| Objective:(1)To observe the co-expression of Nesfatin-1-positive neurons and c-fos immuno-positive neurons in paraventricular hypothalamic nucleus(PVN),ventromedial nucleus of the hypothalamus(VMH),arcuate nucleus(Arc)and lateral parabrachial nucleus(LPBN)in Wistar rats under hypoglycemia;(2)To investigate the effect of Nesfatin-1 injection into LPBN on the glucose counter-regulation response(CRR)in normal rats and the mechanism of regulating blood glucose;(3)To explore the effect of LPBN injection of Nesfatin-1 on the CRR induced by different doses of insulin;(4)To observe the effect of LPBN injection of Nesfatin-1 on convulsion and death induced by hypoglycemia in rats.Methods:(1)Frozen section and immunofluorescence double staining were used to observe the co-expression of Nesfatin-1-positive neurons and c-fos immuno-positive neurons in three nuclei(PVN,VMH,Arc)of hypothalamus in rats under hypoglycemia;(2)Nucleus catheterization,nucleus microinjection and intraperitoneal insulin-induced hypoglycemia were used.By injecting of Nesfatin-1 into unilateral LPBN in rats(50pmol,0.5μL),we observed the effect of Nesfatin-1 on CRR.The incidence of convulsion and death in rats after insulin-induced hypoglycemia was calculated,and the duration of convulsion or death in each group was compared.Results:(1)After intraperitoneal injection of insulin for two hours,the expression of Nesfatin-1 in PVN and Arc was significantly increased,while no significant change was observed in VMH.The expression of Nesfatin-1 in LPBN was also significantly increased.Meanwhile,the co-expression of nesfatin-1 and c-fos in PVN,Arc and LPBN was also significantly higher than that in the control group(25.0±6.99 vs.12.7±3.6,P<0.05;16.4±4.1 vs.11.7±2.5,P<0.05;10.4±1.7 vs.6.9±3.2,P<0.05),while there was no significant difference between VMH and control group(22.0±3.3 vs.21.0±4.2,P > 0.05).(2)After intraperitoneal injection of low-dose insulin for 90 and 120 minutes,the blood glucose of the low-dose insulin+Nesfatin-1 group was significantly higher than that of the low-dose insulin+NS control group(t=2.433,3.045;P=0.045,P=0.022).However,after intraperitoneal injection of high-dose insulin for 30 or 60 minutes,the blood glucose of the high-dose insulin+Nesfatin-1 group was significantly higher than that of the high-dose insulin + NS control group(t=2.967,3.546;P=0.011,P=0.003),but Nesfatin-1 did not continuously increase blood glucose at 90 and 120 minutes.In addition,when Nesfatin-1 group received different doses of insulin intraperitoneally,the blood glucose of the high-dose insulin group was significantly higher than that of the low-dose insulin group at 30 minutes(t=4.363,P=0.003),and the blood glucose of the low-dose insulin group was higher than that of the high-dose insulin group at 90 and 120 minutes,but there was no statistical significance.(3)There was significant difference in the incidence of convulsion and death between the low-dose insulin + Nesfatin-1 group and the low-dose insulin + NS group.However,There was no statistically significant difference in the incidence of convulsion and death between the high-dose insulin + Nesfatin-1 group and the high-dose insulin + NS group.Meanwhile,rats in the low-dose insulin + NS group and the high-dose insulin + NS group had more frequent and severe seizures and died earlier than the experimental group.Conclusion:(1)Insulin-induced hypoglycemia in Wistar rats can activate Nesfatin-1 neurons in LPBN.(2)Nesfatin-1 is involved in the CRR,and the injection of Nesfatin-1 into LPBN can obviously enhance CRR and improve the blood glucose of rats.However,under severe hypoglycemia(intraperitoneal injection of 15IU/kg insulin),Nesfatin-1 cannot completely reverse the hypoglycemia caused by high concentration of insulin.(3)Injection of Nesfatin-1 into LPBN can improve the occurrence of convulsions and deaths caused by severe hypoglycemia.This experiment provide a certain experimental reference for further exploration of the prevention and treatment of Nesfatin-1 in LPBN in patients diabetes,especially type I diabetes with hypoglycemia. |
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