5-羟色胺7受体对排尿影响的研究进展

目前,许多研究证实5-羟色胺（5-HT）受体对控制排尿有着显著作用,其中尤以5-HT1A受体为甚,其次为5-HT7、5-HT2和5-HT3受体,这些受体似乎调控着排尿的各个通路,包括交感神经、副交感神经及体神经通路。其中5-HT7受体对逼尿肌收缩有调控作用。深入开展5-HT7受体对排尿影响的研究,将有助于进一步阐明泌尿反射调控的机制及研发新的有效治疗尿失禁、膀胱过度活动症等疾病的药物。     

5-羟色胺1A受体激动剂改善脊髓损伤大鼠排尿功能障碍的实验研究

目的 研究5-羟色胺-1A(5-HTlA)受体激动剂对慢性脊髓损伤(spinal cord injury,SCI)大鼠排尿障碍的改善作用. 方法 雌性SD大鼠14只,体质量175 ～ 200 g.随机分为2组:实验组7只显微镜下大鼠T10棘突水平行脊髓离断建立脊髓损伤模型,正常对照组7只.8周后乌拉坦(1.3 g/kg)麻醉下,2组大鼠颈静脉和膀胱内置管,连接压力感受器,记录膀胱最大容量、残余尿量、排尿量和尿道外括约肌的肌电活动.静脉注入5-HTI A/7受体激动剂8-羟基-丙胺-四氢萘(8-OH-DPAT,0.03 ～1.00 mg/kg),得出剂量—效应曲线后再给予5HTIA受体抑制剂WAY-100635 (0.3g/kg).观察比较2组大鼠用药前后尿动力学指标的变化. 结果 随着8-OH-DPAT剂量增加,SCI大鼠膀胱容量从(33.2 ±8.3)ml降至(22.8±2.4) ml,排尿量从(0.14±0.08)ml增至(0.38 ±0.09) ml,残余尿量从(3.68±1.36)ml降至(1.84±0.21)ml,而膀胱最高压力从(27.1±3.6)mm Hg(1 mm Hg=0.133 kPa)降至(22.8±2.4) mm Hg,用药前后差异均有统计学意义(P＜0.05).对照组大鼠用药前后排尿情况变化差异无统计学意义.肌电图显示8-OH-DPAT引起SCI大鼠尿道外括约肌强直收缩中出现阶段性的松弛,正常对照组大鼠作用无明显改变. 结论 8-OH-DPAT可以剂量依赖性地部分恢复SCI大鼠的尿道外括约肌协调性松弛,从而降低膀胱容量,增加排尿量,减少残尿量,增加排尿效率,改善排尿障碍.     

5-HT7受体激动剂改善脊髓损伤大鼠排尿障碍

目的:研究五羟色胺-7受体激动剂对慢性脊髓损伤(spinal cord injury,SCI)大鼠排尿障碍的改善作用。方法:体重175～200g之间的SD雌性大鼠,脊髓损伤模型7只,正常对照8只。乌拉坦麻醉下,在正常大鼠和脊髓损伤的模型的大鼠颈静脉和膀胱内置管,连接压力感受器,记录膀胱最大容量、剩余尿量、排尿量和尿道外括约肌的肌电活动。静脉注入5-HT7受体激动剂LP44(3～300μg/kg)得到剂量-效应曲线随后再给予5-HT7受体抑制剂SB269970(100μg/kg)。结果:LP44剂量依赖性的降低SCI大鼠膀胱最大容量,并且增加排尿量、减少剩余尿量从而增加排尿效率。但是不能引起脊髓完好的大鼠排尿的改变。SB269970注入后可以逆转LP44的作用。同时LP44引起脊髓损伤大鼠尿道外括约肌强直收缩中出现阶段性的松弛,而对于脊髓完好的大鼠作用无明显改变。结论:LP44可以剂量依赖性地部分恢复SCI大鼠的尿道外括约肌协调性松弛,从而降低膀胱容量,增加排尿量,减少剩余尿量,结果增加排尿效率,改善排尿障碍。     

曲马多对急性心肌缺血大鼠丘脑束旁核5-HT1A受体mRNA表达的影响

心肌缺血时通过中枢和外周痛觉敏感化机制,心脏交感传入冲动增加,从而有可能诱发心绞痛。但由心肌缺血引发的心绞痛(症状)和急性心肌梗死(损伤)的程度往往不相平行。丘脑束旁核(parafascicular nucleus,Pf)是内脏痛觉感受、调制和整合的重要中枢,Pf内含有丰富的5-HT能神经末梢及其受体。研究表明,5-HT1A受体在大鼠内脏痛模型中发挥着重要的作用,但目前对5-HT1A受体mRNA的研究多局限于外周炎症后脊髓水平的表达,心肌缺血时Pf水平 5-HT1A受体mRNA的表达尚无定论。本研究拟观察曲马多对急性心肌缺血大鼠Pf部位5-HT1A受体mRNA表达的影响。     

大鼠5-羟色胺1A受体超表达细胞株的构建

目的 构建大鼠5-羟色胺1A受体(5-HT1AR)超表达细胞株.方法 从雄性SD大鼠脑组织中提取总RNA,构建真核表达质粒pc-DNA3.1/hisC-Rat-5-HT1AR.提取和纯化该真核表达质粒,并采用脂质转染法转染到人神经母细胞瘤细胞SH-SY5Y细胞.经G418筛选,获得SH-SY5Y-Rat-5-HT1AR细胞株.采用Western blot法鉴定5-HT1AR蛋白表达;光镜下观察细胞的形态;四甲基偶氮唑盐比色法测定细胞活力;细胞免疫荧光染色后,共聚焦显微镜下观察5-HT1AR表达.结果 成功构建了重组质粒pc-DNA3.1/hisC-Rat-5-HT1AR.重组质粒顺利转染SH-SY5Y细胞,构建了稳定表达大鼠5-HT1AR的细胞株SH-SY5Y-Rat-5-HT1AR.SH-SY5Y-Rat-5-HT1AR细胞呈梭状,细长,突起长而少.SH-SY5Y-Rat-5-HT1AR细胞的活力明显低于SH-SY5Y细胞.共聚焦显微镜下5-HT1AR主要在细胞膜表达.结论 成功地构建了大鼠5-HT1AR超表达细胞株.

Abstract：

Objective To establish a cell line with overexpression of rat serotonin1A receptor (5-HT1AR).Methods Human neuroblastoma cells-SH-SY5Y were donated by cancer institute attached to the 4 th Affiliated Hospital, Hebei Medical University. Total RNA was extracted from brain tissues of male SD rats and rat 5-HT1A R was obtained by RT-PCR. Plasmid pc-DNA3. 1/hisC containing the rat 5-HT1AR (pc-DNA3.1/hisC-Rat-5-HT1AR)was constructed and transfected into SH-SY5Y cells. The transfected cells were isolated by G418 selection and SH-SY5Y-Rat-5-HT1A R cells were obtained. Expression of 5-HT1A R was detected by Western blot analysis. Cell viability was evaluated by MTT assay. SH-SY5Y-Rat-5-HT1AR cells were further observed for 5-HT1AR by immuno-fluorescence staining. Results Plasmid pc-DNA3. 1/hisC-Rat-5-HT1AR was successfully constructed by linking Rat-5-HT1A R with pc-DNA3.1/hisC and transfected into SH-SY5Y. The SH-SY5Y-Rat-5-HT1A R cells were more slender than SH-SY5Y cells with less and longer processes. MTT showed that the viability of SH-SY5Y-Rat-5-HT1A R cells was much lower than SH-SY5Y. Rat 5-HT1A R was expressed efficiently on the membrane of SH-SY5Y-Rat-5-HT1A R cells. Conclusion A cell line with overexpress of rat 5-HT1A R is successfully established.     

远位触液神经元5-HT1A受体在大鼠神经病理性痛中的作用

目的 评价远位触液神经元5-羟色胺1A(5-HT1A)受体在大鼠神经病理性痛中的作用.方法 雄性SD大鼠40只,体重230～270 g,采用随机数字表法,将其随机分为4组(n=10):假手术组(S组)、神经病理性痛组(NP组)、二甲基亚砜组(DMSO组)和8-羟基-2-(双-正丙胺基)-四氢萘满组(8-OH-DPAT组).采用坐骨神经慢性压迫法(CCI制备大鼠神经病理性痛模型,S组仅暴露坐骨神经,但不结扎.CCI后第7天,8-OH-DPAT组和DMSO组向远位触液神经元分别缓慢注射5-HT1A受体特异性激动剂8-OH-DPAT或DMSO 1 μl,5 min内注射完毕.分别于CCI前(T0)、CCI后第7天(T1)和给药后3、6 h(T2,3)时,测定缩足潜伏期(PWL)和缩足阈值(PWT).于给药后6 h时处死大鼠,取脑组织,采用免疫荧光标记法检测远位触液核神经元5-HT1A受体的表达.结果 与S组比较,NP组、DMSO组和8-OH-DPAT组T1时PWL缩短,PWT降低(P＜0.01);与DMSO组比较,8-OH-DPAT组T2和T3时PWL延长,PWT升高(P＜0.01).与S组比较,NP组和DMSO组5-HT1A受体表达下调(P＜0.01);与NP组和DMSO组比较,8-OH-DPAT组5-HT1A.受体表达上调(P＜0.01);NP组和DMSO组间5-HT1A受体表达比较差异无统计学意义(P＞0.05).结论 远位触液神经元5-HT1A受体参与了大鼠神经病理性痛的调控.

Abstract：

Objective To evaluate the role of 5-HT1A receptors in distal cerebrospinal fluid (CSF)-contacting neurons in neuropathic pain (NP) in rats. Methods Forty male SD rats weighing 230-270 g were randomly divided into 4 groups (n = 8 each): sham operation group (group S); NP group; dimethyl sulfoxide (DMSO) group and 8-OH-DPAT (a specific 5-HT1A receptor agonist) group. NP was induced by chronic constrictive injury (CCI) in groups NP, DMSO and 8-OH-DPAT. Four silk ligatures were placed on the sciatic nerve at 1 mm intervals . In group S, the sciatic nerve was exposed but not ligated. 8-OH-DPAT and DMSO 1 μl were injected into the region where most of CSF-contacting neurons are present over 5 min on 7th day after CCI in groups 8-OH-DPAT and DMSO respectively. Paw withdrawal latency (PWL) and paw withdrawal threshold (PWT) were measured before CCI, on 7th day after CCI, and at 3 and 6 h after administration. The rats were sacrificed 6 h after administration, and the brain tissues removed for determination of the expression of 5-HT1A receptors in the distal CSF-contacting neurons by immunofluorescence. Results Compared with group S, PWL was significantly shorten and PWT decreased at T, in groups NP, DMSO and 8-OH- DPAT (P ＜ 0.01) . Compared with group DMSO, PWL was significantly prolonged and PWT increased at T2 and T3 in group 8-OH-DPAT ( P ＜ 0.01). The 5-HT1A receptor expression was significantly down-regulated in groups NP and DMSO compared with group S, while up-regulated in group 8-OH-DPAT compared with groups NP and DMSO ( P ＜ 0.01). There was no significant difference in 5-HT1A receptor expression between groups NP and DMSO ( P ＞ 0.05). Conclusion 5-HT1A receptors in distal CSF-contacting neurons are involved in the regulation of NP in rats.     

曲马多对神经病理性痛大鼠中脑远位触液神经元5-HT1A受体表达的影响

目的 探讨曲马多对神经病理性痛大鼠中脑远位触液神经元5-HT1A受体表达的影响.方法 SPF级雄性SD大鼠40只,体重220～280 g,采用坐骨神经慢性压迫法制备大鼠神经病理性痛模型,随机分为5组(n=8):正常对照组(C组)、生理盐水组(NS组)、曲马多组(T组)、神经病理性痛+生理盐水组(NP+NS组)和神经病理性痛+曲马多组(NP+T组).C组不行任何处理;NS组和T组仅暴露坐骨神经,分别腹腔注射生理盐水2 ml/kg或曲马多10 mg/kg;NP+NS组和NP+T组制备神经病理性痛模型,模型制备后第7天分别腹腔注射生理盐水2 ml/kg或曲马多10 mg/kg.除C组外,其余4组于腹腔注射曲马多或生理盐水前(T1)和注射后1 h(T2)时测定热痛阈和机械痛阈.于模型制备后第5天,左侧侧脑室注射30%霍乱毒素亚单位B与辣根过氧化物酶复合物(CB-HRP)3μl以标记远位触液神经元,并测定远位触液神经元5-HT1A表达水平.结果 与C组比较,NP+NS组中脑远位触液神经元5-HT1A受体表达下调(P＜0.05),其余组差异无统计学意义(P＞0.05);与NS组和T组比较,NP+NS组中脑远位触液神经元5-HT1A受体表达下调,热痛阈和机械痛阈降低,NP+T组热痛阈和机械痛阈降低(P＜0.05),中脑远位触液神经元5-HT1A受体表达差异无统计学意义(P＞0.05);与NP+NS组比较,NP+T组中脑远位触液神经元5-HT1A受体表达上调,热痛阈和机械痛阈升高(P＜0.05).结论 曲马多可下调中脑远位触液神经元5-HT1A受体的表达,该作用可能是其减轻大鼠神经病理性痛的机制之一.     

5-羟色胺1A受体mRNA在大鼠慢传输便秘结肠组织中的表达

为观察慢传输型便秘大鼠结肠组织中5-羟色胺（5-HT）1A受体的rnRNA表达,选取体重（200±20）g的SD大鼠24只,随机分为两组,模型组和对照组各12只,模型组采用大黄灌胃法造模,对照组予相同体积的蒸馏水灌胃。完成造模后处死,测肠道炭末推进长度和推进率,取出标本。用RT—PCR法测各个标本5HT 1A受体的mRNA表达,并进行统计学分析。结果显示,模型组大鼠结肠5HTlA受体的mRNA表达（0．77±0．14）高于对照组（0．6l±0．21）,有统计学意义（P〈0．05）。结果表明,慢传输型便秘大鼠结肠中5-HTlA受体表达量上升,提示5-HT1A是影响慢性便秘的因素。     

去卵巢大鼠体温变化与下丘脑视前区5-羟色胺_(1A)受体表达

目的观察大鼠去卵巢后体温变化与下丘脑视前区5-羟色胺1A(5-HT1A)受体亚型表达之间的关系。方法用数字温度计TM902C检测大鼠的肛温,酶联免疫吸附试验测定血清激素水平,根据5-HT1A受体互补DNA序列合成相应的特异性引物,用聚合酶链反应(PCR)法观察大鼠去卵巢后下丘脑视前区5-HT1A受体表达。结果 (1)大鼠去卵巢(OVX) 组自第8周末肛温明显高于假去势(sham)组(P<0．05)。(2)OVX组大鼠血清17β-雌二醇(17β-E2)较sham组下降,黄体生成素(LH)水平升高,差异显著(P<0．05)。(3)OVX组下丘脑视前区5-HT1A受体表达增强,与sham组比较有显著性差异(P <0．05)。结论大鼠卵巢摘除后体温升高,下丘脑视前区5-HT1A受体表达增强,提示5-HT在低雌激素状态下的体温调节中起一定作用。     

慢性束缚应激小鼠海马神经元5-羟色胺1A受体表达的变化

目的 探讨慢性束缚应激小鼠海马神经元5-羟色胺1A(5-HT1A)受体表达的变化.方法 BALB/c种系雄性小鼠40只,6～9月龄,体重25～35 g,采用随机数字表法,将其随机分为2组(n=20):正常对照组(C组)和慢性束缚应激组(S组).S组慢性束缚应激模型制备成功后1d,依次进行悬尾实验、明暗穿箱实验和水迷宫实验,悬尾实验中记录静止时间,明暗穿箱实验中记录明亮箱中停留时间,水迷宫实验中记录逃避潜伏期和穿过平台次数.然后处死小鼠,取海马组织,采用免疫组化法测定CA1区和CA3区神经元5-HT1A受体的表达.结果 与C组比较,S组静止时间和逃避潜伏期延长,明亮箱中停留时间缩短,穿过平台次数减少,海马神经元5-HT1A受体表达下调(P＜0.05或O.01).结论 慢性束缚应激诱发小鼠认知功能障碍的机制与下调海马神经元5-HT1A受体表达有关.     

Preclinical evidence for the anxiolytic activity of 5-HT3 receptor antagonists: A review

From ancient history to present times, mankind has sought for anxiolytics, and various medications have been found and consequently used, at present culminating in heavy benzodiazepine use. Side-effects such as dependence and tolerance have always induced the need, and accordingly the search, for new and better treatments. The 5-HT_1A receptor agonists like buspirone are an example of such new therapeutic agents. Whether or not those compounds will indeed be better as well remains to be seen. Recently another new class of putative anxiolytics has been proposed, the 5-HT₃ receptor antagonists. The present article reviews the evidence for anxiolytic activity of this new class of compounds of animal models of anxiety. Compared to the established anxiolytics (benzodiazepines and, to a lesser extent, 5-HT_1A receptor agonists) 5-HT₃ receptor antagonists have a different anxiolytic profile. They are active in a limited number of animal models, they often are very potent and the ratio between therapeutic activity and side-effects is remarkably large. No evidence for tolerance or rebound effects was found, which makes them an attractive alternative to the benzodiazepines. Preliminary human clinical data are controversial; some investigators have reported positive effects in anxiety, others have not been able to demonstrate this.     

The effects of a 5-HT2A receptor antagonist on blood flow in lumbar disc herniation : application of nucleus pulposus in a canine model

Blood vessel clots are found around the nerve root in patients with lumbar disc herniation. Thrombosis formation in the experimental application of nucleus pulposus to the nerve root has been shown in histological studies. In addition, reduction of blood flow and nerve conduction velocity are induced by the application of nucleus pulposus, which mimics lumbar disc herniation. In patients with lumbar disc herniation, nerve root block, which is thought to increase nerve blood flow, improves radiculopathy. 5-HT_2A receptor antagonists are used in chronic arterial occlusive diseases to improve blood flow and have been reported to work as well as nonsteroidal anti-inflammatory drugs in improving radiculopathy due to lumbar disc herniation in clinical studies. This study investigated the effects of a 5-HT_2A receptor antagonist on blood vessel diameter and blood flow in a canine experimental model of lumbar disc herniation. A total of 13 dogs were used. The animals were divided into three experimental groups and surgery was performed 1 week before measurements. In the nucleus pulposus group (NP; n = 5), the nucleus pulposus was applied to the nerve roots from the ventral side. In the sham group (n = 5), nucleus pulposus was not applied. In the naïve group (n = 3), the animals did not undergo surgery. Measurements of vessel diameter and blood flow were done before and after administration of saline and drugs. The diameters and blood flow volume of the observed blood vessels were measured on video-recordings every 10 min for 65 min. In all groups, vessel diameter and blood flow did not change before or after administration of saline. In the NP and sham groups, vessel diameter and blood flow increased significantly after administration of 5-HTRA compared with the naïve group. 5-HTRA improved blood vessel diameter and blood flow in the nerve roots inflamed by the application of nucleus pulposus but not in the intact nerve roots. 5-HTRA might be a potential agent to improve blood flow in the nerve roots of patients with lumbar disc herniation.     

5-HT3 receptor antagonists do not alter spontaneous contraction of pregnant myometrium in vitro

Background5-HT3 receptor antagonists are effective antiemetics for perioperative use. However, their effects on myometrial contractility remain unknown. We examined whether three different 5-HT3 receptor antagonists could affect the contraction of human myometrium.MethodsSamples of human myometrium were taken from parturients undergoing elective cesarean delivery. Effects of ondansetron, granisetron and tropisetron (over a range of 1–10⁴ ng/mL) on spontaneous contraction (ratios of amplitude, interval, and duration of the contraction) were examined and compared to saline controls (n=6 for each agent).ResultsNone of the three 5-HT3 receptor antagonists significantly affected myometrial contraction.Conclusion5-HT3 receptor antagonists do not affect the contraction of myometrial strips isolated from term pregnant women.     

The 5-HT2c receptor gene Cys23Ser polymorphism influences the intravaginal ejaculation latency time in Dutch Caucasian men with lifelong premature ejaculation

It has been postulated that the persistent short intravaginal ejaculation latency time （IELT） of men with lifelong premature ejaculation （LPE） is related to 5-hydroxytryptamine （HT）2c receptor functioning. The aim of this study was to investigate the relationship of Cys23Ser 5-HT2c receptor gene polymorphism and the duration of IELT in men with LPE. Therefore, a prospective study was conducted in 64 Dutch Caucasian men with LPE. Baseline IELT during coitus was assessed by stopwatch over a 1-month period. All men were genotyped for Cys23Ser 5-HT2c receptor gene polymorphism. Allele frequencies and genotypes of Cys and Ser variants of 5-HT2c receptor gene polymorphism were determined. Association between Cys/Cys and Ser/Ser genotypes and the natural logarithm of the IELT in men with LPE were.investigated. As a result, the geometric mean, median and natural mean IELT were 25.2, 27.0, 33.9s, respectively. Of all men, 20.0%, 10.8%, 23.1% and 41.5% ejaculated within 10, 10-20, 20-30 and 30-60s after vaginal penetration. Of the 64 men, the Cys/Cys and Ser/Ser genotype frequency for the Cys23Ser polymorphism of the 5-HT2c receptor gene was 81% and 19%, respectively. The geometric mean IELT of the wildtypes （Cys/Cys） is significantly lower （22.6s; 95% CI 18.3-27.8s） than in male homozygous mutants （Ser/Ser） （40.4s; 95% CI 20.3-80.4s） （P = 0.03）. It is concluded that Cys23Ser 5-HT2c receptor gene polymorphism is associated with the IELT in men with LPE. Men with Cys/Cys genotype have shorter IELTs than men with Ser/Ser genotypes.     

5-羟色胺1受体对酸灌注脊髓损伤猫膀胱排尿的影响

目的 观察5-羟色胺1(5-HT1)受体激动剂对酸灌注脊髓损伤(SCI)猫膀胱排尿的影响.方法 手术彻底离断雌猫脊髓12只,术后饲养2个月,氯醛糖麻醉猫后予膀胱置管,注入0.5％乙酸溶液同时行膀胱压力测定.静脉给与5-HTlA受体激动剂8-OH-DPAT(0.30 ～30.00μg/kg)或GR-46611 (0.03 ～300.00 μg/kg),最后给予5-HT1A受体拮抗剂WAY-100635(300.00 μg/kg),记录膀胱容量阈值、膀胱容量、残尿量、排尿量和血压,同时记录尿道外括约肌肌电图(EUS-EMG).结果 8-OH-DPAT作用酸灌注脊髓损伤猫后,其膀胱容量阈值、膀胱容量、残尿量等均呈剂量依赖性增加,剂量≥10μg/kg时反应明显,差异有统计学意义.而GR-46611无类似效应.WAY-100635能逆转8-OH-DPAT的大部分效应,但对GR-46611无影响.8-OH-DPAT和GR-46611对EUS-EMG均无作用.结论 5-HTlA受体激动剂可增加生理盐水灌注及酸灌注脊髓损伤猫的膀胱容量,提示以后可能作为临床上治疗脊髓损伤患者的药物,改善慢性脊髓损伤患者的膀胱过度活动及增加脊髓损伤患者的膀胱容量.     

Bulbocavernosus reflex during the micturition cycle in normal male subjects

BACKGROUND: To investigate normal changes in the bulbocavernosus reflex (BCR) during the micturition cycle, we examined the change in BCR during the micturition cycle using an evoked potential reaction of the BCR (BCR-EP). METHODS: Fourteen normal subjects were examined in the study. The BCR-EP was recorded at empty bladder, filled bladder, during voiding and at empty bladder after voiding. To elicit the BCR-EP, the dorsal nerve of the penis was stimulated by two ring electrodes and an electromyogram of the external urethral sphincter was recorded. The maximum amplitude was measured to evaluate changes in the BCR. RESULTS: The amplitude of the BCR was increased by bladder filling and the ratio of the amplitude at filled bladder/amplitude at empty bladder was 1.32 +/- 0.39. The stable BCR-EP elicited by stimulation at empty bladder disappeared during voluntary voiding in 13 of 14 subjects. However, as stimulation was increased in seven subjects, the BCR-EP was again seen clearly. CONCLUSIONS: The BCR varies during the micturition cycle, although in normal subjects this variation occurs within a relatively narrow range. Changes in the BCR out of the normal range (e.g. large acceleration by bladder filling or insufficient inhibition during voiding) could suggest the existence of neurogenic disease.