大麻素受体1拮抗剂利莫那班对肉仔鸡下丘脑食欲调控因子和腺苷酸激活蛋白激酶基因表达的影响
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摘要
研究旨在探讨大麻素受体1(CB1)拮抗剂利莫那班(SR141716)对肉仔鸡下丘脑食欲调控因子和腺苷酸激活蛋白激酶(AMPK)基因表达的影响。选取1日龄体重相近的健康爱拔益加(AA)肉仔鸡20只,7日龄时随机分为2个组,进行第3脑室埋管,恢复3 d。10日龄08:00试验组以1μg剂量脑室注射SR141716,对照组同时注射等量的二甲基亚砜(DMSO)。统计每只鸡在注射后2 h的采食量,然后采集血液和下丘脑样品,测定血清中丙氨酸转氨酶、天冬氨酸转氨酶活性及葡萄糖、尿酸、总胆固醇、高密度脂蛋白胆固醇、低密度脂蛋白胆固醇含量,利用荧光定量PCR技术检测下丘脑中AMPKα1、AMPKα2以及食欲调控相关基因厌食神经肽(POMC)、促食神经肽Y (NPY)、刺鼠色蛋白相关蛋白(AgRP)、可卡因和苯丙胺调节转录物(CART)的基因表达量。结果表明:1)与对照组相比,肉仔鸡脑室注射SR141716后,2 h内的采食量显著降低(P<0.05)。2)与对照组相比,肉仔鸡脑室注射SR141716显著提高了血清中低密度脂蛋白胆固醇含量(P<0.05),对其他血清生化指标无显著影响(P>0.05)。3)与对照组相比,肉仔鸡脑室注射SR141716显著降低了下丘脑AMPKα1、AMPKα2、POMC和AgRP的基因表达量(P<0.05),显著提高了下丘脑CART的基因表达量(P<0.05),对下丘脑NPY的基因表达量无显著影响(P>0.05)。由此可见,脑室注射CB1受体拮抗剂SR141716可降低肉仔鸡食欲,并影响中枢AMPK及其下游食欲调控因子POMC的基因表达;中枢通过抑制促食神经肽AgRP和提高抑食神经肽CART的基因表达,参与肉仔鸡食欲调控。
This study aimed to investigate the effects of cannabinoid receptor 1(CB1) antagonist rimonabant(SR141716) on gene expression of hypothalamic appetite regulation factor and adenosine monophosphate-activated protein kinase(AMPK) of broilers.Twenty healthy one-day-old Arbor Acres(AA) broilers with similar weight were randomly allocated into 2 groups at 7 days of age,and a guide cannula were implanted stereotaxically into the third ventricle,chicks were allowed 3 days of recovery.At 08:00 at 10 days of age,birds in the experimental group were ventricle injected 1 μg SR141716,while other birds in the control group injected the same amount of dimethyl sulfoxide(DMSO) at the same time.The feed intake of each chick in the 2 hours after injection was counted,then the blood and hypothalamic sample was taken,the activities of alanine aminotransferase,aspartate aminotransferase and contents of glucose,uric acid,total cholesterol,high density lipoprotein cholesterol and low density lipoprotein cholesterol in serum were determined.The gene expression of AMPKα1,AMPKα2,pro-opiomelanocortin-α(POMC),neuropeptide Y(NPY),agouti-related protein(AgRP) and cocaine-and amphetamine-regulated transcript(CART) in hypothalamus were detected by real time PCR method.The results showed as follows:1) compared with the control group,the feed intake of broilers was significantly decreased in the 2 hours after ventricle injected SR141716(P<0.05).2) Compared with the control group,the serum low density lipoprotein cholesterol content of broilers was significantly increased after ventricle injected SR141716(P<0.05),but there was no significant effect on other serum biochemical parameters(P > 0.05).3) Compared with the control group,the gene expression of AMPKα1,AMPKα2,POMC and AgRP in hypothalamus of broilers were significantly decreased after ventricle injected SR141716(P<0.05),while the gene expression of CART in hypothalamus was significantly increased(P<0.05),but there was no significant effect on the gene expression of NPY in hypothalamus(P>0.05).In conclusion,ventricle injected CB1 antagonist SR141716 can reduce the feed intake of broilers and affect the gene expression of AMPK and its downstream appetite regulator POMC in central nervous.The central nervous is involved in the appetite regulation of broilers by inhibiting the stimulate neuropeptide AgRP and anorexigenic neuropeptides CART.
This study aimed to investigate the effects of cannabinoid receptor 1(CB1) antagonist rimonabant(SR141716) on gene expression of hypothalamic appetite regulation factor and adenosine monophosphate-activated protein kinase(AMPK) of broilers.Twenty healthy one-day-old Arbor Acres(AA) broilers with similar weight were randomly allocated into 2 groups at 7 days of age,and a guide cannula were implanted stereotaxically into the third ventricle,chicks were allowed 3 days of recovery.At 08:00 at 10 days of age,birds in the experimental group were ventricle injected 1 μg SR141716,while other birds in the control group injected the same amount of dimethyl sulfoxide(DMSO) at the same time.The feed intake of each chick in the 2 hours after injection was counted,then the blood and hypothalamic sample was taken,the activities of alanine aminotransferase,aspartate aminotransferase and contents of glucose,uric acid,total cholesterol,high density lipoprotein cholesterol and low density lipoprotein cholesterol in serum were determined.The gene expression of AMPKα1,AMPKα2,pro-opiomelanocortin-α(POMC),neuropeptide Y(NPY),agouti-related protein(AgRP) and cocaine-and amphetamine-regulated transcript(CART) in hypothalamus were detected by real time PCR method.The results showed as follows:1) compared with the control group,the feed intake of broilers was significantly decreased in the 2 hours after ventricle injected SR141716(P<0.05).2) Compared with the control group,the serum low density lipoprotein cholesterol content of broilers was significantly increased after ventricle injected SR141716(P<0.05),but there was no significant effect on other serum biochemical parameters(P > 0.05).3) Compared with the control group,the gene expression of AMPKα1,AMPKα2,POMC and AgRP in hypothalamus of broilers were significantly decreased after ventricle injected SR141716(P<0.05),while the gene expression of CART in hypothalamus was significantly increased(P<0.05),but there was no significant effect on the gene expression of NPY in hypothalamus(P>0.05).In conclusion,ventricle injected CB1 antagonist SR141716 can reduce the feed intake of broilers and affect the gene expression of AMPK and its downstream appetite regulator POMC in central nervous.The central nervous is involved in the appetite regulation of broilers by inhibiting the stimulate neuropeptide AgRP and anorexigenic neuropeptides CART.
引文
[1]陈璇,杨明,郭鸿彦.大麻植物中大麻素成分研究进展[J].植物学报,2011,46(2):197-205.
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[3]PERTWEE R G.Pharmacology of cannabinoid receptor ligands[J].Current Medicinal Chemistry,1999,6(8):635-664.
[4]TANDA G,GOLDBERG S R.Cannabinoids:reward,dependence,and underlying neurochemical mechanisms—a reviewof recent preclinical data[J].Psychopharmacology,2003,169(2):115-134.
[5]MOLDRICH G,WENGER T.Localization of the CB1cannabinoid receptor in the rat brain.An immunohistochemical study[J].Peptides,2000,21(11):1735-1742.
[6]DEN BOON F S,CHAMEAU P,SCHAAFSMAZHAO Q,et al.Excitability of prefrontal cortical pyramidal neurons is modulated by activation of intracellular type-2 cannabinoid receptors[J].Proceedings of the National Academy of Sciences of the United States of America,2012,109(9):3534-3539.
[7]KYROU I,VALSAMAKIS G,TSIGOS C.The endocannabinoid system as a target for the treatment of visceral obesity and metabolic syndrome[J].Annals of the New York Academy of Sciences,2006,1083(1):270-305.
[8]ARNONE M,MARUANI J,CHAPERON F,et al.Selective inhibition of sucrose and ethanol intake by SR141716,an antagonist of central cannabinoid(CB1)receptors[J].Psychopharmacology,1997,132(1):104-106.
[9]JAMSHIDI N,TAYLOR D A.Anandamide administration into the ventromedial hypothalamus stimulates appetite in rats[J].British Journal of Pharmacology,2001,134(6):1151-1154.
[10]KIRKHAMT C,WILLIAMS C M,FEZZA F,et al.Endocannabinoid levels in rat limbic forebrain and hypothalamus in relation to fasting,feeding and satiation:stimulation of eating by 2-arachidonoyl glycerol[J].British Journal of Pharmacology,2002,136(4):550-557.
[11]KOLA B,HUBINA E,TUCCI S A,et al.Cannabinoids and ghrelin have both central and peripheral metabolic and cardiac effects via AMP-activated protein kinase[J].Journal of Biological Chemistry,2005,280(26):25196-25201.
[12]KIMMS,PAK Y K,JIANG P G,et al.Role of hypothalamic Foxo1 in the regulation of food intake and energy homeostasis[J].Nature Neuroscience,2006,9(7):901-906.
[13]JAILLARD T,ROGER M,GALINIER A,et al.Hypothalamic reactive oxygen species are required for insulin-induced food intake inhibition:an NADPH oxidase-dependent mechanism[J].Diabetes,2009,58(7):1544-1549.
[14]LIVAK K J,SCHMITTGEN T D.Analysis of relative gene expression data using real-time quantitative PCR and the 2-ΔΔCTmethod[J].Methods,2001,25(4):402-408.
[15]PARKER K E,JOHNS H W,FLOROS T G,et al.Central amygdala opioid transmission is necessary for increased high-fat intake following 24-h food deprivation,but not following intra-accumbens opioid administration[J].Behavioural Brain Research,2014,260:131-138.
[16]KIRKHAMT C.Endocannabinoid and non-homeostatic controls of appetite[J].Official Journal of the International Chair on Cardiometabolic Risk,2008,1(3):22-26.
[17]STINCIC T L,HYSON R L.Localization of CB1 cannabinoid receptor mRNA in the brain of the chick(Gallus domesticus)[J].Brain Research,2008,1245:61-73.
[18]ALIZADEH A,ZENDEHDEL M,BABAPOUR V,et al.Role of cannabinoidergic system on food intake in neonatal layer-type chicken[J].Veterinary Research Communications,2015,39(2):151-157.
[19]NOVOSELETSKY N,NUSSINOVITCH A,FRIEDMAN-EINAT M.Attenuation of food intake in chicks by an inverse agonist of cannabinoid receptor 1 administered by either injection or ingestion in hydrocolloid carriers[J].General and Comparative Endocrinology,2011,170(3):522-527.
[20]MAI P,YANG L,TIAN L,et al.Endocannabinoid system contributes to liver injury and inflammation by activation of bone marrow-derived monocytes/macrophages in a CB1-dependent manner[J].Journal of Immunology,2015,195(7):3390-3401.
[21]MUKHOPADHYAY B,SCHUEBEL K,MUKHOPADHYAY P,et al.Cannabinoid receptor 1 promotes hepatocellular carcinoma initiation and progression through multiple mechanisms[J].Hepatology,2015,61(5):1615-1626.
[22]SHI D M,XI Z,YU X F,et al.Inhibiting CB1 receptors improves lipogenesis in an in vitro non-alcoholic fatty liver disease model[J].Lipids in Health and Disease,2014,13:173.
[23]CHANDA D,KIMY H,LI T G,et al.Hepatic cannabinoid receptor type 1 mediates alcohol-induced regulation of bile acid enzyme genes expression via CREBH[J].PLoS One,2013,8(7):e68845.
[24]ZHANG Y F,YUAN Z Q,SONG D G,et al.Effects of cannabinoid receptor 1(brain)on lipid accumulation by transcriptional control of CPT1A and CPT1B[J].Animal Genetics,2014,45(1):38-47.
[25]KOLA B,WITTMAN G,BODNR I,et al.The CB1receptor mediates the peripheral effects of ghrelin on AMPK activity but not on grow th hormone release[J].The FASEB Journal,2013,27(12):5112-5221.
[26]KIMJ,YANG G,KIMY,et al.AMPK activators:mechanisms of action and physiological activities[J].Experimental&Molecular Medicine,2016,48(4):e224.
[27]OH T S,CHO H,CHO J H,et al.Hypothalamic AMPK-induced autophagy increases food intake by regulating NPY and POMC expression[J].Autophagy,2016,12(11):2009-2025.
[28]TACHIBANA T,TAKAGI T,TOMONAGA S,et al.Central administration of cocaine-and amphetamineregulated transcript inhibits food intake in chicks[J].Neuroscience Letters,2003,337(3):131-134.
[29]VERTY A N,BOON W M,MALLET P E,et al.Involvement of hypothalamic peptides in the anorectic action of the CB1 receptor antagonist rimonabant(SR141716)[J].European Journal of Neuroscience,2009,29(11):2207-2216.
[30]COTA D,MARSICANO G,TSCHP M,et al.The endogenous cannabinoid system affects energy balance via central orexigenic drive and peripheral lipogenesis[J].Journal of Clinical Investigation,2003,112(3):423-431.
[31]OSEI-HYIAMAN D,DEPETRILLO M,HARVEYWHITE J,et al.Cocaine-and amphetamine-related transcript is involved in the orexigenic effect of endogenous anandamide[J].Neuroendocrinology,2005,81(4):273-282.
[32]KRASHES MJ,KODA S,YE C P,et al.Rapid,reversible activation of AgRP neurons drives feeding behavior in mice[J].Journal of Clinical Investigation,2011,121(4):1424-1428.
[33]STAROWICZ K,NIGAMS,DI MARZO V.Biochemistry and pharmacology of endovanilloids[J].Pharmacology&Therapeutics,2007,114(1):13-33.
[34]WU J G,ZHU C J,YANG L S,et al.N-oleoylglycineinduced hyperphagia is associated with the activation of agouti-related protein(AgRP)neuron by cannabinoid receptor type 1(CB1R)[J].Journal of Agricultural and Food Chemistry,2017,65(5):1051-1057.
[2]GAONI Y,MECHOULAMR.Isolation,structure,and partial synthesis of an active constituent of hashish[J].Journal of Analytical and Applied Chemistry,1964,86(8):1646-1647.
[3]PERTWEE R G.Pharmacology of cannabinoid receptor ligands[J].Current Medicinal Chemistry,1999,6(8):635-664.
[4]TANDA G,GOLDBERG S R.Cannabinoids:reward,dependence,and underlying neurochemical mechanisms—a reviewof recent preclinical data[J].Psychopharmacology,2003,169(2):115-134.
[5]MOLDRICH G,WENGER T.Localization of the CB1cannabinoid receptor in the rat brain.An immunohistochemical study[J].Peptides,2000,21(11):1735-1742.
[6]DEN BOON F S,CHAMEAU P,SCHAAFSMAZHAO Q,et al.Excitability of prefrontal cortical pyramidal neurons is modulated by activation of intracellular type-2 cannabinoid receptors[J].Proceedings of the National Academy of Sciences of the United States of America,2012,109(9):3534-3539.
[7]KYROU I,VALSAMAKIS G,TSIGOS C.The endocannabinoid system as a target for the treatment of visceral obesity and metabolic syndrome[J].Annals of the New York Academy of Sciences,2006,1083(1):270-305.
[8]ARNONE M,MARUANI J,CHAPERON F,et al.Selective inhibition of sucrose and ethanol intake by SR141716,an antagonist of central cannabinoid(CB1)receptors[J].Psychopharmacology,1997,132(1):104-106.
[9]JAMSHIDI N,TAYLOR D A.Anandamide administration into the ventromedial hypothalamus stimulates appetite in rats[J].British Journal of Pharmacology,2001,134(6):1151-1154.
[10]KIRKHAMT C,WILLIAMS C M,FEZZA F,et al.Endocannabinoid levels in rat limbic forebrain and hypothalamus in relation to fasting,feeding and satiation:stimulation of eating by 2-arachidonoyl glycerol[J].British Journal of Pharmacology,2002,136(4):550-557.
[11]KOLA B,HUBINA E,TUCCI S A,et al.Cannabinoids and ghrelin have both central and peripheral metabolic and cardiac effects via AMP-activated protein kinase[J].Journal of Biological Chemistry,2005,280(26):25196-25201.
[12]KIMMS,PAK Y K,JIANG P G,et al.Role of hypothalamic Foxo1 in the regulation of food intake and energy homeostasis[J].Nature Neuroscience,2006,9(7):901-906.
[13]JAILLARD T,ROGER M,GALINIER A,et al.Hypothalamic reactive oxygen species are required for insulin-induced food intake inhibition:an NADPH oxidase-dependent mechanism[J].Diabetes,2009,58(7):1544-1549.
[14]LIVAK K J,SCHMITTGEN T D.Analysis of relative gene expression data using real-time quantitative PCR and the 2-ΔΔCTmethod[J].Methods,2001,25(4):402-408.
[15]PARKER K E,JOHNS H W,FLOROS T G,et al.Central amygdala opioid transmission is necessary for increased high-fat intake following 24-h food deprivation,but not following intra-accumbens opioid administration[J].Behavioural Brain Research,2014,260:131-138.
[16]KIRKHAMT C.Endocannabinoid and non-homeostatic controls of appetite[J].Official Journal of the International Chair on Cardiometabolic Risk,2008,1(3):22-26.
[17]STINCIC T L,HYSON R L.Localization of CB1 cannabinoid receptor mRNA in the brain of the chick(Gallus domesticus)[J].Brain Research,2008,1245:61-73.
[18]ALIZADEH A,ZENDEHDEL M,BABAPOUR V,et al.Role of cannabinoidergic system on food intake in neonatal layer-type chicken[J].Veterinary Research Communications,2015,39(2):151-157.
[19]NOVOSELETSKY N,NUSSINOVITCH A,FRIEDMAN-EINAT M.Attenuation of food intake in chicks by an inverse agonist of cannabinoid receptor 1 administered by either injection or ingestion in hydrocolloid carriers[J].General and Comparative Endocrinology,2011,170(3):522-527.
[20]MAI P,YANG L,TIAN L,et al.Endocannabinoid system contributes to liver injury and inflammation by activation of bone marrow-derived monocytes/macrophages in a CB1-dependent manner[J].Journal of Immunology,2015,195(7):3390-3401.
[21]MUKHOPADHYAY B,SCHUEBEL K,MUKHOPADHYAY P,et al.Cannabinoid receptor 1 promotes hepatocellular carcinoma initiation and progression through multiple mechanisms[J].Hepatology,2015,61(5):1615-1626.
[22]SHI D M,XI Z,YU X F,et al.Inhibiting CB1 receptors improves lipogenesis in an in vitro non-alcoholic fatty liver disease model[J].Lipids in Health and Disease,2014,13:173.
[23]CHANDA D,KIMY H,LI T G,et al.Hepatic cannabinoid receptor type 1 mediates alcohol-induced regulation of bile acid enzyme genes expression via CREBH[J].PLoS One,2013,8(7):e68845.
[24]ZHANG Y F,YUAN Z Q,SONG D G,et al.Effects of cannabinoid receptor 1(brain)on lipid accumulation by transcriptional control of CPT1A and CPT1B[J].Animal Genetics,2014,45(1):38-47.
[25]KOLA B,WITTMAN G,BODNR I,et al.The CB1receptor mediates the peripheral effects of ghrelin on AMPK activity but not on grow th hormone release[J].The FASEB Journal,2013,27(12):5112-5221.
[26]KIMJ,YANG G,KIMY,et al.AMPK activators:mechanisms of action and physiological activities[J].Experimental&Molecular Medicine,2016,48(4):e224.
[27]OH T S,CHO H,CHO J H,et al.Hypothalamic AMPK-induced autophagy increases food intake by regulating NPY and POMC expression[J].Autophagy,2016,12(11):2009-2025.
[28]TACHIBANA T,TAKAGI T,TOMONAGA S,et al.Central administration of cocaine-and amphetamineregulated transcript inhibits food intake in chicks[J].Neuroscience Letters,2003,337(3):131-134.
[29]VERTY A N,BOON W M,MALLET P E,et al.Involvement of hypothalamic peptides in the anorectic action of the CB1 receptor antagonist rimonabant(SR141716)[J].European Journal of Neuroscience,2009,29(11):2207-2216.
[30]COTA D,MARSICANO G,TSCHP M,et al.The endogenous cannabinoid system affects energy balance via central orexigenic drive and peripheral lipogenesis[J].Journal of Clinical Investigation,2003,112(3):423-431.
[31]OSEI-HYIAMAN D,DEPETRILLO M,HARVEYWHITE J,et al.Cocaine-and amphetamine-related transcript is involved in the orexigenic effect of endogenous anandamide[J].Neuroendocrinology,2005,81(4):273-282.
[32]KRASHES MJ,KODA S,YE C P,et al.Rapid,reversible activation of AgRP neurons drives feeding behavior in mice[J].Journal of Clinical Investigation,2011,121(4):1424-1428.
[33]STAROWICZ K,NIGAMS,DI MARZO V.Biochemistry and pharmacology of endovanilloids[J].Pharmacology&Therapeutics,2007,114(1):13-33.
[34]WU J G,ZHU C J,YANG L S,et al.N-oleoylglycineinduced hyperphagia is associated with the activation of agouti-related protein(AgRP)neuron by cannabinoid receptor type 1(CB1R)[J].Journal of Agricultural and Food Chemistry,2017,65(5):1051-1057.
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