海军军医大学学报  2025, Vol. 46 Issue (1): 72-78   PDF    
慢性应激对小鼠下丘脑食欲调节因子的影响
尹立峰1,2, 刘庆2, 朱健2, 王晨旭2, 郑天宇2, 朱永恒1, 莫烽锋2     
1. 上海海洋大学食品学院, 上海 201306;
2. 海军军医大学(第二军医大学)海军医学系海军营养与食品卫生学教研室, 上海 200433
摘要: 目的 观察小鼠在慢性应激时及应激停止后下丘脑食欲调节因子的变化,并初步探索应激依赖性的食欲变化机制。方法 将32只雄性C57BL/6小鼠随机分为对照(Ctrl)组及慢性不可预知温和应激(CUMS)组,每组16只。CUMS组小鼠予以CUMS(持续11周)建立应激模型,Ctrl组正常饲养。记录小鼠摄食量和体重。通过悬尾实验和强迫游泳实验验证CUMS模型是否构建成功。第12周时从两组各随机取8只小鼠收集全脑、下丘脑组织待测;两组各剩余的8只小鼠重新编组为应激停止(C-CUMS)组和应激停止对照(C-Ctrl)组,继续观察3周后,取全脑、下丘脑组织待测。通过qPCR和免疫组织化学染色方法检测小鼠下丘脑中食欲调节因子食欲素1型受体(OX1R)、瘦素受体(LEPR)和刺鼠相关蛋白(AgRP)的表达情况。结果 应激第2~11周,CUMS组小鼠累积摄食量高于Ctrl组(均P<0.05),11周内两组小鼠体重差异无统计学意义(均P>0.05)。11周后CUMS组小鼠悬尾不动时间和水中漂浮时间均长于Ctrl组小鼠(均P<0.01),提示造模成功。造模成功后,CUMS组小鼠下丘脑组织中促食欲因子AgRP mRNA、OX1R mRNA表达量高于Ctrl组(均P<0.01),抑食欲因子LEPR mRNA表达量低于Ctrl组(P<0.01);CUMS组小鼠下丘脑弓状核中AgRP蛋白表达水平高于Ctrl组(P<0.05),LEPR蛋白表达水平低于Ctrl组(P<0.01)。应激停止3周后,C-CUMS组小鼠累积摄食量少于C-Ctrl组(P<0.05),体重低于C-Ctrl组(P<0.05)。C-CUMS组小鼠LEPR mRNA表达量高于C-Ctrl组(P<0.01),AgRP mRNA、OX1R mRNA表达量与C-Ctrl组相比差异无统计学意义(均P>0.05);C-CUMS组小鼠AgRP蛋白表达水平与C-Ctrl组比较差异无统计学意义(P>0.05),LEPR蛋白表达水平高于C-Ctrl组(P<0.01)。结论 CUMS导致小鼠食欲增加,其机制可能与LEPR和AgRP的功能调节有关;应激停止后其食欲下降,机制可能涉及LEPR的功能调节。
关键词: 慢性不可预知温和应激    食欲调节因子    下丘脑    食欲素1型受体    瘦素受体    刺鼠相关蛋白    
Effects of chronic stress on hypothalamic appetite-regulating factors in mice
YIN Lifeng1,2, LIU Qing2, ZHU Jian2, WANG Chenxu2, ZHENG Tianyu2, ZHU Yongheng1, MO Fengfeng2     
1. College of Food Science and Technology, Shanghai Ocean University, Shanghai 201306, China;
2. Department of Naval Nutrition and Food Hygiene, Faculty of Naval Medicine, Naval Medical University (Second Military Medical University), Shanghai 200433, China
Abstract: Objective To explore the effects of chronic stress and stress cessation on hypothalamic appetite regulators in mice, and to explore the stress-dependent mechanism of appetite change. Methods A total of 32 male C57BL/6 mice were randomly divided into control (Ctrl) group (n=16) and chronic unpredictable mild stress (CUMS) group (n=16). The mice in the CUMS group were given CUMS to establish the stress model, and those in the Ctrl group were fed normally. The food intake and weight of mice were recorded. The CUMS model was verified through tail suspension experiments and forced swimming experiments. Eight mice in the Ctrl group and 8 mice in the CUMS group were randomly sacrificed at the 12th week. The Ctrl group was re-grouped into the cessation-control (C-Ctrl) group (n=8), the CUMS group was re-grouped into the cessation-stress (C-CUMS) group (n=8), and the mice were sacrificed at the 15th week. The mRNA and protein levels of appetite-regulating factors, including orexin 1 receptor (OX1R), leptin receptor (LEPR) and agouti-related protein (AgRP) in the hypothalamus, were detected by quantitative polymerase chain reaction and immunochemistry. Results From week 2 to week 11 of stress, the food intake of the mice in the CUMS group was significantly higher than that in the Ctrl group (all P < 0.05), while there was no significant difference in body weight between the 2 groups within 11 weeks (all P > 0.05). Compared with the Ctrl group, the immobility durations of forced swimming and tail suspension of the CUMS group were markedly longer after 11 weeks (both P < 0.01), indicating successful modeling. AgRP and OX1R mRNA expression in the hypothalamus of the CUMS group was significantly increased (both P < 0.01), while LEPR mRNA expression was significantly decreased (P < 0.01); AgRP protein in the hypothalamic arcuate nucleus of the CUMS group was significantly higher than that of the Ctrl group (P < 0.05), and LEPR protein was markedly lower than that of the Ctrl group (P < 0.01). However, after 3 weeks of stress cessation, the C-CUMS group had less food intake and lower body weight than the C-Ctrl group (both P < 0.05). The LEPR mRNA of the C-CUMS group was significantly increased (P < 0.01), while AgRP and OX1R mRNA were not significantly different (both P > 0.05). There was no significant difference in AgRP protein levels between the C-CUMS group and the C-Ctrl group (P > 0.05), while LEPR protein level of the C-CUMS group was significantly higher than that of the C-Ctrl group (P < 0.01). Conclusion CUMS can lead to increased appetite in mice, which may involve the functional regulation of LEPR and AgRP. After the stress cessation, the appetite decreases, which may involve the functional regulation of LEPR.
Key words: chronic unpredictable mild stress    appetite regulating factors    hypothalamus    orexin 1 receptor    leptin receptor    agouti-related protein    

应激又称为压力,是当身体应对超出自身所能承受的刺激时所表现出的一种系统性的、非特异性的反应[1]。伴随着社会生活和工作节奏的加快,当代人各个层面的压力正在不断增加,越来越多的人出现各种躯体和精神方面的异常症状。关于应激对食欲的影响已有大量研究,然而应激源停止(或脱离应激环境)后食欲的变化及机制的相关研究甚少。下丘脑是机体神经中枢影响摄食行为的核心,也是应激影响的主要脑区,其中紧邻第三脑室的弓状核是下丘脑中影响食欲的一个重要核团,包含多个与食欲调控有关的影响因子[2]。在众多动物应激模型中,慢性不可预知温和应激(chronic unpredictable mild stress,CUMS)与人类心身疾病具有高度相似性,因此本研究采用此模型,通过观察C57BL/6小鼠体重和摄食量变化,检测下丘脑食欲调节因子食欲素1型受体(orexin 1 receptor,OX1R)、瘦素受体(leptin receptor,LEPR)和刺鼠相关蛋白(agouti-related protein,AgRP)的表达,探讨应激依赖性的食欲变化机制。

1 材料和方法 1.1 动物模型的构建与分组

C57BL/6小鼠,雄性,6~8周龄,体重18~22 g,由上海杰思捷实验动物有限公司[动物生产许可证号:SCXK(沪)2018-0004]提供。将小鼠随机分为对照(Ctrl)组(n=16)和应激(CUMS)组(n=16)。小鼠在12 h光照/12 h黑暗循环、温度20~25 ℃、相对湿度65%~75%、自由摄食和饮水的条件下饲养1周后进行实验。

CUMS组小鼠每周随机进行7种应激(味道刺激、昼夜颠倒、束缚、潮湿垫料、空笼、频闪、强光照射36 h[3]),连续11周,建立CUMS模型。对照组正常饲养。记录小鼠摄食量和体重。通过悬尾实验和强迫游泳实验检测小鼠抑郁样行为,验证模型是否建立成功。模型建立成功后(第12周),从两组各随机取8只小鼠收集全脑、下丘脑组织;两组各剩余的8只小鼠重新编组为应激停止(C-CUMS)组和应激停止对照(C-Ctrl)组,继续观察3周后,于第15周取全脑、下丘脑组织。全脑组织立即放入多聚甲醛溶液固定,随后进行石蜡包埋、切片;下丘脑组织立即放于-80 ℃冰箱保存待测。

1.2 qPCR检测小鼠下丘脑食欲调节因子mRNA的表达

取小鼠下丘脑组织放于1.5 mL离心管中,加入1 mL RNAiso Plus裂解液(货号9109,日本TaKaRa公司),使用组织研磨机研磨;匀浆后置于冰上,加入200 μL氯仿(货号67-66-3,昆山金城试剂有限公司),用漩涡振荡器剧烈震荡15 s后,4 ℃、16 000×g离心15 min;取上清液,加入500 μL异丙醇(货号67-63-0,上海麦克林生化科技股份有限公司),混合均匀,4 ℃静置10 min后,4 ℃、16 000×g离心15 min;弃上清液,加入1 mL 75%乙醇(山东利尔康医疗科技股份有限公司)漂洗后,4 ℃ 6 300×g离心5 min;弃上清液,室温干燥25 min至沉淀透明,加入DEPC水[货号B501005-0500,生工生物工程(上海)股份有限公司]溶解RNA。将获得的RNA按照反转录试剂盒(货号RR036A,日本TaKaRa公司)的步骤操作,获得cDNA。以β-肌动蛋白(β-actin)为内参,采用qPCR法检测小鼠下丘脑食欲调节因子OX1RLEPRAgRP mRNA的表达。β-actin正向引物5'-GTGCTATGTTGCTCTAGACTTCG-3',反向引物5'-ATGCCACAGGATTCCATACC-3';OX1R正向引物5'-CAGGCTTTGTGCAAGGTCAT-3',反向引物5'-CTTGAACAACAGTGGGTGGC-3';LEPR正向引物5'-ATGACGCAGGGCTGTATGTC-3',反向引物5'-TGGACTGTTGGGAAGTTGGTAG-3';AgRP正向引物5'-GGAAGCAGTCACGTGTGGACCC-3',反向引物5'-AGTGGAGCCACGCCCATCTGG-3'。按照试剂盒(货号RR420A,日本TaKaRa公司)说明书配制扩增体系(20 μL):TB Green Premix Ex Taq 10 μL,正、反向引物(10 μmol/L)各0.4 μL,ROX Reference Dye Ⅱ(50×)0.4 μL,DNA模板2 μL,灭菌水6.8 μL。将扩增体系分别加入八联管,使用QuantStudio 1荧光定量PCR仪(美国赛默飞公司)进行检测。采用2-ΔΔCt法计算目的基因的相对表达量。

1.3 免疫组织化学染色检测小鼠下丘脑AgRP和LEPR蛋白的表达

取小鼠全脑组织切片,经二甲苯、无水乙醇、90%乙醇、80%乙醇、70%乙醇脱蜡至水后,用PBS缓慢振荡洗涤5次、每次3 min;室温下于通透液中浸泡15 min,用PBS缓慢振荡洗涤6次、每次5 min;通过煮沸法进行抗原修复,将切片置于沸水浴中20 min,冷却至室温后用PBS缓慢振荡洗涤6次、每次5 min;加入内源性过氧化物酶阻断剂(3%过氧化氢溶液)孵育15 min,用PBS缓慢振荡洗涤5次、每次3 min;采用山羊血清在室温下孵育10 min;加入稀释好的一抗(LEPR抗体:货号20966-1-AP,武汉三鹰生物技术有限公司;AgRP抗体:货号ab254558,英国Abcam公司),4 ℃孵育过夜;第2天在室温下复温30 min,用PBS缓慢振荡洗涤5次、每次3 min,洗去未结合的一抗;加入对应一抗种属的二抗试剂(货号PV-9001,北京中杉金桥生物技术有限公司),室温孵育25 min,用PBS缓慢振荡洗涤5次、每次3 min;加入HRP标记的链霉卵白素试剂,室温孵育15 min,用PBS缓慢振荡洗涤5次、每次3 min;避光加入按比例稀释的DAB显色剂,室温孵育,在显微镜下观察5~15 min,发现变色后放入自来水中终止显色,用PBS缓慢振荡洗涤5次、每次3 min;用苏木精染核30 s;加入5%冰乙酸分化30 s;流水冲洗1 h;70%、80%、90%、100%乙醇分别冲洗30 s;二甲苯浸泡1 min;中性树脂封片。

1.4 统计学处理

应用GraphPad Prism 8.3.0软件进行数据分析和绘图。呈正态分布的计量资料以x±s表示,任意两组间的比较采用独立样本t检验。检验水准(α)为0.05。

2 结果 2.1 CUMS对小鼠摄食量及体重的影响

CUMS组小鼠第2~11周累积摄食量均高于Ctrl组(均P<0.05,图 1A),11周内两组小鼠体重差异无统计学意义(均P>0.05,图 1B)。有趣的是,停止应激3周后,第15周时C-CUMS组小鼠累积摄食量少于C-Ctrl组(P<0.05,图 1C)。第11周时CUMS组和Ctrl组小鼠体重差异无统计学意义(n=16,P>0.05,图 1B),但C-CUMS组第11~15周体重均低于C-Ctrl组(n=8,均P<0.05,图 1D)。以上结果表明CUMS对小鼠的食欲有重要影响。

图 1 各组小鼠累积摄食量和体重的变化 Fig 1 Changes of cumulative food intake and body weight of mice in each group A, C: Changes of cumulative food intake; B, D: Changes of body weight. *P < 0.05, **P < 0.01 vs Ctrl group at the same week; P < 0.05, △△P < 0.01 vs C-Ctrl group at the same week. n=16 in Ctrl and CUMS group, n=8 in C-Ctrl and C-CUMS group, x±s. Ctrl: Control; CUMS: Chronic unpredictable mild stress; C-Ctrl: Cessation-control; C-CUMS: Cessation-CUMS.

2.2 CUMS模型验证

采用悬尾实验和强迫游泳实验评价小鼠抑郁样行为,CUMS组小鼠的悬尾不动时间和水中漂浮时间均长于Ctrl组小鼠[(42.82±8.07)s vs(26.48±6.44)s,(223.01±14.27)s vs(165.12±20.39)s,均P<0.01]。以上实验结果表明小鼠出现了明显的抑郁样行为,CUMS模型构建成功。

2.3 CUMS对小鼠下丘脑组织中食欲调节因子表达的影响

qPCR检测结果(表 1)显示,CUMS组小鼠下丘脑组织中OX1R mRNA和AgRP mRNA表达量较Ctrl组升高(均P<0.01),LEPR mRNA表达量较Ctrl组下降(P<0.01)。停止应激3周后,C-CUMS组与C-Ctrl组小鼠下丘脑组织中OX1R mRNA和AgRP mRNA表达量差异均无统计学意义(均P>0.05),但C-CUMS组LEPR mRNA表达量较C-Ctrl组升高(P<0.01)。以上实验结果表明,CUMS导致下丘脑组织中促食欲因子OX1R mRNA和AgRP mRNA表达量升高、抑食欲因子LEPR mRNA表达量降低,应激停止后LEPR mRNA表达水平提高,提示食欲调节因子在CUMS导致的食欲变化中起着重要作用。

表 1 qPCR检测各组小鼠下丘脑组织中食欲调节因子的mRNA相对表达量 Tab 1 mRNA expression of appetite-regulating factors in hypothalamus of mice in each group detected by qPCR   

免疫组织化学染色结果显示,CUMS组小鼠下丘脑弓状核中AgRP蛋白表达水平较Ctrl组升高(P<0.05),而C-CUMS组与C-Ctrl组比较小鼠下丘脑弓状核中AgRP蛋白表达水平差异无统计学意义(P>0.05),见图 2。以上结果表明CUMS激活了小鼠下丘脑弓状核中的AgRP神经元。

图 2 各组小鼠下丘脑弓状核AgRP免疫组织化学染色结果 Fig 2 Immunohistochemical staining of AgRP in hypothalamic arcuate nucleus of mice in each group A-D: Representative immunohistochemical staining images (A: Ctrl group; B: CUMS group; C: C-Ctrl group; D: C-CUMS group); E: AgRP positive area in Ctrl group and CUMS group; F: AgRP positive area in C-Ctrl group and C-CUMS group. *P < 0.05. n=3, x±s. AgRP: Agouti-related protein; Ctrl: Control; CUMS: Chronic unpredictable mild stress; C-Ctrl: Cessation-control; C-CUMS: Cessation-CUMS.

免疫组织化学染色结果显示,CUMS组小鼠下丘脑弓状核中LEPR蛋白表达水平较Ctrl组下降(P<0.01),而C-CUMS组小鼠下丘脑弓状核中LEPR蛋白表达水平较C-Ctrl组升高(P<0.01),见图 3。以上结果提示应激停止后小鼠下丘脑弓状核中LEPR可能在小鼠食欲变化中发挥关键作用。

图 3 各组小鼠下丘脑弓状核LEPR免疫组织化学染色结果 Fig 3 Immunohistochemical staining of LEPR in hypothalamic arcuate nucleus of mice in each group A-D: Representative immunohistochemical staining images (A: Ctrl group; B: CUMS group; C: C-Ctrl group; D: C-CUMS group); E: LEPR positive area in Ctrl group and CUMS group; F: LEPR positive area in C-Ctrl group and C-CUMS group. **P < 0.01. n=3, x±s. LEPR: Leptin receptor; Ctrl: Control; CUMS: Chronic unpredictable mild stress; C-Ctrl: Cessation-control; C-CUMS: Cessation-CUMS.

3 讨论

研究者会根据研究目的选择合适的应激模型,比如CUMS主要涉及味道刺激、昼夜颠倒、束缚、潮湿垫料、空笼、频闪、强光照射36 h等比较温和的刺激,实验时间较长,一般持续2~4个月[3];慢性束缚应激每日给予小鼠3~4 h的束缚刺激,应激周期控制在2~3周[4];睡眠剥夺应激每日对小鼠进行8~12 h的睡眠剥夺,应激周期为1~2周[5]。本实验选择CUMS为应激模型,既很好地模拟了机体在应激状态下心理和生理的变化,又避免了造模本身对躯体的直接损伤。

应激对食欲的影响具有不确定性,有研究表明应激会导致食欲下降[6],也有研究表明应激会导致食欲上升[7]。本研究结果发现,应激第2~11周CUMS模型小鼠的摄食量较对照组小鼠升高,而在第12周应激停止后C-CUMS组小鼠摄食量有降低的趋势,并在第15周表现出与C-Ctrl组差异有统计学意义,表明CUMS对小鼠的食欲有重要影响。

下丘脑是大脑中调节全身内稳态的重要区域,是机体中枢维持能量平衡的核心,对食欲调节有重要影响[8]。已确定与食欲相关的调节因子如AgRP和促食欲素等具有促进摄食的作用,而阿片黑素促皮质激素原(pro-opiomelanocortin,POMC)、黑皮质素及瘦素具有抑制摄食的作用[9-10]。下丘脑中的AgRP是增加食欲的主要调节因子,在摄食和能量平衡中起着重要作用[11]。本研究中,CUMS组小鼠AgRP mRNA表达量和蛋白表达水平均高于Ctrl组,而C-CUMS组与C-Ctrl组比较差异无统计学意义,提示CUMS小鼠食欲调节机制可能与AgRP有关。

食欲素是一种在神经系统内广泛分布的神经肽类物质,与其受体结合后激活下游信号通路,进而参与调节摄食的生理过程[12]。食欲素以2种形式存在,即食欲素A(orexin-A)和食欲素B(orexin-B)。食欲素受体有2种亚型,即OX1R和食欲素2型受体(orexin 2 receptor,OX2R)。OX1R只对食欲素A具有亲和力,而OX2R对食欲素A和食欲素B具有相同的亲和力。与食欲素B和OX2R相比,食欲素A和OX1R在调节摄食行为方面的作用更大。有趣的是,食欲素在增加食物摄入量的同时也会增加能量消耗,导致体重下降[13-14]。本研究中CUMS组小鼠OX1R mRNA表达量高于Ctrl组,C-CUMS组与C-Ctrl组OX1R mRNA表达量的差异无统计学意义。这一结果与CUMS导致小鼠摄食量增高的情况一致,而小鼠体重无明显变化。由此推测,CUMS后小鼠食欲上升、体重却无明显变化,食欲素很可能发挥了关键作用。

瘦素是白色脂肪细胞分泌的一种蛋白质,被认为是多种生理过程(包括食欲和能量代谢)的主要调节因子之一。它通过与多种受体结合的方式调节食物的摄入,可与下丘脑中的LEPR相结合抑制食欲,也可通过多种信号转导途径作用于POMC神经元,促其产生抑制食欲的α-黑素细胞刺激素,同时亦可作用于AgRP神经元并抑制其产生促进食欲的AgRP,引起食欲降低、机体能量消耗增加[15-17]。本研究中CUMS组小鼠LEPR mRNA和蛋白的表达水平均低于Ctrl组,而C-CUMS组LEPR mRNA和蛋白的表达水平均高于C-Ctrl组,表明应激停止后小鼠食欲下降的机制可能涉及LEPR的功能调节,但相关调节通路还有待进一步研究。一般生理状态下体重的升降与摄食量多少呈正相关,本研究发现应激导致小鼠摄食量增加时体重却没有升高,可能跟应激诱导的代谢稳态失衡有关。小鼠应激时摄取的依然是标准饲料,不能补偿应激所消耗的能量,所以体重没有增加。我们推测应激时AgRP可能抑制了瘦素与LEPR的结合,产生了促进食欲的作用。

综上所述,本研究初步证明CUMS导致小鼠食欲上升,其机制可能与LEPR和AgRP的功能调节有关;应激停止后小鼠食欲下降且体重降低,LEPR可能在此过程中发挥了重要作用。

参考文献
[1]
陈一尘, 罗一君. 应激的评估及其对进食行为的影响[J]. 心理学进展, 2019, 9(10): 1755-1766. DOI:10.12677/AP.2019.910213
[2]
SIEMIAN J N, ARENIVAR M A, SARSFIELD S, et al. Hypothalamic control of interoceptive hunger[J]. Curr Biol, 2021, 31(17): 3797-3809.e5. DOI:10.1016/j.cub.2021.06.048
[3]
ABDELMEGUID N E, HAMMAD T M, ABDEL-MONEIM A M, et al. Effect of epigallocatechin-3-gallate on stress-induced depression in a mouse model: role of interleukin-1β and brain-derived neurotrophic factor[J]. Neurochem Res, 2022, 47(11): 3464-3475. DOI:10.1007/s11064-022-03707-9
[4]
王霞, 王少贤, 方朝义, 等. 慢性束缚应激大鼠下丘脑弓状核食欲调控因子的变化[J]. 中国病理生理杂志, 2019, 35(3): 473-478. DOI:10.3969/j.issn.1000-4718.2019.03.015
[5]
刘陶, 郁婧, 钱召强, 等. 两种睡眠剥夺模型对小鼠情绪和学习记忆的影响[J]. 陕西师范大学学报(自然科学版), 2018, 46(5): 77-82. DOI:10.15983/j.cnki.jsnu.2018.05.354
[6]
ORTOLANI D, GARCIA M C, MELO-THOMAS L, et al. Stress-induced endocrine response and anxiety: the effects of comfort food in rats[J]. Stress, 2014, 17(3): 211-218. DOI:10.3109/10253890.2014.898059
[7]
ZHAO Y L, GAO P H, XIAO Q, et al. Effects of stress on appetite in rats through hypothalamus and stomach ghrelin pathway[J]. Lab Anim Comparat Med, 2021, 41(3): 207-214. DOI:10.12300/j.issn.1674-5817.2020.160
[8]
YOO E S, YU J, SOHN J W. Neuroendocrine control of appetite and metabolism[J]. Exp Mol Med, 2021, 53(4): 505-516. DOI:10.1038/s12276-021-00597-9
[9]
SMITKA K, PROCHAZKOVA P, ROUBALOVA R, et al. Current aspects of the role of autoantibodies directed against appetite-regulating hormones and the gut microbiome in eating disorders[J]. Front Endocrinol (Lausanne), 2021, 12: 613983. DOI:10.3389/fendo.2021.613983
[10]
LEI M M, WU S Q, SHAO X B, et al. Creating leptin-like biofunctions by active immunization against chicken leptin receptor in growing chickens[J]. Domest Anim Endocrinol, 2015, 50: 55-64. DOI:10.1016/j.domaniend.2014.09.001
[11]
KRASHES M J, KODA S, YE C, et al. Rapid, reversible activation of AgRP neurons drives feeding behavior in mice[J]. J Clin Invest, 2011, 121(4): 1424-1428. DOI:10.1172/JCI46229
[12]
SONG Y, LI J, LI H, et al. The role of ventral tegmental area orexinergic afferents in depressive-like behavior in a chronic unpredictable mild stress (CUMS) mouse model[J]. Biochem Biophys Res Commun, 2021, 579: 22-28. DOI:10.1016/j.bbrc.2021.09.062
[13]
BERNER L A, BROWN T A, LAVENDER J M, et al. Neuroendocrinology of reward in anorexia nervosa and bulimia nervosa: beyond leptin and ghrelin[J]. Mol Cell Endocrinol, 2019, 497: 110320. DOI:10.1016/j.mce.2018.10.018
[14]
TESKE J A, MAVANJI V. Energy expenditure: role of orexin[J]. Vitam Horm, 2012, 89: 91-109. DOI:10.1016/B978-0-12-394623-2.00006-8
[15]
GARCIA N W, PFENNIG K S, BURMEISTER S S. Leptin manipulation reduces appetite and causes a switch in mating preference in the plains spadefoot toad (Spea bombifrons)[J]. PLoS One, 2015, 10(4): e0125981. DOI:10.1371/journal.pone.0125981
[16]
FENG H, ZHENG L, FENG Z, et al. The role of leptin in obesity and the potential for leptin replacement therapy[J]. Endocrine, 2013, 44(1): 33-39. DOI:10.1007/s12020-012-9865-y
[17]
BARRIOS-CORREA A A, ESTRADA J A, CONTRERAS I. Leptin signaling in the control of metabolism and appetite: lessons from animal models[J]. J Mol Neurosci, 2018, 66(3): 390-402. DOI:10.1007/s12031-018-1185-0