大鼠DMV注射AVP对胃运动和胃酸分泌的影响及其机制的研究
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摘要
大鼠束缚-浸水应激(restraint water-immersion stress, RWIS)条件下,胃运动和胃酸分泌机能均发生紊乱。其中在调控胃肠功能的较高级中枢下丘脑前部,视上核(SON)、室旁核(PVN)神经元活动最强烈,在初级中枢延髓,迷走神经背核(DMV)神经元活动最强烈,这些结果提示,可能有SON→DMV→胃的神经调控环路的存在。若有,其神经递质及其受体是什么,至今尚不明确。已知SON、PVN主要由精氨酸加压素(AVP)能和催产素(OT)能神经元组成,已有文献报道:OT能神经元有纤维末梢止于DMV;向DMV注射OT可抑制胃运动,刺激胃酸分泌,说明OT可通过DMV对胃机能进行调控,那么,AVP能神经元是否存在于DMV并对胃机能进行调控呢?至今未见有文献报道。本研究通过向DMV微量注射AVP,观察对胃运动和胃酸分泌的影响来对这一问题进行探讨,并对其作用机制进行了初步研究。
本文首先探讨了向DMV注射AVP对胃运动的影响,实验分两步:
第一步:观察大鼠右侧DMV微量注射AVP及AVP受体阻断剂对胃运动的影响。实验分四组:一组(对照组)右侧DMV注射生理盐水(0.2μL),二组右侧DMV注射0.18nmol AVP (0.2μL),三组右侧DMV注射0.018 nmol AVP(0.2μL),四组右侧DMV预先注射0.32 nmol AVP V1a受体阻断剂SR49059 (0.2μL),再向DMV注射0.018 nmol AVP(0.2μL)。用幽门部放置水囊的方法测定大鼠胃运动,观察注药前后的胃运动变化。统计指标包括注射前5min、注射后25min内胃的收缩幅度、时程以及胃运动指数,同时记录呼吸、血压、心电。结果:两种不同剂量的AVP被注射到右侧DMV后5min内,胃运动、呼吸、心率受到显著性抑制,平均血压无明显变化;随着时间的推移,AVP的抑制作用逐渐消失。但生理盐水被注射到右侧DMV后与注射前相比较,胃运动、呼吸、血压、心率均无明显改变。右侧DMV预先注射AVP V1a受体阻断剂SR49059,AVP对胃运动的抑制作用完全消除,这表明DMV中的AVP敏感神经元确实可抑制胃运动,且AVP的这种抑制作用是通过AVP V1a受体实现的。
第二步是在第一步的基础上,通过预先股静脉注射植物性神经节胆碱能N1型受体阻断剂六烃季铵(8μmol,1mL),再向DMV注射AVP,用同样方法观察胃运动的变化,初步探讨了AVP在DMV内调控胃运动的神经元类型。结果:预先静脉注射六烃季铵后,胃运动几乎完全消失,平均动脉压显著降低,呼吸频率、心率无明显变化;待胃运动稍有恢复后,再向DMV内微量注射AVP,胃运动无明显变化。表明DMV内AVP敏感神经元对胃运动的调控是通过节前胆碱能神经元来实现的。
本研究还探讨了AVP通过DMV对胃酸分泌的调控,实验分两组:一组(对照组)右侧DMV注射生理盐水(0.2μL),二组右侧DMV注射0.18 nmol AVP (0.2μL),通过食道插管灌流37℃生理盐水,幽门插管收集并用精密pH计测定灌流液,统计灌流液中的H+分泌量。结果:与注药前相比,注药后生理盐水组胃液H+分泌量无明显变化,AVP组胃液H+分泌量显著增加。这表明大鼠DMV微量注射AVP可促进胃液H+的分泌。
结论:大鼠DMV内微量注射AVP可抑制胃运动,促进胃酸分泌。其调控机制可能为:AVP与DMV神经元胞体膜或树突膜上的AVP V1a型受体结合,通过节前胆碱能-节后NANC能抑制性迷走神经通路实现对胃运动的抑制性调控作用,通过节前胆碱能-节后胆碱能兴奋性神经通路实现对胃酸分泌的兴奋性调控作用。
We previously used the Restraint Water-Immersion Stress (RWIS)model in rats to study the neuronal pathways activated during the RWIS. After different durations of RWIS, neuronal activation, demonstrated by Fos-immunoreactivity (Fos-IR), was found significantly increased in specific brain areas, such as the medullary visceral zone [mainly dorsal motor nucleus of the vagus (DMV), nucleus of solitary tract (NTS), area postrema (AP), and nucleus ambiguus(NA)] and the hypothalamus [mainly supraoptic nucleus (SON) and paraventricular nucleus (PVN)]. Among which DMV and SON neuronal activities degree were the most. Whether the vasopressinergic pathway from the PVN/SON to the DMV plays a role in regulating the gastric function? To clarify the question, the aim of the present study was to investigate the effect of arginine-vasopressin (AVP) microinjected into the DMV on gastric motor and secretory functions.
The study was divided into three parts.
Experiment 1, to investigate the effect of AVP in DMV on gastric motility.
(1) Two doses (0.018 nmol and 0.18 nmol) of AVP (0.2μL) were microinjected into the DMV to investigate the effect of AVP.
(2) Control experiment. Normal saline (0.2μL ) was microinjected into the DMV .
Experiment 2, to investigate the mechanism involved in the effect of AVP on regulating the gastric motility.
Two groups of rats with six in each were pretreated with SR49059 (0.32 nmol in 0.2μL into DMV) 15min or hexamethonium (8μmol in 1ml,i.v. ) 30min before the AVP was administrated.
Experiment 3, to investigate the effect of AVP in DMV on gastric acid secretion,.
(1) AVP (0.18 nmol in 0.2μL) was microinjected into the DMV to investigate the effect of AVP .
(2) Control experiment. Normal saline (0.2μL) was microinjected into the DMV.
A latex balloon connected with a pressure transducer was inserted into the pylorus to record the gastric motility. Gastric acid output was collected every 20 min after saline flush with the method of esophageal perfusion, the pH value of the collected juice was measured by precision pH meter.The blood pressure, electrocardiogram and respiration of the rats were continuously monitored by the BL-420 Biological Experimental System.
Microinjection of AVP (0.018nmol and 0.18nmol in 0.2μL) microinjected into right DMV significantly inhibited the gastric motility. The inhibitory motility effect of AVP (0.18μmol in 0.2μL) was completely blocked by SR49059 (0.32nmol in 0.2μL), the specific AVP V1a receptor antagonist, and abolished by hexamethonium (8μmol in 1ml, i.v.), the neuronal nicotinic cholinergic receptor antagonist.
Conclusion: These results indicated that, AVP inhibited the gastric motility and excited the acid secretion by activating the specific V1a AVP receptors in DMV, and the cholinergic neuron pathway may be involved.
本文首先探讨了向DMV注射AVP对胃运动的影响,实验分两步:
第一步:观察大鼠右侧DMV微量注射AVP及AVP受体阻断剂对胃运动的影响。实验分四组:一组(对照组)右侧DMV注射生理盐水(0.2μL),二组右侧DMV注射0.18nmol AVP (0.2μL),三组右侧DMV注射0.018 nmol AVP(0.2μL),四组右侧DMV预先注射0.32 nmol AVP V1a受体阻断剂SR49059 (0.2μL),再向DMV注射0.018 nmol AVP(0.2μL)。用幽门部放置水囊的方法测定大鼠胃运动,观察注药前后的胃运动变化。统计指标包括注射前5min、注射后25min内胃的收缩幅度、时程以及胃运动指数,同时记录呼吸、血压、心电。结果:两种不同剂量的AVP被注射到右侧DMV后5min内,胃运动、呼吸、心率受到显著性抑制,平均血压无明显变化;随着时间的推移,AVP的抑制作用逐渐消失。但生理盐水被注射到右侧DMV后与注射前相比较,胃运动、呼吸、血压、心率均无明显改变。右侧DMV预先注射AVP V1a受体阻断剂SR49059,AVP对胃运动的抑制作用完全消除,这表明DMV中的AVP敏感神经元确实可抑制胃运动,且AVP的这种抑制作用是通过AVP V1a受体实现的。
第二步是在第一步的基础上,通过预先股静脉注射植物性神经节胆碱能N1型受体阻断剂六烃季铵(8μmol,1mL),再向DMV注射AVP,用同样方法观察胃运动的变化,初步探讨了AVP在DMV内调控胃运动的神经元类型。结果:预先静脉注射六烃季铵后,胃运动几乎完全消失,平均动脉压显著降低,呼吸频率、心率无明显变化;待胃运动稍有恢复后,再向DMV内微量注射AVP,胃运动无明显变化。表明DMV内AVP敏感神经元对胃运动的调控是通过节前胆碱能神经元来实现的。
本研究还探讨了AVP通过DMV对胃酸分泌的调控,实验分两组:一组(对照组)右侧DMV注射生理盐水(0.2μL),二组右侧DMV注射0.18 nmol AVP (0.2μL),通过食道插管灌流37℃生理盐水,幽门插管收集并用精密pH计测定灌流液,统计灌流液中的H+分泌量。结果:与注药前相比,注药后生理盐水组胃液H+分泌量无明显变化,AVP组胃液H+分泌量显著增加。这表明大鼠DMV微量注射AVP可促进胃液H+的分泌。
结论:大鼠DMV内微量注射AVP可抑制胃运动,促进胃酸分泌。其调控机制可能为:AVP与DMV神经元胞体膜或树突膜上的AVP V1a型受体结合,通过节前胆碱能-节后NANC能抑制性迷走神经通路实现对胃运动的抑制性调控作用,通过节前胆碱能-节后胆碱能兴奋性神经通路实现对胃酸分泌的兴奋性调控作用。
We previously used the Restraint Water-Immersion Stress (RWIS)model in rats to study the neuronal pathways activated during the RWIS. After different durations of RWIS, neuronal activation, demonstrated by Fos-immunoreactivity (Fos-IR), was found significantly increased in specific brain areas, such as the medullary visceral zone [mainly dorsal motor nucleus of the vagus (DMV), nucleus of solitary tract (NTS), area postrema (AP), and nucleus ambiguus(NA)] and the hypothalamus [mainly supraoptic nucleus (SON) and paraventricular nucleus (PVN)]. Among which DMV and SON neuronal activities degree were the most. Whether the vasopressinergic pathway from the PVN/SON to the DMV plays a role in regulating the gastric function? To clarify the question, the aim of the present study was to investigate the effect of arginine-vasopressin (AVP) microinjected into the DMV on gastric motor and secretory functions.
The study was divided into three parts.
Experiment 1, to investigate the effect of AVP in DMV on gastric motility.
(1) Two doses (0.018 nmol and 0.18 nmol) of AVP (0.2μL) were microinjected into the DMV to investigate the effect of AVP.
(2) Control experiment. Normal saline (0.2μL ) was microinjected into the DMV .
Experiment 2, to investigate the mechanism involved in the effect of AVP on regulating the gastric motility.
Two groups of rats with six in each were pretreated with SR49059 (0.32 nmol in 0.2μL into DMV) 15min or hexamethonium (8μmol in 1ml,i.v. ) 30min before the AVP was administrated.
Experiment 3, to investigate the effect of AVP in DMV on gastric acid secretion,.
(1) AVP (0.18 nmol in 0.2μL) was microinjected into the DMV to investigate the effect of AVP .
(2) Control experiment. Normal saline (0.2μL) was microinjected into the DMV.
A latex balloon connected with a pressure transducer was inserted into the pylorus to record the gastric motility. Gastric acid output was collected every 20 min after saline flush with the method of esophageal perfusion, the pH value of the collected juice was measured by precision pH meter.The blood pressure, electrocardiogram and respiration of the rats were continuously monitored by the BL-420 Biological Experimental System.
Microinjection of AVP (0.018nmol and 0.18nmol in 0.2μL) microinjected into right DMV significantly inhibited the gastric motility. The inhibitory motility effect of AVP (0.18μmol in 0.2μL) was completely blocked by SR49059 (0.32nmol in 0.2μL), the specific AVP V1a receptor antagonist, and abolished by hexamethonium (8μmol in 1ml, i.v.), the neuronal nicotinic cholinergic receptor antagonist.
Conclusion: These results indicated that, AVP inhibited the gastric motility and excited the acid secretion by activating the specific V1a AVP receptors in DMV, and the cholinergic neuron pathway may be involved.
引文
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