| Objective: Diabetes mellitus(DM)is a metabolic disorder characterized by chronic hyperglycemia caused by various causes.It has become the third largest disease in the world following tumors,cardiovascular and cerebrovascular diseases and is a serious threat to human health.Diabetes can cause multiple system damage with the prolongation of the disease course,leading to tissue lesions and dysfunction,which seriously affects the quality of life of diabetic patients.The current role of Ghrelin and Obestatin in diabetic rats is mostly concentrated on their effects on the peripheral digestive system and regulation of blood glucose levels.So far,there are few reports on the effects of central ghrelin and obestatin on gastric motility and food intake in diabetic rats.What are the effects of Ghrelin and Obestatin on gastric motility and feeding in diabetic rats and what’s the possible mechanism? Are there hippocampus/ arcuate nucleus of hypothalamus(ARC)Ghrelin / Obestatin neural pathways in brain and are there any changes in these pathways in early stage of diabetic state? What is the role of this change in the disorders in feeding and gastric function in diabetic rats? In this study,the above problems will be explored by means of central nuclear localization,gastric motion recording in awake rats,gastric emptying,food intake test,fluorescence retrograde tracing and electrophysiology.Methods: Rat model of diabetes was prepared: After one weeks of adaptive feeding,300 rats were injected intraperitoneally with 65 mg/kg streptozotocin.After 72 h,the fasting blood glucose was measured by tail vein blood.The fasting blood glucose was higher than 7.0 mmol/L and the postprandial blood glucose was higher than 11.1 mmol/ L modeled successfully for diabetic rats,a total of 260 diabetic rats were prepared,and the rats with failed modeling were discarded.The expression of Ghrelin/Obestatin mRNA in hippocampus/ARC of normal and diabetic rats was detected by PCR.Fluorescence gold retrograde tracking combined with fluorescent immunohistochemistry was performed.Hippocampus-ARC Ghrelin neural pathway and ARC-hippocampal Obestatin neural pathway in normal and diabetic rats were investigated;The effects of Ghrelin microinjection into ARC on the discharge activity of gastric traction(GD)neurons and Obestatin microinjection into hippocampus on the discharge activity of GD neurons were observed;Microinjection of drugs in specific nuclues,gastric motility recording in vivo,gastric emptying measurement and food intake test were applied to study the effect of Ghrelin/Obestatin in hippocampal-ARC pathway on gastric motion and feeding in diabetic rats.Results: The results of PCR showed that compared with normal rats,the expression of ghrelin mRNA in hippocampus of diabetic rats increased significantly,while the expression of Obestatin mRNA in ARC decreased significantly.Fluorescence gold retrograde tracing combined with fluorescent immunohistochemistry showed that some Ghrelin positive neurons in hippocampus sent nerve fibers to ARC.Compared with normal rats,the number of FG/Ghrelin double-labeled neurons in hippocampus of diabetic rats is significantly increased.At the same time,some of the Obestatin immunopositive neurons in ARC sent nerve fibers to the hippocampus and the number of FG/Obestatin double-labeled neurons in ARC of diabetic rats was significantly reduced compared with normal rats;Ghrelin excited GD-E neurons and inhibited GD-I neurons in ARC.The effects of Ghrelin on GD neurons in ARC could be completely blocked by Ghrelin receptor blocker Des-Lys-3-GHRP6.Obestatin inhibited the discharge activity of GD neurons in hippocampus.The inhibitory effect of Obestatin on the activity of GD neurons could be partial blocked by corticotropin-releasing factor receptor 1(CRFR1).Ghrelin could promote gastric motility,gastric emptying and feeding in a dose-dependent manner;Des-Lys-3-GHRP6 completely blocked the above effects of ghrelin;Obestatin inhibited gastric motility,gastric emptying and feeding in a dose-dependent manner.NBI27914 partially blocked the effects of Obestatin.The effects of Ghrelin on the discharge activity of GD neurons in ARC and the promoting effect on gastric motility,gastric emptying and feeding in diabetic rats were stronger than that of normal rats while the corresponding effects of Obestatin in hippocampus were weaker than that of normal rats.Conclusion: There are Ghrelin and Obestatin nerve and function pathways between hippocampus and ARC.The two pathways regulated gastric motility,gastric emptying and feeding in both normal and diabetic rats.The enhanced hippocampus-ARC Ghrelin neural pathway and weakened ARC-hippocampus Obestatin neural pathway participated the pathogenesis of hyperphagia and gastric hyperkinesia in early diabetes mellitus and GHS-R and CRFR1 mediated the effects of Ghrelin and Obestatin respectively. |
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