甘丙肽家族及其受体的功能

甘丙肽家族包含甘丙肽(galanin)、甘丙肽信息相关肽(galanin-message-associated peptide,GMAP)、甘丙肽样肽(galanin-like peptide,GALP)和alarin。目前已经克隆了三种甘丙肽受体,分别是GalR1、GalR2、GalR3,它们都是G蛋白偶联受体。三种受体具有不同的分布特征,介导不同的生理过程。甘丙肽及其受体在生物体内中枢神经系统和外周神经系统中分布广泛,参与学习和记忆、焦虑行为、痛觉调节、摄食活动、渗透平衡、神经损伤修复和神经保护、胃肠道活动以及皮肤炎症处理等多种生理过程。这些生理功能提示甘丙肽及其受体可能在多种疾病的病理过程中发挥着潜在的作用,如阿尔茨海默氏病、癫痫、酗酒、糖尿病、神经性疼痛、抑郁症和癌症。     

促肾上腺皮质激素释放因子的胃肠道作用

自1981年确定促肾上腺皮质激素释放因子(CRF)的化学结构以来,已在其分布与功能的方面开展了大量工作。在胃肠道已发现CRF 免疫活性物质。由于其它下正脑肽(如促甲状腺素释放激素,生长抑素等)在胃肠道有一定作用,人们很自然地想考察一下CRF 是否也有类似作用。波兰学者S.J.Konturek 等用狗做了三组(A,B 和C)实验。A 组狗预先制备胃瘘、海氏小胃和胰瘘、     

受体活性修饰蛋白研究进展

受体活性修饰蛋白(receptor activity-modifying proteins,RAMPs)属于单跨膜蛋白家族,分三个结构域,RAMP的N端和跨膜区决定本身的功能和受体表型,胞内C端对于配体的信号传导和受体循环有重要作用。目前发现有三个成员:RAMP1、RAMP2和RAMP3。RAMPs通过改变G蛋白偶联受体的糖基化,作用于配体结合区域来调节受体表型。RAMP1与降钙素受体样受体(calcitonin receptor like receptor,CRLR)结合表现出降钙素基因相关肽(calcitonin gene-related peptide,CGRP)受体表型:RAMP2和RAMP3与CRLR结合则对肾上腺髓质素(adrenomedullin,AM)表现高亲和力,与降钙素受体(calcitonin receptor,CTR)结合则作为胰淀粉样酶(amylin,AMY)受体。由此可见,RAMPs不仅调节受体与配体结合,还影响细胞内的蛋白相互作用调节细胞内信号传导来影响细胞的增殖、迁移、分化等生物学特性。RAMPs还对心血管系统的病理生理有重要调节作用。     

肠易激综合症患者结肠组织中促肾上腺皮质激素释放因子受体表达的研究

目的 检测肠易激综合症（IBS）患者结肠组织中促肾上腺皮质激素释放因子受体（CRFR）1及CRFR2的表达,探讨其与IBS发病的关系,从而为靶向治疗IBS提供依据.方法 分别采集15例腹泻型IBS（D-IBS）患者、15例便秘型IBS（C-IBS）患者和10例正常健康人结肠组织标本,采用Elivision（TM）PLUS/HRP免疫组化染色方法和Western blot测定CRFR1、CRFR2在各组结肠组织中的蛋白表达.结果 实验结果显示,正常对照组CRFR1和CRFR2免疫组化染色以浅黄色为主,分布范围局限,平均光密度值分别为（0.254±0.099）和（0.201±0.030）;D-IBS组中CRFR1以深或棕黄色为主,分布范围较为广泛,平均光密度值为（0.384±0.048）,显著高于C-IBS组（0.144±0.077）及正常对照组（P〈0.01）;C-IBS组中CRFR2染色以深或棕黄色为主,分布范围较为广泛,平均光密度值为（0.322±0.022）,显著高于D-IBS组（0.162±0.023）（P〈0.01）及正常对照组（P〈0.05）.Western blot结果显示,D-IBS组CRFR1蛋白表达明显高于C-IBS组和正常对照组;C-IBS组CRFR2表达高于D-IBS组和正常对照组.结论 不同亚型IBS患者其CRFR表达的亚型亦不同,提示不同亚型IBS的发生可能与患者结肠组织中CRFR1、CRFR2的表达水平有关.     

神经、内分泌和免疫系统间的关系——神经免疫调节

目录一、前言二、应激与免疫三、免疫细胞上的受体(一)类固醇激素受体(二)儿茶酚胺受体(三)组织胺受体(四)肽类受体四、阿片肽与免疫功能的关系(一)免疫细胞上的阿片样和非阿片样受体1.阿片受体(1)淋巴细胞(2)单核细胞和巨噬细胞     

白细胞介素26及其受体研究进展

IL-26又称AK155属于IL-10家族成员,主要是由记忆性T细胞、NK细胞和单核细胞产生;其受体IL-26R是由IL-20R1(CRF2-8)和IL-10R2(CRF2-4)组成的异二聚体。IL-26的生物学功能还不完全清楚,初步研究表明IL-26可能在免疫调节、抗病毒感染、调节细胞增生及细胞凋亡、抗肿瘤等方面发挥作用。     

High-altitude hypoxia induces disorders of the brainendocrine-immune network through activation of corticotropin-releasing factor and its type-1 receptors

High-altitude hypoxia can induce physiological dysfunction and mountain sickness,but the underlying mechanism is not fully understood.Corticotrophin-releasing factor(CRF) and CRF type-1 receptors(CRFR1) are members of the CRF family and the essential controllers of the physiological activity of the hypothalamo-pituitary-adrenal(HPA) axis and modulators of endocrine and behavioral activity in response to various stressors.We have previously found that high-altitude hypoxia induces disorders of the brain-endocrine-immune network through activation of CRF and CRFR1 in the brain and periphery that include activation of the HPA axis in a time-and dose-dependent manner,impaired or improved learning and memory,and anxiety-like behavioral change.Meanwhile,hypoxia induces dysfunctions of the hypothalamo-pituitary-endocrine and immune systems,including suppression of growth and development,as well as inhibition of reproductive,metabolic and immune functions.In contrast,the small mammals that live on the Qinghai-Tibet Plateau alpine meadow display low responsiveness to extreme high-altitudehypoxia challenge,suggesting well-acclimatized genes and a physiological strategy that developed during evolution through interactions between the genes and environment.All the findings provide evidence for understanding the neuroendocrine mechanisms of hypoxia-induced physiological dysfunction.This review extends these findings.     

促肾上腺皮质激素释放激素及去甲肾上腺素对高原鼠兔体液免疫的抑制作用

采用第三脑室注入CRF 及N E 的方法观察对高原鼠兔(Ochotona curzoniae) 体液免疫的影响。结果表明: 第三脑室注入CRF 1 Lg 可抑制抗体生成, 比对照下降29.2%(P<0.01) , 而在第三脑室注入CRF 受体阻断剂α-helical CRF2 (9-41) 50 Lg 后再注入CRF 1 Lg 则可取消CRF 对抗体生成的抑制作用; 第三脑室注入5 nM NE, 与对照相比, 抗体水平下降38.85%(P < 0.01) , 而使用62OHDA 损毁脑内交感神经系统则使抗体水平升高24.31% (P <0.01)。这些结果表明, 高原鼠兔中枢CRF 升高对体液免疫有抑制作用, 中枢交感神经系统对体液免疫也具有紧张性抑制作用。     

逍遥散对慢性束缚应激模型大鼠相关脑区CRF基因表达的影响

目的:观察慢性束缚应激大鼠相关脑区CRF mRNA(下丘脑、垂体、海马、皮层)含量变化以及逍遥散对其影响.方法:用RT-PCR和图像分析方法测定相关脑区CRF mRNA含量变化.结果:应激组较正常对照组在下丘脑CRF-1基因表达下调(P<0.01).在下丘脑逍遥散组较应激组CRF-1基因表达显著下调(P<0.01),CRF-2基因表达显著上调(P<0.01);在海马区逍遥散组CRF-2基因表达较模型组上调(P<0.05);在皮层逍遥散组CRF-1基因表达较应激组则显著上调(P<0.01).结论:逍遥散组对慢性束缚应激中枢神经肽CRF的调节位点在下丘脑、垂体、海马和皮层,充分证实逍遥散的调节靶点与下丘脑、边缘系统及皮层中枢有关.     

应激反应中心率加快可能与促肾上腺皮质激素释放因子有关

促肾上腺皮质激素释放因子(CRF)为41肽,广泛分布于中枢神经系统,它是垂体ACTH的主要生理调节因子。一般认为它是协调、整合内分泌和应激反应的重要介质。通常血压升高通过压力反射作用可使心率减慢,而在应激情况下,往往不但动脉压升高,而且心率明显加快。这种心率加快是否与CRF有关?最近Fisher给大鼠侧脑室内注射CRF,观察了它在应激反应中对心率的影响。     

CRF receptor type 1 mediates continual hypoxia-induced CRF peptide and CRF mRNA expression increase in hypothalamic PVN of rats

We demonstrated previously that hypoxia activated CRF and CRF mRNA in PVN, and CRF receptor 1 (CRFR1) mRNA in rat pituitary. The aim of the study is to test whether the hypoxia-activated CRF and CRF mRNA is associated with triggering CRFR1. Rats were exposed to hypobaric hypoxia at altitude of 2 and 5 km. CRF and CRF mRNA were assayed by immunostaining and in situ hybridization. CRFR1 mRNA was assayed by RT-PCR. Results showed that 5 km continual hypoxia increased CRF and CRF mRNA in PVN, CRFR1 mRNA in pituitary, and plasma corticosterone. The hypoxia-increased CRF, CRF mRNA, CRFR1 mRNA, and corticosterone were blocked by CRFR1 antagonist (CP-154,526), suggesting that CRFR1 in PVN and pituitary are responsible for the hypoxia-increased CRF and CRF mRNA in PVN.     

CRF receptor antagonist astressin-B reverses and prevents alopecia in CRF over-expressing mice

Corticotropin-releasing factor (CRF) signaling pathways are involved in the stress response, and there is growing evidence supporting hair growth inhibition of murine hair follicle in vivo upon stress exposure. We investigated whether the blockade of CRF receptors influences the development of hair loss in CRF over-expressing (OE)-mice that display phenotypes of Cushing''s syndrome and chronic stress, including alopecia. The non-selective CRF receptors antagonist, astressin-B (5 µg/mouse) injected peripherally once a day for 5 days in 4–9 months old CRF-OE alopecic mice induced pigmentation and hair re-growth that was largely retained for over 4 months. In young CRF-OE mice, astressin-B prevented the development of alopecia that occurred in saline-treated mice. Histological examination indicated that alopecic CRF-OE mice had hair follicle atrophy and that astressin-B revived the hair follicle from the telogen to anagen phase. However, astressin-B did not show any effect on the elevated plasma corticosterone levels and the increased weights of adrenal glands and visceral fat in CRF-OE mice. The selective CRF₂ receptor antagonist, astressin₂-B had moderate effect on pigmentation, but not on hair re-growth. The commercial drug for alopecia, minoxidil only showed partial effect on hair re-growth. These data support the existence of a key molecular switching mechanism triggered by blocking peripheral CRF receptors with an antagonist to reset hair growth in a mouse model of alopecia associated with chronic stress.     

Immunocytochemical localization of CRF in the ovine hypothalamus

A population of neuronal cell bodies and their fiber pathways have been elucidated within the ovine hypothalamus. The immunoreactive neurons were located in the anterior and dorsal hypothalamus interspersed throughout the paraventricular nucleus. These perikarya were only observed when an antiserum that was generated against the C-terminal of CRF was employed. A dense fiber projection traversed the medial-basal hypothalamus and ended within the palisade-contact zone of the median eminence and neural stem. Fibers were revealed by antisera generated against both the N-terminal and the C-terminal of CRF. Antisera pre-absorbed with synthetic CRF failed to yield immunoreactivity.     

Peripheral CRF activates myenteric neurons in the proximal colon through CRF(1) receptor in conscious rats

Corticotropin-releasing factor (CRF) injected peripherally induces clustered spike-burst activity in the proximal colon through CRF(1) receptors in rats. We investigated the effect of intraperitoneal CRF on proximal colon ganglionic myenteric cell activity in conscious rats using Fos immunohistochemistry on the colonic longitudinal muscle/myenteric plexus whole mount preparation. In vehicle-pretreated rats, there were only a few Fos immunoreactive (IR) cells per ganglion (1.2 +/- 0.6). CRF (10 microg/kg ip) induced Fos expression in 19.6 +/- 2.1 cells/ganglion. The CRF(1)/CRF(2) antagonist astressin (33 microg/kg ip) and the selective CRF(1) antagonist CP-154,526 (20 mg/kg sc) prevented intraperitoneal CRF-induced Fos expression in the proximal colon (number of Fos-IR cells/ganglion: 2.7 +/- 1.2 and 1.0 +/- 1.0, respectively), whereas atropine (1 mg/kg sc) had no effect. Double labeling of Fos with protein gene product 9.5 revealed the neuronal identity of activated cells that were encircled by varicose fibers immunoreactive to vesicular acetylcholine transporter. Fos immunoreactivity was mainly present in choline acetyltransferase-IR nerve cell bodies but not in the NADPH-diaphorase-positive cells. These results indicate that peripheral CRF activates myenteric cholinergic neurons in the proximal colon through CRF(1) receptor.     

Disruption of the CRF/CRF1 receptor stress system exacerbates the somatic signs of opiate withdrawal

Escape from the extremely stressful opiate withdrawal syndrome may motivate opiate seeking and taking. The corticotropin-releasing factor receptor-1 (CRF1) pathway mediates behavioral and endocrine responses to stress. Here, we report that genetic inactivation (CRF1-/-) as well as pharmacological antagonism of the CRF/CRF1 receptor pathway increased and prolonged the somatic expression of opiate withdrawal. Opiate-withdrawn CRF1-/- mice also showed aberrant CRF and dynorphin expression in the paraventricular nucleus of the hypothalamus (PVN) and the striatum, indicating profound impairments in stress-responsive brain circuitry. Intake of nonstressful amounts of corticosterone effectively reduced the exaggerated somatic reactions of CRF1-/- mice to opiate withdrawal. Exogenous corticosterone also restored "wild-type-like" patterns of CRF and dynorphin gene expression in the PVN and the striatum of opiate-withdrawn CRF1-/- mice, respectively. The present findings unravel a key role for the hypothalamus-pituitary-adrenal (HPA) system and brain extra-hypothalamic CRF/CRF1 receptor circuitry in somatic, molecular, and endocrine alterations induced by opiate withdrawal.     

Corticotropin-releasing factor in humans. I. CRF stimulation in normals and CRF radioimmunoassay

The biological activity of ovine (o) and human (h) corticotropin-releasing factor (CRF) in normal volunteers was investigated, using bolus injections with different CRF dosages. There was a significant increase of ACTH, beta-endorphin and cortisol after the injection of all dosages. Repetitive stimulation and continuous infusion of hCRF lead to repetitive release of identical amounts of ACTH or constant elevation of ACTH levels. oCRF and hCRF serum immunoreactivity was measured with specific radioimmunoassays after bolus injection, pulsatile administration and infusion of CRF. The half-time of serum disappearance after acute injection studies was calculated as 9 min for hCRF dand 18 min for oCRF. The 'metabolic clearance' of hCRF calculated using the infusion study was 2.72 ml/min X kg. Endogenous CRF immunoreactivity was detectable in 14 patients during insulin hypoglycemia and in 86 out of 97 pregnant females. Furthermore, CRF could be extracted from human placenta. The chromatographic pattern of extracted placenta CRF, pregnancy serum CRF and CRF standard preparation was identical. Furthermore, CRF immunoreactivity was detectable in some patients with different causes of ACTH hypersecretion.     

Topographical distribution of CRF immunoreactivity in the pigeon brain

The distribution of corticotropin-releasing factor (CRF) immunoreactivity was demonstrated by immunocytochemistry in intact and colchicine-treated pigeons. Colchicine injections were administered at different times related to the circadian activity of the CRF-adrenocorticotropin (ACTH)-corticosterone axis. Three CRF antisera were used, two directed against synthetic rat CRF and one directed against synthetic ovine CRF. No fundamental differences appeared in the pigeon brain with respect to the specific CRF antiserum used. The most effective colchicine injection times corresponded to hypersecretion in the corticotropic axis. CRF-immunopositive neurons were scattered throughout the pigeon brain. In addition to the paraventricular hypothalamic system, which is involved in adenohypophysial ACTH regulation, several other hypothalamic and extrahypothalamic areas showed CRF neurons. The distribution suggests that CRF may also act as a modulator and a neurotransmitter. Two hypothalamic paraventricular nucleus-median eminence CRF pathways are described here. Moreover, CRF-immunopositive reactions were observed in specific areas of cerebral ventricle walls, suggesting that CRF may be released into the cerebral fluid.     

Differential profile of CRF receptor distribution in the rat stomach and duodenum assessed by newly developed CRF receptor antibodies

Peripheral corticotropin-releasing factor (CRF) receptor ligands inhibit gastric acid secretion and emptying while stimulating gastric mucosal blood flow in rats. Endogenous CRF ligands are expressed in the upper gastrointestinal (GI) tissues pointing to local expression of CRF receptors. We mapped the distribution of CRF receptor type 1 (CRF1) and 2 (CRF2) in the rat upper GI. Polyclonal antisera directed against the C-terminus of the CRF receptor protein were generated in rabbits and characterized by western blotting and immunofluorescence using CRF1- and CRF2-transfected cell lines and in primary cultured neurons from rat brain cortex. A selective anti-CRF1 antiserum (4467a-CRF1) was identified and used in parallel with another antiserum recognizing both CRF1 and CRF2 (4392a-CRF1&2) to immunostain gastric tissue sections. Antiserum 4467a-CRF1 demonstrated specific immunostaining in a narrow zone in the upper oxyntic gland within the stomach corpus. Conversely, 4392a-CRF1&2 labeled cells throughout the oxyntic gland and submucosal blood vessels. Pre-absorption with the specific antigen peptide blocked immunostaining in all experiments. Doublestaining showed co-localization of 4392a-CRF1&2 but not 4467a-CRF1 immunoreactivity with H/K-ATPase and somatostatin immunostaining in parietal and endocrine cells of the oxyntic gland. No specific staining was observed in the antrum with either antisera, whereas only antiserum 4392a-CRF1&2 showed modest immunoreactivity in the duodenal mucosa. Finally, co-localization of CRF2 and urocortin immunoreactivity was found in the gastric glands. These results indicate that both CRF receptor subtypes are expressed in the rat upper GI tissues with a distinct pattern and regional differences suggesting differential function.     

Expression and effects of metabotropic CRF1 and CRF2 receptors in rat small intestine

Corticotropin-releasing factor (CRF)-like peptides mediate their effects via two receptor subtypes, CRF1 and CRF2; these receptors have functional implication in the motility of the stomach and colon in rats. We evaluated expression and functions of CRF1 and CRF2 receptors in the rat small intestine (i.e., duodenum and ileum). CRF(1-2)-like immunoreactivity (CRF(1-2)-LI) was localized in fibers and neurons of the myenteric and submucosal ganglia. CRF(1-2)-LI was found in nerve fibers of the longitudinal and circular muscle layers, in the mucosa, and in mucosal cells. Quantitative RT-PCR showed a stronger expression of CRF2 than CRF1 in the ileum, whereas CRF1 expression was higher than CRF2 expression in the duodenum. Functional studies showed that CRF-like peptides increased duodenal phasic contractions and reduced ileal contractions. CRF1 antagonists (CP-154,526 and SSR125543Q) blocked CRF-like peptide-induced activation of duodenal motility but did not block CRF-like peptide-induced inhibition of ileal motility. In contrast, a CRF2 inhibitor (astressin2-B) blocked the effects of CRF-like peptides on ileal muscle contractions but did not influence CRF-like peptide-induced activation of duodenal motility. These results demonstrate the presence of CRF(1-2) in the intestine and demonstrate that, in vitro, CRF-like peptides stimulate the contractile activity of the duodenum through CRF1 receptor while inhibiting phasic contractions of the ileum through CRF2 receptor. These results strongly suggest that CRF-like peptides play a major role in the regulatory mechanisms that underlie the neural control of small intestinal motility through CRF receptors.     

Characterization of CRF1 receptor antagonists with differential peripheral vs central actions in CRF challenge in rats

The aim of this study was to investigate peripheral and central roles of corticotropin-releasing factor (CRF) in endocrinological and behavioral changes. Plasma adrenocorticotropin (ACTH) concentration was measured as an activity of hypothalamic-pituitary-adrenal (HPA) axis. As behavioral changes, locomotion and anxiety behavior were measured after CRF challenge intravenously (i.v.) for the peripheral administration or intracerebroventricularly (i.c.v.) for the central administration. Plasma ACTH concentration was significantly increased by both administration routes of CRF; however, hyperlocomotion and anxiety behavior were induced only by the i.c.v. administration. In the drug discovery of CRF₁ receptor antagonists, we identified two types of compounds, Compound A and Compound B, which antagonized peripheral CRF-induced HPA axis activation to the same extent, but showed different effects on the central CRF signal. These had similar in vitro CRF₁ receptor binding affinities (15 and 10 nM) and functional activities in reporter gene assay (15 and 9.5 nM). In the ex vivo binding assays using tissues of the pituitary, oral treatment with Compound A and Compound B at 10 mg/kg inhibited [¹²⁵I]-CRF binding, whereas in the assay using tissues of the frontal cortex, treatment of Compound A but not Compound B inhibited [¹²⁵I]-CRF binding, indicating that only Compound A inhibited central [¹²⁵I]-CRF binding. In the peripheral CRF challenge, increase in plasma ACTH concentration was significantly suppressed by both Compound A and Compound B. In contrast, Compound A inhibited the increase in locomotion induced by the central CRF challenge while Compound B did not. Compound A also reduced central CRF challenge-induced anxiety behavior and c-fos immunoreactivity in the cortex and the hypothalamic paraventricular nucleus. These results indicate that the central CRF signal, rather than the peripheral CRF signal would be related to anxiety and other behavioral changes, and CRF₁ receptor antagonism in the central nervous system may be critical for identifying drug candidates for anxiety and mood disorders.