从脑胃ghrelin通路探讨应激对大鼠食欲的影响

doi:10.12300/j.issn.1674-5817.2020.160

摘要/Abstract

摘要： 目的从脑胃饥饿素（ghrelin）通路研究应激影响食欲的分子机制。方法 32只雄性Wistar大鼠随机分为对照组和应激组,每组16只,每笼2只。采用夹尾应激方法每日2次,连续21d刺激大鼠,监测各组大鼠的摄食量及体质量;在实验第7、21天每组各选取8只大鼠,ELISA法检测血清酰基化ghrelin质量浓度;蛋白质印迹法检测大鼠胃及下丘脑中ghrelin蛋白表达,以及下丘脑中腺苷酸活化蛋白激酶α2（Amp-activated protein kinase α2,AMPKα2）表达水平;实时荧光定量PCR检测胃中ghrelin、生长激素促分泌素受体（growth hormone secretagogue receptor,GHSR）、瘦素（leptin）,以及下丘脑中ghrelin、AMPKα2、神经肽Y（neuropeptide Y,NPY）、刺鼠基因相关蛋白（agouti-related protein,AgRP）mRNA水平。结果实验第7天应激组每笼大鼠的总摄食量多于对照组（P<0.05）,第21天两组大鼠的总摄食量未见明显差异（P>0.05）;在第7、21天,两组大鼠的体质量均无明显差异（P>0.05）。应激可以明显上调血清中酰基化ghrelin水平（P<0.01）,胃及下丘脑中ghrelin蛋白（P<0.05）以及下丘脑中AMPKα2蛋白（P<0.01）的表达也均明显上调。应激还能明显上调胃内ghrelin及GHSR mRNA水平（P<0.01）,下调胃内leptin mRNA水平（P<0.01）;同时上调下丘脑内ghrelin、AMPKα2、NPY及AgRP mRNA水平（P<0.01）。结论夹尾应激能导致大鼠中枢及外周ghrelin水平的上调,可能对大鼠食欲暂时性增加有影响。

关键词: 应激, 摄食, 饥饿素, 大鼠

Abstract: Objective To study the molecular mechanism of stress on appetite through hypothalamus and stomach ghrelin pathway. Methods Thirty-two male Wistar rats were randomly divided into a control group and a stress group with 16 rats in each group and 2 rats in each cage. Stress was induced by tail pinch for consecutive 21 days. Food intake and body weight of the rats in each group were recorded. Serum ghrelin concentrations of eight rats in each group were detected by ELISA on day 7 and day 21. Ghrelin protein expression in stomach and hypothalamus as well as Amp-activated protein kinase α2 (AMPKα2) protein expression in hypothalamus of rats were measured by Western blotting. The mRNA levels of ghrelin, growth hormone secretagogue receptor (GHSR) and leptin in stomach as well as ghrelin, AMPKa2, neuropeptide Y (NPY), agouti-related protein (AgRP) in hypothalamus were measured by real-time quantitative PCR. Results On day 7, the total food intake of rats in each cage in the stress group was higher than that in the control group (P < 0.05), and there was no significant difference in total food intake between the two groups on day 21 (P > 0.05). On day 7 and day 21, there was no significant difference in body weight between the two groups (P > 0.05). Stress upregulated the serum levels of acylated ghrelin (P < 0.01), the ghrelin protein expression levels in stomach and hypothalamus (P < 0.05), and the AMPKα2 protein expression level in hypothalamus (P < 0.01). Stress also upregulated the ghrelin and GHSR mRNA expression levels in stomach (P < 0.01), downregulated the leptin mRNA expression level in stomach (P < 0.01), and upregulated the ghrelin, AMPKα2, NPY and AgRP mRNA expression levels in hypothalamus (P < 0.01). Conclusion Tail pinch-induced stress can increase central and peripheral ghrelin levels in rats, and act as a temporary appetite enhancer.

Key words: Stress, Appetite, Ghrelin, Rats

中图分类号:

Q95-33
R-332

赵怡玲, 高鹏飞, 肖茜, 张岚, 潘丹青, 徐月妹. 从脑胃ghrelin通路探讨应激对大鼠食欲的影响[J]. 实验动物与比较医学, 2021, 41(3): 207-214.

ZHAO Yiling, GAO Pengfei, XIAO Qian, ZHANG Lan, PAN Danqing, XU Yuemei. Effects of Stress on Appetite in Rats Through Hypothalamus and Stomach Ghrelin Pathway[J]. Laboratory Animal and Comparative Medicine, 2021, 41(3): 207-214.

图/表 5

参考文献

[1] DALLMAN M F, PECORARO N, AKANA S F, et al.Chronic stress and obesity: a new view of “comfort food”[J]. Proc Natl Acad Sci USA, 2003, 100(20):11696-11701. DOI:10.1073/pnas.1934666100.
[2] OLIVER G, WARDLE J.Perceived effects of stress on food choice[J]. Physiol Behav, 1999, 66(3):511-515. DOI:10.1016/s0031-9384(98)00322-9.
[3] KLATZKIN R R, GAFFNEY S, CYRUS K, et al.Stress-induced eating in women with binge-eating disorder and obesity[J]. Biol Psychol, 2018, 131:96-106. DOI:10.1016/j.biopsycho.2016.11.002.
[4] SINHA R.Chronic stress, drug use, and vulnerability to addiction[J]. Ann N Y Acad Sci, 2008, 1141:105-130. DOI:10.1196/annals.1441.030.
[5] PECORARO N, REYES F, GOMEZ F, et al.Chronic stress promotes palatable feeding, which reduces signs of stress: feedforward and feedback effects of chronic stress[J]. Endocrinology, 2004, 145(8):3754-3762. DOI:10.1210/en.2004-0305.
[6] KOJIMA M, HOSODA H, DATE Y, et al.Ghrelin is a growth-hormone-releasing acylated peptide from stomach[J]. Nature, 1999, 402(6762):656-660. DOI:10.1038/45230.
[7] DATE Y, KOJIMA M, HOSODA H, et al.Ghrelin, a novel growth hormone-releasing acylated peptide, is synthesized in a distinct endocrine cell type in the gastrointestinal tracts of rats and humans[J]. Endocrinology, 2000, 141(11):4255-4261. DOI:10.1210/endo.141.11.7757.
[8] 施新猷. 现代医学实验动物学[M]. 北京: 人民军医出版社, 2000.
[9] LIVAK K J, SCHMITTGEN T D.Analysis of relative gene expression data using real-time quantitative PCR and the 2-^△△Ct method[J]. Methods, 2001, 25(4):402-408. DOI:10.1006/meth.2001.1262.
[10] CHAO A M, JASTREBOFF A M, WHITE M A, et al.Stress, cortisol, and other appetite-related hormones: Prospective prediction of 6-month changes in food cravings and weight[J]. Obesity (Silver Spring), 2017, 25(4):713-720. DOI:10.1002/oby.21790.
[11] CAHILL S, TUPLIN E, HOLAHAN M R.Circannual changes in stress and feeding hormones and their effect on food-seeking behaviors[J]. Front Neurosci, 2013, 7:140. DOI:10.3389/fnins.2013.00140.
[12] STANLEY S, WYNNE K, MCGOWAN B, et al.Hormonal regulation of food intake[J]. Physiol Rev, 2005, 85(4):1131-1158. DOI:10.1152/physrev.00015.2004.
[13] ANDREWS Z B, LIU Z W, WALLLINGFORD N, et al.UCP₂ mediates ghrelin's action on NPY/AgRP neurons by lowering free radicals[J]. Nature, 2008, 454(7206):846-851. DOI:10.1038/nature07181.
[14] BLASIAK A, GUNDLACH A L, HESS G, et al.Interactions of circadian rhythmicity, stress and orexigenic neuropeptide systems: implications for food intake control[J]. Front Neurosci, 2017, 11:127. DOI:10.3389/fnins.2017.00127.
[15] LIM C T, KOLA B, KORBONITS M.AMPK as a mediator of hormonal signalling[J]. J Mol Endocrinol, 2010, 44(2):87-97. DOI:10.1677/JME-09-0063.
[16] AKAMIZU T, KANGAWA K.The physiological significance and potential clinical applications of ghrelin[J]. Eur J Intern Med, 2012, 23(3):197-202. DOI:10.1016/j.ejim.2011.12.001.
[17] YANG C G, WANG W G, YAN J, et al.Gastric motility in ghrelin receptor knockout mice[J]. Mol Med Rep, 2013, 7(1):83-88. DOI:10.3892/mmr.2012.1157.
[18] CHAO A, GRILO C M, WHITE M A, et al.Food cravings, food intake, and weight status in a community-based sample[J]. Eat Behav, 2014, 15(3):478-482. DOI:10.1016/j.eatbeh.2014.06.003.
[19] VOIGT J P, FINK H.Serotonin controlling feeding and satiety[J]. Behav Brain Res, 2015, 277:14-31. DOI:10.1016/j.bbr.2014.08.065.
[20] ZHANG Y, CHUA S.Leptin function and regulation[J]. Compr Physiol, 2017, 8(1):351-369. DOI:10.1002/cphy.c160041.
[21] TAMARGO-GÓMEZ I, MARI O G. AMPK: regulation of metabolic dynamics in the context of autophagy[J]. Int J Mol Sci, 2018, 19(12):E3812. DOI:10.3390/ijms19123812.
[22] LAMBERT G W, STRAZNICKY N E, LAMBERT E A, et al.Sympathetic nervous activation in obesity and the metabolic syndrome: causes, consequences and therapeutic implications[J]. Pharmacol Ther, 2010, 126(2):159-172. DOI:10.1016/j.pharmthera.2010.02.002.
[23] VERHULST P J, DEPOORTERE I.Ghrelin's second life: from appetite timulator to glucose regulator[J]. World J Gastroenterol, 2012, 18(25):3183-3195. DOI:10.3748/wjg.v18.i25.3183.
[24] PORPORATO P E, FILIGHEDDU N, REANO S, et al.Acylated and unacylated ghrelin impair skeletal muscle atrophy in mice[J]. J Clin Invest, 2013, 123(2):611-622. DOI:10.1172/JCI39920.
[25] BARIM A O, AYDIN S, COLAK R, et al.Ghrelin, paraoxonase and arylesterase levels in depressive patients before and after citalopram treatment[J]. Clin Biochem, 2009, 42(10-11):1076-1081. DOI:10.1016/j.clinbiochem.2009.02.020.
[26] OZSOY S, BESIRLI A, ABDULREZZAK U, et al.Serum ghrelin and leptin levels in patients with depression and the effects of treatment[J]. Psychiatry Investig, 2014, 11(2):167-172. DOI:10.4306/pi.2014.11.2.167.
[27] KURT E, GULER O, SERTESER M, et al.The effects of electroconvulsive therapy on ghrelin, leptin and cholesterol levels in patients with mood disorders[J]. Neurosci Lett, 2007, 426(1):49-53. DOI:10.1016/j.neulet.2007.08.018.
[28] ISHITOBI Y, KOHNO K, KANEHISA M, et al.Serum ghrelin levels and the effects of antidepressants in major depressive disorder and panic disorder[J]. Neuropsychobiology, 2012, 66(3):185-192. DOI:10.1159/000339948.
[29] NAKASHIMA K, AKIYOSHI J, HATANO K, et al.Ghrelin gene polymorphism is associated with depression, but not panic disorder[J]. Psychiatr Genet, 2008, 18(5):257. DOI:10.1097/YPG.0b013e328306c979.
[30] FRITZ E M, SINGEWALD N, DE BUNDEL D.The good, the bad and the unknown aspects of ghrelin in stress coping and stress-related psychiatric disorders[J]. Front Synaptic Neurosci, 2020, 12:594484. DOI:10.3389/fnsyn.2020.594484.
[31] JEONG J Y, LEE D H, KANG S S.Effects of chronic restraint stress on body weight, food intake, and hypothalamic gene expressions in mice[J]. Endocrinol Metab (Seoul), 2013, 28(4):288-296. DOI:10.3803/EnM.2013.28.4.288.
[32] JIANG S Z, EIDEN L E.Activation of the HPA axis and depression of feeding behavior induced by restraint stress are separately regulated by PACAPergic neurotransmission in the mouse[J]. Stress, 2016, 19(4):374-382. DOI:10.1080/10253890.2016.1174851.
[33] LI B, XU Y M, PAN D Q, et al.Effect of immobilization stress on the appetite and stomach ghrelin expression in maternal mice[J]. Int J Clin Exp Pathol, 2015, 8(12):15993-15999.
[34] PATTERSON Z R, DUCHARME R, ANISMAN H, et al.Altered metabolic and neurochemical responses to chronic unpredictable stressors in ghrelin receptor-deficient mice[J]. Eur J Neurosci, 2010, 32(4):632-639. DOI:10.1111/j.1460-9568.2010.07310.x.