【作者】 陈曦；

【导师】 蒋正尧；

【作者基本信息】 青岛大学 ， 生理学， 2012， 博士

【摘要】 Nesfatin-1是来源于NUCB2的一种抑食肽。NUCB2的mRNA在下丘脑和脑干的摄食调控相关核团均有表达,如弓状核(ARC)、室旁核(PVN)、下丘脑外侧区(LHA)、孤束核(NTS)等。已知在自由进食的大鼠侧脑室、第三、第四脑室、小脑延髓池、室旁核注射nesfatin-1可显著抑制摄食。为明确nesfatin-1在大脑中的作用位点,应在核团局限注射观察其抑制摄食的作用。下丘脑和脑干的神经环路参与nesfatin-1抑制摄食的机制,其内的葡萄糖感受性神经元参与低血糖引发的摄食调节以及血糖水平的稳态调节。很多摄食相关肽可通过调制葡萄糖感受性神经元的兴奋性参与摄食调节。并且进食也可影响摄食相关肽的分泌。目的：探讨nesfatin-1在下丘脑参与摄食调控的机制以及观察进食是否影响血浆nesfatin-1的水平。方法：在体细胞外电生理记录,行为学方法,ELASA等实验方法。实验结果：1.与对照组核团内注射生理盐水相比,在PVN内核团注射50pmol的nesfatin-1可抑制大鼠1-5小时内的累积摄食量,抑制百分率分别为58.6%,61.4%,44.5%,41.9%,30.4%。2.LHA内注射50pmol的nesfatin-1可显著抑制1-5小时内的累积摄食量,但抑制的幅度比PVN要小,1-5小时的抑制百分率分别为38.9%,34.0%,22.3%,25.4%,17.6%。3.腹内侧核(VMN)内注射50pmol的nesfatin-1仅可抑制第2小时累积摄食量,且抑制幅度小于PVN和LHA。4.PVN内注射50pmol的nesfatin-1可显著抑制2-6小时的累积饮水量,但在LHA和VMN中却无此效应。5.在PVN中记录到97个自发性放电的神经元,64个对微量加压注射葡萄糖溶液有反应,其中27个鉴定为葡萄糖兴奋性(GE)神经元,37个鉴定为葡萄糖抑制性(GI)神经元。Nesfatin-1可兴奋73.0%的GI神经元,而对13.5%的GI神经元则具有抑制作用,对13.5%的GI神经元没有作用。因此nesfatin-1对PVN中GI神经元主要为兴奋作用,可将其放电频率从20.6±4.8 Hz增加至32.7±6.5 Hz(P<0.01)。6.在LHA中记录到118个自发放电的神经元中,GI神经元占28.8%,葡萄糖将其频率从5.3±1.0 Hz降低至2.4±0.5Hz(P<0.01)。在34个GI神经元中,nesfatin-1可抑制其中的24个,4个神经元则被兴奋。Nesfatin-1可将GI神经元的放电频率从4.8±1.5 Hz降低至1.6±0.4 Hz(P<0.01)。7.在VMN记录到的神经元中,35个(35/77,45.4%)为GE神经元,6个(6/77,7.7%)为GI神经元。Nesfatin-1对其中的GE神经元(27/35,77.1%)的主要作用为兴奋作用,可将其放电频率从3.4±0.2 Hz增加至6.7±0.4 Hz(n=27,P<0.01)。另有3个GE神经元可被抑制,对其中的5个GE神经元没有作用。8.在MSG处理的小鼠的侧脑室中注射nesfatin-1,与对照组相比,可显著抑制第2,3,5小时内的累积摄食量和第2,3,4,5小时内累积饮水量。与盐水处理组侧脑室注射nesfatin-1相比,nesfatin-1并未改变MSG处理组小鼠效应最强的第2小时的累积摄食量和累积饮水量。9.口服生理盐水、葡萄糖溶液、蛋白质粉溶液、脂肪乳溶液并不影响人血浆nesfatin-1的水平。结论：1. Nesfatin-1可能至少部分是通过作用于下丘脑PVN. LHA、VMN中的葡萄糖感受神经元,调制其兴奋性来发挥抑制摄食的作用。2. Nesfatin-1可通过作用于下丘脑室旁核发挥抑制饮水的作用。3.ARC在nesfatin-1抑制摄食的机制中只是起到部分作用。4.进食不同类型食物不影响血浆nesfatin-1水平。更多还原

【Abstract】 Nesfatin-1, derived from the precursor NEFA/nucleobindin 2 (NUCB2), was recently identified as anorexigenic signal. NUCB2 mRNA expression was detected in rat hypothalamic and brainstem nuclei implicated in the regulation of food intake, notably in the ARC, PVN, LHA and NTS. Nesfatin-1 has been shown to inhibit food intake after lateral, third, or fourth brain ventricle, cisterna magna administration, or PVN injection in ad libitum fed rats. With regards to the understanding of nesfatin-1’s brain sites of action, additional microinjection studies will be necessary to define specific nuclei responsive to nesfatin-1 to get insight into the differential effects on food intake. Neural mechanisms involved in nesfatin-1’s anorexigenic effect encompass the recruitment of several hypothalamic and medullary anorexigenic pathways. Glucosensing neurons located in the hypothalamus and lower brainstem are involved in glucoprivic feeding and homeostatic control of blood glucose. Feeding related peptides modulate glucose sensing neurons to control food intake. Food intake also affects the secretion of feeding related peptides.Object:To shed light on the mechanisms by which nesfatin-1 exerts its satiety-promoting actions and to investigate the influence of food intake on plasm nesfatin-1’s level.Methods:in vivo extracellular recording, behavioral test and ELASA were performed in the present studies.Results:1. Nesfatin-1 injection (50 pmol) into the PVN significantly reduced cumulative food intake by 58.6%,61.4%,44.5%,41.9% and 30.4%, respectively, compared with vehicle-injected controls at the end of 1,2,3,4 and 5 h after injection into the PVN.2. Nesfatin-1 (50 pmol) significantly reduced feeding at the end of 1-5 h after injection into the LHA but of a less magnitude than that observed in the PVN. Nesfatin-1 decreased cumulative food intake by 38.9%,34.0%,22.3%,25.4% and 17.6%, respectively, compared with vehicle-injected controls at the end of 1,2,3,4 and 5h after injection into the LHA. 3. The VMN exhibited a delayed inhibitory response to 50 pmol nesfatin-1, observed at the end of 2 h postinjection. The response was markedly less than the feeding response at the end of 2 h after PVN and LHA injection.4. Nesfatin-1 (50 pmol) significantly reduced cumulative water intake at the end of 2-6 h after injection into the PVN but not in the LHA and VMN.5. Among 97 recorded neurons in PVN,64 units responded to micro-injection of glucose. 27 were GE neurons and 37 were GI neurons. The main effect was excitatory after intra-PVN administration of nesfatin-1 in GI neurons. Micro-injection of nesfatin-1 significantly increased the spontaneous firing rate of GI neurons from 20.6±4.8 Hz to 32.7±6.5 Hz (P<0.01). In 37 GI neurons,73.0% were excited,13.5% were inhibited, and 13.5% were not affected.6. Of 118 neurons in the LHA tested,28.8% were inhibited by glucose (GI neurons). Glucose decreased the firing rate from 5.3±1.0 Hz to 2.4±0.5Hz (P<0.01). Of 34 GI neurons in the LHA tested, inhibitory effects were seen in 24 of GI neurons while the 4 cells were excited in response to nesfatin-1. Micro-injection of nesfatin-1 significantly decreased the spontaneous firing rate of GI neurons from 4.8±1.5 Hz to 1.6±0.4 Hz (P<0.01).7. In VMN,35 (35/77,45.4%) were GE neurons and 6 (6/77,7.7%) were GI neurons. The main effects were excitatory after intra-VMN administration of nesfatin-1 in GE neurons (27/35,77.1%). Micro-injection of nesfatin-1 significantly increased the spontaneous firing rate of GE neurons from 3.4±0.2 to 6.7±0.4 Hz (n=27, P<0.01). Of 35 GE neurons in VMN,3 were inhibited, and 5 were not affected by nesfatin-1.8. I.c.v injection of nesfatin-1 significantly reduced cumulative food intake at the end of 2,3,5 h and decreased cumulative water intake at the end of 2,3,4,5 h in MSG-treated mice. Compared with saline-treated mice, nesfatin-1 didn’t change the cumulative food and water intake in MSG-treated mice at the end of 2h after injection.9. Oral saline, glucose solution, protein powder solution and fat emulsion did not affect human plasma nesfatin-1 level. Conclusion:1. Nesfatin-1 may exert at least a part of its physiological actions on the control of food intake as a direct result of its role in controlling the excitability of glucosensing neurons in the LHA, VMN and PVN.2. Nesfatin-1 inhibits water intake in the PVN.3. ARC plays a partial role in nesfatin-1’s regulation of food intake.4. Macronutrients do not influence postprandial plasma nesfatin-1 level.更多还原