Acetylcholine In Diabetic Rats Research Articles

Screening of the active fractions from the Coreopsis tinctoria Nutt. Flower on diabetic endothelial protection and determination of the underlying mechanism

Ethnopharmacological relevanceThe Coreopsis tinctoria Nutt. flower (CTF) has been used traditionally in China for treating hypertension and diabetes as well as reducing body weight and blood fat. However, the vascular protection effect of the CTF has not been studied to date. Aim of the studyThis study aimed to screen and identify bioactive fractions from the CTF with a diabetic endothelial protection effect and to clarify the underlying mechanism. Materials and methodsThe vascular protection effect of Fraction A was studied in high-fat diet and streptozocin-induced diabetic models. The endothelial protection effect of Fraction A-2 was further studied in an in vitro vascular endothelial dysfunction model induced by high glucose. In a high glucose-induced human umbilical vein endothelial cell (HUVEC) model, Fractions A-2-2 and A-2-3 were screened, and their detailed mechanisms of endothelial protection were studied. Liquid chromatography mass spectrometry (LC-MS) was used to identify the main components in Fractions A-2-2 and A-2-3. ResultsFraction A treatment significantly improved the endothelium-dependent vasodilation of the mesenteric artery induced by acetylcholine in diabetic rats. The maximum relaxation was 79.82 ± 2.45% in the control group, 64.36 ± 9.81% in the model group, and 91.87 ± 7.38% in the Fraction A treatment group (P < 0.01). Fraction A treatment also decreased rat tail pressure compared with the model group at the 12th week. The systolic blood pressure was 152.7 5 ± 16.99 mmHg in the control group, 188.50 ± 5.94 mmHg in the model group, and 172.60 ± 14.31 mmHg in the Fraction A treatment group (P < 0.05). The mean blood pressure was 128.50 ± 13.79 mmHg in the control group, 157.00 ± 6.06 mmHg in the model group, and 144.80 ± 11.97 mmHg in the Fraction A treatment group (P < 0.05).In an in vitro vascular endothelium-dependent vasodilation dysfunction model induced by high glucose, Fraction A-2 improved the vasodilation of the mesenteric artery. The maximum relaxation was 82.15 ± 16.24% in the control group, 73.29 ± 14.25% in the model group, and 79.62 ± 13.89% in the Fraction A-2 treatment group (P < 0.05).In a high glucose-induced HUVEC model, Fraction A-2-2 and Fraction A-2-3 upregulated the expression of IRS-1, Akt, and eNOS and increased the levels of p-IRS-1Ser307, p-Akt Ser473, and p-eNOSSer1177 and also decreased the expression of NOX4, TNF-α, IL-6, sVCAM, sICAM, and NF-κB (P < 0.01). With the intervention of AG490 and LY294002, the above effects of Fraction A-2-2 and Fraction A-2-3 were inhibited (P < 0.01). LC-MS data showed that in Fraction A-2-2 and Fraction A-2-3, there were 10 main components: flavanocorepsin; polyphenolic; flavanomarein; isochlorogenic acid A; dicaffeoylquinic acid; coreopsin; marein; coreopsin; luteolin-7-O-glucoside; and 3′,5,5’,7-tetrahydroxyflavanone-O-hexoside. ConclusionThe protective effect of the CTF on diabetic endothelial dysfunction may be due to its effect on the JAK2/IRS-1/PI3K/Akt/eNOS pathway and the related oxidative stress and inflammation. The results strongly suggested that Fraction A-2-2 and Fraction A-2-3 were the active fractions from the CTF, and the CTF might be a potential option for the prevention of vascular complications in diabetes.

Cepharanthine improves glycemic status and slows the progression of renal and vascular injury in streptozotocin‐induced diabetic rats

We have previously demonstrated that Cepharanthine improves glycemic status and reduces renal injury in streptozotocin‐induced diabetic rats via antioxidant and anti‐inflammatory mechanisms. However, the hypoglycemic mechanism of Cepharanthine remains unclear. We hypothesize that Cepharanthine reduces blood glucose in steptozotocin‐induced diabetic rats via restoring pancreatic β‐cells mass and insulin secretion. Diabetes was induced by a single i.p. injection of streptozotocin (50 mg/kg) in male Sprague Dawley rats. Control and diabetic rats were treated with Cepharanthine (10 mg/kg/day, i.p.) for six weeks (n=6–8). Cepharanthine treatment significantly reduced blood glucose levels and increased plasma insulin levels in diabetic rats (blood glucose was 290± 67 in Cepharanthine treated diabetic rats vs. 550±17 mg/dl in non‐treated diabetic rats, P&lt;0.05). The percentage of TUNEL positive cells was significantly elevated in the pancreas of diabetic rats and these changes was reduced by Cepharanthine treatment (P&lt;0.05). Immunohistochemcial analysis revealed a decrease in pancreatic islet‐specific glucose‐6‐phosphatase catalytic subunit related protein (IGRP) and an increase in pancreatic and duodenal homeobox 1 (PDX‐1) expression levels in Cepharanthine treated vs. non treated diabetic rats (P&lt;0.05). Cepharanthine also improved vascular endothelial function as it significantly attenuated the impairment in aortic ring relaxation to acetylcholine in diabetic rats. Moreover, Cepharanthine reduced markers of renal injury and inflammation and slowed the progression of renal fibrosis as evidenced by the reduction in levels of albuminuria, podocalyxin excretion, renal ICAM‐1 and TGF‐β in diabetic rats. The reduction in renal inflammation and injury was associated with decreased in renal caspase 3 expression in Cepharanthine treated diabetic rats. These data suggest that Cepharanthine halts the progression of streptozotocin‐induced diabetic complications in rats via reduced pancreatic β‐cells apoptosis and increased β‐cells maturation and subsequently insulin release probably via the regulation of the PDX‐1 and IGRP transcription factors of pancreatic β‐cells.Support or Funding InformationThe work was supported by bridge funding grant from Augusta University, Augusta, Georgia to Dr. Ahmed A. ElmarakbyThis abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Alteration in amyloid β42, phosphorylated tau protein, interleukin 6, and acetylcholine during diabetes-accelerated memory dysfunction in diabetic rats: correlation of amyloid β42 with changes in glucose metabolism.

BackgroundDiabetes accelerates memory dysfunction in a continuous, slowly pathological process. Studies suggest that the time course of certain biomarkers can characterize the pathological course of the disease to provide information for early intervention. Thus, there is an urgent need for validated biomarkers to characterize the cognitive impairment induced by DM. We aimed to detect changes in cerebrospinal fluid biomarkers such as amyloid β42, phosphorylated tau protein, interleukin 6, and acetylcholine in diabetic rats over time, and to analyse the relationship between diabetes and cognitive impairment.MethodsRats were injected once intraperitoneally with 1% of streptozotocin to establish a diabetic model. Index changes were investigated longitudinally and all were measured at the end of the experiment at day 75. Aβ42, P-tau, IL-6, and ACh levels in CSF, insulin levels in plasma, and Aβ42 levels in plasma and brain tissue were measured by ELISA.ResultsCompared with control, the diabetic model showed ACh in CSF to be decreased by day 15, continuing lower out to day 75. Aβ42 changes in brain and blood showed the same trends but exhibited minima at different time points: day 30 in CSF and day 15 in plasma. After the minimum, Aβ42 in cerebrospinal fluid rose and levelled off lower than in the control group, whereas Aβ42 in plasma rose and went above the controls at day 30, slowly trending upwards for the remainder of the experiment. P-tau protein in CSF in diabetic rats showed an increasing trend, becoming significantly different from the controls at day 60 and day 75. Aβ42 in CSF was strongly negatively correlated with blood glucose at day 15 and was negatively correlated with insulin in serum, particularly at day 45.ConclusionOur longitudinal research model suggest that changes in the measured biomarkers appear before learning and memory impairments do. Aβ42 and ACh in the diabetes model group clearly changed from day 0 to day 45, and then P-tau and IL-6 varied significantly from day 45 to day 75. The reduced ACh levels observed possibly correlated with the factors common to changes in Aβ42, P-tau, and IL-6.

Effects of gemfibrozil treatment on vascular reactivity of streptozotocin-diabetic rat aorta.

The effects of gemfibrozil treatment on plasma lipids, lipid peroxides and vascular reactivity of aorta were investigated in diabetic rats. Rats were divided randomly into two groups: control and diabetic. Diabetes was induced by a single intraperitoneal injection of streptozotocin (45 mg kg(-1)). Twelve weeks after the induction of diabetes, some of the control and diabetic rats were started treatment with gemfibrozil (100 mg kg(-1) daily; gavage) for 2 weeks. Blood glucose, plasma triglyceride, cholesterol, low-density lipoprotein (LDL) cholesterol and thiobarbituric acid reactive substances (TBARS) levels were markedly increased and gemfibrozil treatment restored these parameters in diabetic rats. However high-density lipoprotein (HDL) cholesterol levels did not differ in all experimental groups. In diabetic rats, the endothelium-dependent relaxations to acetylcholine were decreased when compared with control rats. Gemfibrozil treatment restored the endothelium-dependent responses to acetylcholine in diabetic rats. The endothelium-independent relaxation responses to sodium nitroprusside were not altered in all groups. These findings suggest that gemfibrozil treatment has beneficial effects against cardiovascular and metabolic complications of diabetes via its hypolipidaemic and antioxidant properties.

Attenuation of nitric oxide- and prostaglandin-independent vasodilation of retinal arterioles induced by acetylcholine in streptozotocin-treated rats

Diabetes alters retinal hemodynamics, but little is known about the impact of diabetes on the role of endothelium-derived hyperpolarizing factor (EDHF) in the regulation of retinal circulation. Therefore, we examined how diabetes affects the nitric oxide- and prostaglandin-independent vasodilation of retinal arterioles induced by acetylcholine. Male Wistar rats were treated with streptozotocin (80 mg/kg, i.p.) and experiments were performed 6–8 weeks later. Under artificial ventilation, rats were treated with tetrodotoxin (100 μg/kg, i.v.) to eliminate any nerve activity and prevent movement of the eye. Methoxamine was used to maintain adequate systemic circulation. Fundus images were captured by a digital camera that was equipped with a special objective lens. The vasodilator responses of retinal arterioles were assessed by measuring changes in diameters of the vessels. In streptozotocin-induced diabetic rats and the age-matched controls, acetylcholine increased diameters of retinal arterioles in a dose-dependent manner. The vasodilator responses to acetylcholine in diabetic rats were smaller than those in control rats. The nitric oxide- and prostaglandin-independent vasodilation of retinal arterioles observed under treatment with combination of N G-nitro- l-arginine methyl ester (30 mg/kg, i.v.) and indomethacin (5 mg/kg, i.v.) were also attenuated by diabetes. Diabetes did not alter the dilator responses of retinal arterioles to sodium nitroprusside and forskolin. These results suggest that diabetes impairs EDHF-mediated vasodilation of retinal arterioles induced by acetylcholine. The impaired EDHF-mediated vasodilation may contribute to alteration of retinal hemodynamics in diabetes.

Insulin reverses impaired acetylcholine-induced dilatation of the rat basilar artery during diabetes mellitus

The goal of this study was to determine whether treatment of diabetic rats with insulin reverses impaired nitric oxide synthase-dependent reactivity of the basilar artery in vivo. We measured the diameter of the basilar artery in non-diabetic rats, diabetic (streptozotocin; 50–60 mg/kg i.p.) rats, and diabetic rats treated with insulin implants in response to acetylcholine and nitroglycerin before and following topical application of N G-monomethyl- l-arginine ( l-NMMA). Reactivity of the basilar artery was measured 2–3 months after injection of streptozotocin. We found that topical application of acetylcholine (1.0 and 10 μM) produced dose-related dilatation of the basilar artery in non-diabetic rats, which was inhibited (>80%) by topical application of l-NMMA (10 μM). In diabetic rats, the magnitude of vasodilation produced by acetylcholine was significantly less than in non-diabetic rats. Further, topical application of l-NMMA did not affect dilatation of the basilar artery in diabetic rats in response to acetylcholine. In insulin treated diabetic rats, dilatation of the basilar artery in response to acetylcholine was significantly greater than that observed in either non-diabetic or diabetic rats. In contrast, dilatation of the basilar artery in response to nitroglycerin was similar in insulin treated diabetic rats, non-diabetic rats and diabetic rats. Topical application of l-NMMA only partially inhibited dilatation of the basilar artery in response to acetylcholine in diabetic rats treated with insulin. Thus, the findings of this study suggest that impaired acetylcholine-induced dilatation of the basilar artery during diabetes mellitus can be restored by treatment with insulin. Further, it appears that the contribution of cellular pathways, in addition to nitric oxide synthase, may contribute to dilatation of the basilar artery in response to acetylcholine in rats treated with insulin.

Attenuation of Depressor Response Induced by Platelet Activating Factor and Acetylcholine in Streptozotocin-Induced Diabetic Rats.

The in vivo and in vitro effects of platelet-activating factor (PAF, 1-O-hexadecyl-2-acetyl-sn-glycero-3-phosphorylcholine) and acetylcholine (ACh) on vascular relaxation responses were examined in streptozotocin-induced diabetic rats. Intravenous injection of PAF and ACh (0.03 to 10 micrograms/kg) decreased the mean blood pressure in both control and diabetic rats in a dose-dependent fashion. Initial blood pressure in diabetic rats did not significantly differ from that in control rats. However, depressor responses induced by PAF and ACh in diabetic rats were attenuated more than those in control rats. In perfused mesenteric arterial bed preconstricted with methoxamine (10(-5) - 10(-4) M), PAF (10(-11) -3 x 10(-10) M) produced a concentration-dependent relaxation. However, this relaxation was significantly attenuated in the diabetic preparation compared with the control preparation. ACh also produced a concentration-dependent vasodilation in perfused mesenteric arterial bed. The concentration-response curve for the relaxation of the mesenteric arterial bed to ACh in diabetic preparation was shifted to the right compared with that in control preparation. A pretreatment with oxyhaemoglobin (10(-6) M) also shifted the concentration-response curves for relaxation to ACh in both control and diabetic preparation to the right. There was no difference in relaxation induced by sodium nitroprusside between the diabetic and the control preparation.(ABSTRACT TRUNCATED AT 250 WORDS)

Changes in Sensitivity of the Rat Stomach Fundus to Various Drugs in Streptozotocin-Induced Diabetic Rats

The changes in sensitivity of the rat stomach fundus to acetylcholine (ACh), norepinephrine (NE), isoproterenol (ISO) and to vasoactive intestinal peptide (VIP) were examined in control rats and in streptozotocin (STZ)-induced diabetic rats. The dose-response curves for drugs were constructed 8 weeks after treatment with STZ. The dose-response curve for ACh in diabetic rats was shifted to the left as compared to the control curve, whereas the dose-response curves for NE, ISO and VIP were shifted to the right. These results suggest that functional changes in the autonomic nervous systems of the rat stomach fundus may occur in STZ-induced diabetic rats.

Changes in sensitivity of the rat stomach fundus to various drugs in streptozotocin-induced diabetic rats.

The changes in sensitivity of the rat stomach fundus to acetylcholine (ACh), norepinephrine (NE), isoproterenol (ISO) and to vasoactive intestinal peptide (VIP) were examined in control rats and in streptozotocin (STZ)-induced diabetic rats. The dose-response curves for drugs were constructed 8 weeks after treatment with STZ. The dose-response curve for ACh in diabetic rats was shifted to the left as compared to the control curve, whereas the dose-response curves for NE, ISO and VIP were shifted to the right. These results suggest that functional changes in the autonomic nervous systems of the rat stomach fundus may occur in STZ-induced diabetic rats.