normal rats. Moreover, mRNA levels of phosphoenolpyruvate carboxykinase (PEPCK) in liver from STZ-diabetic rats were reversed in a dose-dependent manner by the repeated oral treatment of Hon-Chi three times daily for two weeks. Otherwise, hyperphagia in STZ-diabetic rats was markedly reversed by similar repeated treatment of Hon-Chi. The obtained results suggest that oral administration of Hon-Chi could decrease hepatic gluconeogenesis to lower plasma glucose in diabetic rats lacking insulin.

Suppression of transforming growth factor-beta1 gene expression by Danggui buxue tang, a traditional Chinese herbal preparation, in retarding the progress of renal damage in streptozotocin-induced diabetic rats.

Danggui buxue tang (DBT), a preparation containing Angelica sinensis (danggui) and Astragalus membranaceus (huangqi) at a ratio of 1 : 5, is used widely in China for stimulating red blood cell production and enhancing cardiovascular function. The present study was undertaken to characterize the effects of this preparation on diabetic nephropathy using streptozotocin-diabetic rats as a model. Streptozotocin-dependent alterations in renal weight/body weight ratio, urinary albumin and beta (2)-microglobulin concentrations, urinary albumin excretion rate, and creatinine clearance were ameliorated after eight weeks of treatment with either DBT or the angiotensin-converting enzyme inhibitor, benazepril. DBT, but not benazepril, partially attenuated the increases in blood glucose, triglycerides and cholesterol in STZ-diabetic rats. Additionally, the increased expression of transforming growth factor-beta (1) mRNA in the renal cortex due to streptozotocin-induced diabetes was modestly attenuated by these treatments. However, eight weeks of treatment with DBT failed to modify the concentration of angiotensin II in plasma or kidney, indicating that the ability of the preparation to retard the progression of kidney disease was not attributable to inhibition of the renin-angiotensin system. We propose that DBT alleviates renal alterations in diabetes and slows the progression of diabetic nephropathy by suppressing transforming growth factor-beta (1) mRNA expression. The preparation may therefore be useful as an adjuvant therapy for controlling diabetes and its complications.

Increase of insulin sensitivity by stevioside in fructose-rich chow-fed rats.

The intake of dietary fructose has undergone a marked increase around the world, especially the developed countries, in recent times. Stevioside, a glycoside contained in the leaves of Stevia rebaudiana Bertoni (Compositae), was used to screen the effect induced by a diet containing 60% fructose on insulin resistance in rats. Single oral administration of stevioside for 90 min decreased plasma glucose concentrations in a dose-dependent manner in rats receiving fructose-rich chow for four weeks. In addition, insulin action on glucose disposal rate was measured using the glucose-insulin index, the product of the areas under the curve of glucose, and insulin during the intraperitoneal glucose tolerance test. Oral administration of stevioside (5.0 mg/kg) in rats given four weeks of fructose-rich chow for 90 min reversed the value of glucose-insulin index, indicating that stevioside has the ability to improve insulin sensitivity in this insulin-resistant animal model. Time for the loss of plasma glucose lowering response to tolbutamide (10.0 mg/kg, i. p.) in fructose-rich chow fed rats was also markedly delayed by repeated stevioside treatment three times daily compared to the vehicle-treated group. The plasma glucose-lowering activity of tolbutamide was introduced to account for varying levels of endogenous insulin secretion, and is widely used as the indicator of insulin resistance development. Thus, it provided the supportive data that repeated oral administration of stevioside delayed the development of insulin resistance in rats on a high-fructose diet. Increased insulin sensitivity by stevioside administration was further identified using the plasma glucose-lowering action of exogenous insulin in streptozotocin-induced diabetic rats (STZ-diabetic rats). Oral administration of stevioside at 0.2 mg/kg three times daily into STZ-diabetic rats for ten days increased the response to exogenous insulin. Taken together, this demonstrated that oral administration of stevioside improves insulin sensitivity, and seems suitable as an adjuvant for diabetic patients and/or those that consume large amounts of fructose.

Activation of opioid mu-receptors by loperamide to improve interleukin-6-induced inhibition of insulin signals in myoblast C2C12 cells.

AIMS/HYPOTHESIS: This study investigated the role of opioid mu-receptor activation in the improvement of insulin resistance. METHODS: Myoblast C2C12 cells were cultured with IL-6 to induce insulin resistance. Radioactive 2-deoxyglucose (2-DG) uptake was used to evaluate the effect of loperamide on insulin-stimulated glucose utilisation. Protein expression and phosphorylation in insulin-signalling pathways were detected by immunoblotting. RESULTS: The insulin-stimulated 2-DG uptake was reduced by IL-6. Loperamide reversed this uptake, and the uptake was inhibited by blockade of opioid mu-receptors. Insulin resistance induced by IL-6 was associated with impaired expression of the insulin receptor (IR), IR tyrosine autophosphorylation, IRS-1 protein content and IRS-1 tyrosine phosphorylation. Also, an attenuated p85 regulatory subunit of phosphatidylinositol 3-kinase, Akt serine phosphorylation and the protein of glucose transporter subtype 4 were observed in insulin resistance. Loperamide reversed IL-6-induced decrement of these insulin signals. CONCLUSIONS/INTERPRETATION: Opioid mu-receptor activation may improve IL-6-induced insulin resistance through modulation of insulin signals to reverse the responsiveness of insulin. This provides a new target in the treatment of insulin resistance.

Improvement of insulin resistance by panax ginseng in fructose-rich chow-fed rats.

In an attempt to probe a new target for handling insulin resistance, we used Panax ginseng root to screen the effect on insulin resistance induced by fructose-rich chow in rats. Insulin action on glucose disposal rate was measured using the glucose-insulin index, which is the product of the areas under the curve of glucose and insulin during the intraperitoneal glucose tolerance test. Oral administration of Panax ginseng root (125.0 mg/kg) into rats three times daily for three days after receiving fructose-rich chow for four weeks reversed the increased glucose-insulin index, indicating that Panax ginseng root has the ability to improve insulin sensitivity. In addition, the plasma glucose concentrations in rats repeatedly treated with Panax ginseng root were not elevated as markedly as those of the vehicle-treated group during the fructose-rich chow-feeding period. Also, the time in which the plasma glucose-lowering response to tolbutamide (10.0 mg/kg, i. p.) receded in fructose-rich chow fed rats was markedly delayed by repeated Panax ginseng root treatment compared to the vehicle-treated group. The plasma glucose-lowering activity of tolbutamide is believed to depend on the secretion of endogenous insulin, which is widely used as an indicator of insulin resistance development. Thus, it provided supportive data that oral administration of Panax ginseng root could delay the development of insulin resistance in rats. In conclusion, our results suggest that oral administration of Panax ginseng root improves insulin sensitivity and may be used as an adjuvant therapy for treating diabetic patients with insulin resistance.

Activation of imidazoline receptors in adrenal gland to lower plasma glucose in streptozotocin-induced diabetic rats.

AIMS/HYPOTHESIS: The present study investigated the effect of agmatine, an endogenous ligand of imidazoline receptors, on plasma glucose in streptozotocin-induced diabetic rats (STZ-diabetic rats). METHODS: Plasma glucose was assessed by the glucose oxidase method. Plasma insulin and beta-endorphin-like immunoreactivity in plasma or adrenal medulla were measured by enzyme-linked immunosorbent assay. Systolic blood pressure was determined by the tail-cuff method. The mRNA levels of glucose transporter subtype 4 (GLUT4) in soleus muscle and phosphoenolpyruvate carboxykinase (PEPCK) in liver were detected by northern blotting. Protein levels of GLUT4 in soleus muscle and hepatic PEPCK were estimated using western blotting analysis. RESULTS: After intravenous injection into fasting STZ-diabetic rats for 30 min, agmatine decreased plasma glucose in a dose-dependent manner without changing systolic blood pressure. At the same time, plasma beta-endorphin-like immunoreactivity also increased in STZ-diabetic rats receiving the same treatment. Plasma glucose was significantly elevated in STZ-diabetic rats by an intravenous injection of clonidine at a dose sufficient to decrease systolic blood pressure. Involvement of I(1)-imidazoline receptors and/or alpha2-adrenoceptors in this effect of agmatine was thus unlikely. The lowering of plasma glucose and increase of plasma beta-endorphin-like immunoreactivity by agmatine were abolished by pretreating the rats with BU-224 at a dose sufficient to block I(2)-imidazoline receptors. Both effects of agmatine were also abolished in adrenalectomised STZ-diabetic rats. Moreover, agmatine enhanced beta-endorphin-like immunoreactivity release from the isolated adrenal medulla of STZ-diabetic rats, an effect also blocked by BU-224. Release of beta-endorphin from the adrenal glands by I(2)-imidazoline receptor activation seems responsible for the plasma glucose-lowering action of agmatine. This was supported by the fact that intravenous injection of naloxone or naloxonazine at doses sufficient to block opioid mu-receptors inhibited the action of agmatine. In addition to lowering plasma glucose, repeated intravenous injection of agmatine into STZ-diabetic rats for 4 days also increased mRNA and protein levels of GLUT4 in soleus muscle. The same treatment also reversed the higher mRNA and protein levels of PEPCK in liver of STZ-diabetic rats. CONCLUSIONS/INTERPRETATION: Our results suggest that agmatine may activate I(2)-imidazoline receptors in the adrenal gland. This enhances secretion of beta-endorphin, which can activate opioid mu-receptors to increase GLUT4 gene expression and/or suppress hepatic PEPCK gene expression, resulting in a lowering of plasma glucose in diabetic rats lacking insulin. The results provide a potential new target for intervention in type 1 diabetes.

Mediation of beta-endorphin in exercise-induced improvement in insulin resistance in obese Zucker rats.

BACKGROUND: Aerobic exercise including treadmill running has long been used to successfully treat and/or prevent insulin resistance and type-2 diabetes. Increase of plasma beta-endorphin is observed with exercise. The present study was designed to clarify the role of endogenous beta-endorphin in exercise-induced improvement in insulin resistance. METHODS: We used a moderate exercise program consisting of treadmill running at 20 m/min and 0% grade for 1 h/day, 7 days/week, for 8 weeks. Plasma glucose concentration was assessed by the glucose oxidase method. The enzyme-linked immunosorbent assay was performed to quantify the plasma level of beta-endorphin-like immunoreactivity (BER). The glucose disposal rate (GDR) was measured by the hyperinsulinemic euglycemic clamp technique. Changes of the insulin signaling in isolated soleus muscle were then detected by immunoprecipitation and immunoblotting. RESULTS: An increase of plasma BER in parallel with the reduction of plasma glucose was obtained in exercise-trained obese Zucker rats. Different from a marked reduction in sedentary obese rats, the value of insulin-stimulated GDR obtained from the exercised obese rats was reversed to near that of the sedentary lean group, eight weeks after the last period of exercise. This effect of exercise was inhibited by naloxone or naloxonazine at doses sufficient to block opioid micro-receptors. Signaling-related defects in the soleus muscle of sedentary obese Zucker rats, which impaired glucose transporter subtype 4 (GLUT 4), included decreased phosphorylation of insulin receptor substrate (IRS)-1, as well as an attenuated p85 regulatory subunit of phosphatidylinositol 3-kinase (PI3 kinase) and Akt serine phosphorylation. In contrast, exercise training failed to modify the levels of insulin receptor (IR), IRS-1, and IR tyrosine autophosphorylation in obese Zucker rats. CONCLUSION: Enhanced insulin sensitivity via exercise training might be mediated by endogenous beta-endorphin through an increase of postreceptor insulin signaling related to the IRS-1-associated PI3-kinase step that leads to the enhancement of GLUT 4 translocation and improved glucose disposal in obese Zucker rats.

Increase of PTEN gene expression in insulin resistance.

Phosphatase and tensin homologue deleted from chromosome ten (PTEN) has recently been characterized as a regulator of insulin sensitivity in the insulin target tissue. However, whether PTEN gene expression is changed in insulin resistance remains unclear. We observed that both the mRNA and protein level of PTEN in soleus muscle isolated from the obese Zucker rats (Fa/Fa) were increased compared to the age-matched lean group. Similarly, both the mRNA and protein level of PTEN in soleus muscle of the fructose-fed lean Zucker rats (Fa/Fa) showing the higher glucose-insulin index were higher than that of the regular chow fed group. These results suggest that increase of PTEN gene expression seems to be related to the development of insulin resistance.

Infusion of beta-endorphin improves insulin resistance in fructose-fed rats.

In an attempt to probe the effect of beta-endorphin on insulin resistance, we used Wistar rats that were fed fructose-rich chow to induce insulin resistance. Insulin action on glucose disposal rate (GDR) was measured using the hyperinsulinemic euglycemic clamp technique, in which glucose (variable), insulin (40 mU/kg/min), and beta-endorphin (6 ng/kg/min) or vehicle were initiated simultaneously and continued for 120 min. A marked reduction in insulin-stimulated GDR was observed in fructose-fed rats compared to normal control rats. Infusion of beta-endorphin reversed the value of GDR, which was inhibited by naloxone and naloxonazine each at doses sufficient to block opioid mu-receptors. Opioid mu-receptors may therefore be activated by beta-endorphin to improve insulin resistance. Next, soleus muscle was isolated to investigate the effect of beta-endorphin on insulin signals. Insulin resistance in rats induced by excess fructose was associated with the impaired insulin receptor (IR), tyrosine autophosphorylation, and insulin receptor substrate (IRS)-1 protein content in addition to the significant decrease in IRS-1 tyrosine phosphorylation in soleus muscle. This impaired glucose transportation was also due to signaling defects that included an attenuated p85 regulatory subunit of phosphatidylinositol 3-kinase (PI3-kinase) and Akt serine phosphorylation. However, IR protein levels were not markedly changed in rats with insulin resistance. beta-endorphin infusion reversed the fructose-induced decrement in the insulin-signaling cascade with increased GDR. Apart from IR protein levels, infusion of beta-endorphin reversed the decrease in protein expression for the IRS-1, p85 regulatory subunit of PI3-kinase, and Akt serine phosphorylation in soleus muscle in fructose-fed rats. The decrease in insulin-stimulated protein expression of glucose transporter subtype 4 (GLUT 4) in fructose-fed rats returned to near-normal levels after beta-endorphin infusion. Infusion of beta-endorphin may improve insulin resistance by modulating the insulin-signaling pathway to reverse insulin responsiveness.

Signals in the activation of opioid mu-receptors by loperamide to enhance glucose uptake into cultured C2C12 cells.

In an attempt to understand the signal pathways of opioid mu-receptors for glucose metabolism, we used loperamide to investigate the glucose uptake into the myoblast C2C12 cells. Loperamide enhanced the uptake of radioactive deoxyglucose into C2C12 cells in a concentration-dependent manner that was abolished in cells pre-incubated with naloxone or naloxonazine at concentrations sufficient to block opioid mu-receptors. Pharmacological inhibition of phospholipase C (PLC) by U73122 resulted in a concentration-dependent decrease in loperamide-stimulated uptake of radioactive deoxyglucose into C2C12 cells. This inhibition of glucose uptake by U73122 was specific since the inactive congener, U73343, failed to modify loperamide-stimulated glucose uptake. Moreover, both chelerythrine and GF 109203X diminished the action of loperamide at concentrations sufficient to inhibit protein kinase C (PKC). The obtained data suggest that an activation of opioid mu-receptors in C2C12 cells by loperamide may increase glucose uptake via the PLC-PKC pathway.

Loperamide increases glucose ultilization in streptozotocin-induced diabetic rats.

1. Loperamide has an ability to lower the plasma glucose concentration in streptozotocin (STZ)-induced diabetic rats. In the present study, we investigated the molecular mechanisms by which loperamide regulates plasma glucose concentrations in the absence of insulin. 2. Loperamide, at a dose sufficient (17.6 microg/kg) to activate mu-opioid receptors, significantly decreased plasma glucose levels in STZ-diabetic rats. The mRNA and protein levels of glucose transporter 4 (GLUT-4) in soleus muscle, detected by northern and western blotting, respectively, were increased after repeated intravenous administration of loperamide (17.6 micro g/kg) to STZ-diabetic rats over 3 days. Moreover, similar treatment with loperamide (17.6 microg/kg) for 3 days reversed the elevated mRNA and protein levels of phosphoenolpyruvate carboxykinase (PEPCK) in the liver of STZ-diabetic rats to near the levels seen in normal rats. 3. These results suggest that activation of mu-opioid receptors by loperamide can increase glucose utilization in peripheral tissues and/or reverse the higher gene expression of PEPCK to inhibit hepatic gluconeogenesis, thereby lower plasma glucose in diabetic rats lacking insulin.

Increase in adenosine A1 receptor gene expression in the liver of streptozotocin-induced diabetic rats.

BACKGROUND: Adenosine A1 receptor (A1-AR) activation can lower plasma glucose in diabetic rats lacking insulin. We investigated the change in A1-AR gene expression in diabetic rats. METHODS: The incorporation of [U-(14)C]-glucose into glycogen was carried out to evaluate the effect of N(6)-cyclopentyladenosine (CPA) on glucose utilization in vitro. The plasma glucose concentration was assessed by the glucose oxidase method. The mRNA and protein levels of A1-AR in isolated liver were detected by Western blotting analysis and Northern blotting analysis, respectively. RESULTS: The effect of CPA, an agonist of A1-AR, on glycogen incorporation in hepatocytes isolated from streptozotocin-induced diabetic rats (STZ-diabetic rats) was more marked than that from the normal rats. However, similar glycogen synthesis was not modified by 12-O-tetradecanoylphorbol-13-acetate (TPA), an activator of protein kinase C, in the isolated hepatocytes from both the normal and the STZ-diabetic rats. A change in response at the receptor level can thus be considered. The mean level of liver mRNA transcripts encoding A1-AR was increased in STZ-diabetic rats to about 250% of that in normal rats. Exogenous insulin at a dose sufficient to normalize the plasma glucose of STZ-diabetic rats reversed the mRNA level of A1-AR in the liver after a four-day treatment. Similar results were also observed in STZ-diabetic rats that received treatment with phlorizin for four days. Moreover, the protein level of A1-AR was higher in the liver of STZ-diabetic rats than that in the normal rats. Similar treatment with exogenous insulin or phlorizin reversed the elevated protein level of A1-AR in the liver of STZ-diabetic rats to near the normal level. Therefore, correction of hyperglycemia in STZ-diabetic rats can reverse the higher gene expression of A1-AR in liver. CONCLUSIONS: The obtained results suggest that an increase in plasma glucose is responsible for the higher gene expression of A1-AR in the liver of STZ-diabetic rats.

Release of beta-endorphin by caffeic acid to lower plasma glucose in streptozotocin-induced diabetic rats.

The role of alpha 1A -adrenoceptors in the regulation of opioid secretion from the adrenal glands of streptozotocin-induced diabetic rats (STZ-diabetic rats) was examined in an attempt to determine the mechanism of plasma glucose-lowering action of caffeic acid. In agreement with a previous report, we showed that caffeic acid produced a dose-dependent lowering of the plasma glucose concentration in STZ-diabetic rats along with an increase of plasma beta-endorphin-like immunoreactivity (BER). These actions of caffeic acid were abolished by pretreatment with WB 4101 or RS 17 056 at doses sufficient to block alpha 1A -adrenoceptors. In addition, naloxone and naloxonazine at doses effective for blocking opioid micro -receptors abolished the plasma glucose-lowering action of caffeic acid. Also, unlike that in wild-type diabetic mice, caffeic acid failed to produce a plasma glucose lowering effect in opioid micro -receptor knockout diabetic mice. We observed that caffeic acid could enhance BER release from isolated rat adrenal medulla in a concentration-dependent manner; inhibitors of alpha 1A -adrenoceptors such as WB 4101 and RS 1705 abolished this action. Investigations of the signal pathways further supported that activation of alpha 1A -adrenoceptor is responsible for the stimulatory effect of caffeic acid on BER secretion from the adrenal medulla. In the presence of U73312, a specific inhibitor of phospholipase C, the caffeic acid-induced increase of BER was reduced in a concentration-dependent manner, but it was not affected by U73343, the negative control of U73312. Chelerythrine and GF 109203X also diminished the action of caffeic acid at concentrations sufficient for inhibiting protein kinase C. Moreover, bilateral adrenalectomy in STZ-diabetic rats resulted in the loss of this plasma glucose-lowering effect of caffeic acid, and there was no increase in plasma BER with caffeic acid. Therefore, beta-endorphin release from the adrenal gland appears to be responsible for the lowering of plasma glucose in STZ-diabetic rats induced by caffeic acid, through the activation of alpha 1A -adrenoceptors.

Plasma glucose-lowering effect of beta-endorphin in streptozotocin-induced diabetic rats.

The effect of beta-endorphin on plasma glucose levels was investigated in streptozotocin-induced diabetic rats (STZ-diabetic rats). A dose-dependent lowering of plasma glucose was observed in the fasting STZ-diabetic rat fifteen minutes after intravenous injection of beta-endorphin. The plasma glucose-lowering effect of beta-endorphin was abolished by pretreatment with naloxone or naloxonazine at doses sufficient to block opioid mu-receptors. Also, unlike wild-type diabetic mice, beta-endorphin failed to induce its plasma glucose-lowering effect in the opioid mu-receptor knock-out diabetic mice. In isolated soleus muscle, beta-endorphin enhanced the uptake of radioactive glucose in a concentration-dependent manner. Stimulatory effects of beta-endorphin on glycogen synthesis were also seen in hepatocytes isolated from STZ-diabetic rats. The blockade of these actions by naloxone and naloxonazine indicated the mediation of opioid mu-receptors. In the presence of U73312, the specific inhibitor of phospholipase C (PLC), the uptake of radioactive glucose into isolated soleus muscle induced by beta-endorphin was reduced in a concentration-dependent manner, but it was not affected by U73343, the negative control of U73312. Moreover, chelerythrine and GF 109203X diminished the stimulatory action of beta-endorphin on the uptake of radioactive glucose at a concentration sufficient to inhibit protein kinase C (PKC). The data obtained suggest that activating opioid mu-receptors by beta-endorphin may increase glucose utilization in peripheral tissues via the PLC-PKC pathway to lower plasma glucose in diabetic rats lacking insulin.