外源性褪黑素对慢性应激抑郁模型大鼠行为学及血清相关激素含量的影响

目的通过观察不同剂量的外源性褪黑素对慢性应激抑郁模型大鼠行为学及血清中5-HT、CRH、ACTH、CORT含量的影响,探讨外源性褪黑素对抑郁症的治疗作用。方法 48只SD大鼠随机分为4组:空白对照组、模型对照组、褪黑素高剂量组、褪黑素低剂量组,每组12只。采用慢性轻度不可预见性应激和孤养方法复制慢性应激抑郁模型,灌胃给予治疗组不同剂量(5.0、10.0 mg·kg-1)的褪黑素加以干预,以体重、糖水试验、旷野试验观察实验前后大鼠行为学的改变。应用酶联免疫吸附法对大鼠血清中5-HT、CRH、ACTH、CORT的含量进行检测。结果从第7天开始,模型组、褪黑素低量组、褪黑素高剂量组的体重增长趋势开始降低,从第14天开始,模型对照组大鼠体重有下降趋势;从第14天开始褪黑素低剂量组、褪黑素高剂量组的糖水摄入量增长趋势降低,模型对照组的糖水摄入量呈下降趋势;第21天,与模型对照组相比,褪黑素低剂量和褪黑素高剂量组大鼠的水平和垂直运动次数都有显著性增多。与空白对照组相比,模型对照组血清5-HT的含量降低、血清CRH、ACTH、CORT含量升高,与模型对照组相比,褪黑素低剂量组和褪黑素高剂量组的血清5-HT含量均显著升高,血清CRH、CORT、ACTH含量均显著降低。结论外源性褪黑素对慢性应激抑郁模型大鼠抑郁行为有改善作用,也可以提高慢性应激抑郁模型大鼠血清中5-HT的含量,降低慢性应激抑郁模型大鼠血清中CRH、ACTH、CORT的含量,但它们之间的具体量效关系有待进一步研究。     

Role of the basolateral amygdala in mediating the effects of the fatty acid amide hydrolase inhibitor URB597 on HPA axis response to stress

The endocannabinoid system is an important regulator of neuroendocrine and behavioral adaptation in stress related disorders thus representing a novel potential therapeutic target. The aim of this study was to determine the effects of the fatty acid amide hydrolase (FAAH) inhibitor URB597 on stress mediators of HPA axis and to study the role of the basolateral amygdala (BLA) in responses to forced swim stress.Systemic administration of URB597 (0.1 and 0.3 mg/kg) reduced the forced swim stress-induced activation of HPA axis. More specifically, URB597 decreased stress-induced corticotropin-releasing hormone (CRH) mRNA expression in the paraventricular nucleus (PVN) of the hypothalamus, and pro-opiomelanocortin (POMC) mRNA expression dose-dependently in pituitary gland without affecting plasma corticosterone levels. URB597 treatment also attenuated stress-induced neuronal activation of the amygdala and PVN, and increased neuronal activation in the locus coeruleus (LC) and nucleus of solitary tract (NTS). Injection of the CB1 receptor antagonist AM251 (1 ng/side) in the BLA significantly attenuated URB597-mediated effects in the PVN and completely blocked those induced in the BLA.These results suggest that the BLA is a key structure involved in the anti-stress effects of URB597, and support the evidence that enhancement of endogenous cannabinoid signaling by inhibiting FAAH represents a potential therapeutic strategy for the management of stress-related disorders.     

Characterization of a novel subtype of hippocampal interneurons that express corticotropin‐releasing hormone

A subset of corticotropin‐releasing hormone (CRH) neurons was previously identified in the hippocampus with unknown function. Here we demonstrate that hippocampal CRH neurons represent a novel subtype of interneurons in the hippocampus, exhibiting unique morphology, electrophysiological properties, molecular markers, and connectivity. This subset of hippocampal CRH neurons in the mouse reside in the CA1 pyramidal cell layer and tract tracing studies using AAV‐Flex‐ChR2‐tdTomato reveal dense back‐projections of these neurons onto principal neurons in the CA3 region of the hippocampus. These hippocampal CRH neurons express both GABA and GAD67 and using in vitro optogenetic techniques, we demonstrate that these neurons make functional connections and release GABA onto CA3 principal neurons. The location, morphology, and importantly the functional connectivity of these neurons demonstrate that hippocampal CRH neurons represent a unique subtype of hippocampal interneurons. The connectivity of these neurons has significant implications for hippocampal function. © 2015 Wiley Periodicals, Inc.     

Adrenal function in Smith-Lemli-Opitz syndrome

Smith-Lemli-Opitz syndrome (SLOS) is a multiple malformation syndrome due to mutations of the 7-dehydrocholesterol reductase gene (DHCR7), which leads to a deficiency of cholesterol synthesis and an accumulation of 7-dehydrocholesterol. The SLOS clinical spectrum ranges from multiple major malformations to a mild phenotype with minor anomalies and intellectual disability. Several children with SLOS and adrenal insufficiency have been described. We performed ovine corticotropin (oCRH) testing in 35 SLOS patients and 16 age- and gender-matched controls. We reviewed prior adrenocorticotropin (ACTH) stimulation tests of our SLOS patients (19 of 35 available) and reviewed results of ACTH stimulation tests from 10 additional SLOS patients. Results from oCRH testing showed that patients with SLOS had significantly higher ACTH baseline values than healthy controls (24.8 ± 15.3 pg/ml vs. 17.8 ± 7.5 pg/ml, P = 0.034). However, no statistically significant differences were noted for peak ACTH values (74.4 ± 35.0 pg/ml vs. 64.0 ± 24.9 pg/ml, P = 0.303) and for baseline (14.2 ± 7.8 mcg/dl vs. 14.2 ± 6.3 mcg/dl, P = 0.992) and peak cortisol values (28.2 ± 7.9 mcg/dl vs. 24.8 ± 8.1 mcg/dl, P = 0.156). The area-under-the-curve (AUC) was not significantly different in SLOS patients compared to controls for both ACTH (250.1 ± 118.7 pg/ml vs. 195.3 ± 96.6 pg/ml, P = 0.121) as well as cortisol secretion (83.1 ± 26.1 mcg/dl vs. 77.8 ± 25.9 mcg/dl, P = 0.499). ACTH stimulation test results were normal in 28 of 29 tests. The individual with the abnormal test results had subsequent normal oCRH tests. The slightly increased baseline ACTH level seen during oCRH testing may be due to compensated adrenocortical insufficiency. However, we were able to show that our patients with SLOS had an adequate glucocorticoid response, and thus, in mild to moderate cases of SLOS stress steroid coverage may not be warranted.     

大鼠胸部严重创伤后早期下丘脑CRH表达的实验研究

目的 探讨大鼠胸部撞击伤应激反应中,下丘脑表达和分泌促肾上腺皮质激素释放激素(CRH)的变化规律及其反馈调节机制。方法 应用BIM-Ⅲ型多功能小型生物撞击机致伤动物,采用免疫组化和RT-PCR等技术,检测大鼠下丘脑CRH的分布及含量变化规律。分组：对照组、撞击伤后15、30、60、120、360、720分钟组、撞击伤处理组(用CRH1R特异性阻滞剂CP-154526处理)。结果 正常情况下,CRH少量分布于下丘脑室旁核和视上核;撞击伤后,室旁核和视上核的CRH染色明显增加;撞击伤后60分钟,下丘脑CRH mRNA含量明显增加,且呈逐渐增加的趋势,720分钟开始回落;用CP-154526处理的大鼠,撞击伤后,下丘脑CRH含量的增加明显减少。结论 在创伤性应激反应早期,CRH通过与下丘脑神经元CRH1R的结合,选择性地调节下丘脑神经元CRH基因的表达。     

大鼠腹腔巨噬细胞表达促肾上腺皮质激素释放激素的实验研究

目的 观察促肾上腺皮质激素释放激素(CRH)在大鼠腹腔巨噬细胞的表达变化规律.方法 体外培养大鼠腹腔巨噬细胞,采用RT-PCR及ELISA技术检测正常及脂多糖(LPS)、佛波脂(PMA)刺激后,细胞CRH mRNA转录水平及CRH多肽合成水平.结果 RT-PCR结果显示,正常大鼠腹腔巨噬细胞内有一定水平的CRH mRNA,LPS及PMA刺激后,细胞内CRH mRNA水平均显著增高(P＜0.01).在多肽水平,正常大鼠腹腔巨噬细胞可表达低水平的CRH[约(0.07±0.01)ng/107cells],LPS及PMA促进了CRH多肽的合成(P＜0.01).结论 正常大鼠腹腔巨噬细胞可合成表达CRH,其水平在活化后大鼠腹腔巨噬细胞显著上调.     

柴胡疏肝散及其提取物对慢性应激大鼠下丘脑CRH含量的影响

目的探讨柴胡疏肝散及其提取物对大鼠下丘脑促肾上腺皮质激素释放激素(CRH)含量的影响。方法采用慢性多相性应激模型,放免方法检测下丘脑CRH含量。结果与正常组相比,模型组下丘脑CRH含量显著升高;而水提醇沉液和原方高剂量组可下调下丘脑CRH含量。结论柴胡疏肝散、水提醇沉液可能通过调节丘脑CRH含量调节HPA起抗抑郁作用。     

强肌健力饮对肾阳虚大鼠CRH、ACTH、Cor水平的影响

目的:观察强肌健力饮对肾阳虚大鼠下丘脑组织中促肾上腺皮质激素释放激素(CRH)及血浆促肾上腺皮质激素(ACTH)、皮质醇(Cor)水平的影响,进一步探讨该方对中医肾阳虚证的防治机理.方法:将大鼠分为正常对照组、肾阳虚模型组、强肌健力饮低、中、高剂量组、右归丸阳性对照组,采用氢化可的松制备肾阳虚大鼠模型.观察动物的一般状态及其胸腺和肾上腺指数,采用放射免疫分析检测CRH、ACTH、Cor的含量.结果:①肾阳虚模型组大鼠体重及胸腺指数、肾上腺指数明显低于正常对照组(P＜0.01),CRH、ACTH、Cor含量均比正常对照组显著降低(P＜0.01).②强肌健力饮各剂量组CRH、ACTH、Cor含量均比肾阳虚模型组显著升高(P＜0.05～0.01).结论:肾阳虚时下丘脑-垂体-肾上腺轴合成、分泌和调控功能低下,而强肌健力饮能够修复该轴功能的损伤,表明该方药具有下丘脑、垂体、肾上腺轴多层次的调节作用.     

Increased transvascular transport of WGA-peroxidase after chronic perinatal stress in the hippocampal microvasculature of the rat

Brain endothelial ultrastructural properties contribute to maintain proper blood-brain barrier (BBB) function. Several physiological and pathological conditions have been shown to alter BBB permeability to blood-borne molecules, acute and chronic stress among them. In the rat, early life stress increased transvascular transport of Evans blue, however, the route of tracer extravasation is not fully known; therefore the aim of the present experiment was to describe the ultrastructural changes in endothelial cells subsequent to chronic perinatal stress in order to ascertain the route for transvascular transport of an electrodense tracer. Pregnant Wistar rats and their litters were used. Four pregnant rats were subjected to forced swimming between gestational days 10 to 20. After delivery, half of the control litters underwent 180 min maternal separation from postnatal day 2 to 20. Controls were kept free of any stress manipulation. At sacrifice between postnatal days 1 to 30 subjects were given intracardially the lectin wheat germ agglutinin conjugated to horseradish peroxidase (WGA-HRP). WGA-HRP stained hippocampi were processed for ultrastructural analysis, transmission electron micrographs were obtained and endothelial ultrastructural parameters quantified using the ImageJ software. Both stress procedures accelerated gross microvessel development by decreasing capillary wall thickness and endothelial microvilli. However, early-life stress also neutralized endothelial glycocalyx, increased vesicle-mediated transport and tended to promote the formation of secondary lysosomes containing endocytosed WGA-HRP vesicles, all parameters of altered endothelial cell function. Tight junction development in both stress groups was similar to the control pups.     

Overexpression of corticotropin-releasing hormone in transgenic mice and chronic stress-like autonomic and physiological alterations

To gain a greater insight into the relationship between hyperactivity of the corticotropin-releasing hormone (CRH) system and autonomic and physiological changes associated with chronic stress, we developed a transgenic mouse model of central CRH overproduction. The extent of central and peripheral CRH overexpression, and the amount of bioactive CRH in the hypothalamus were determined in two lines of CRH-overexpressing (CRH-OE) mice. Furthermore, 24 h patterns of body temperature, heart rate, and activity were assessed using radiotelemetry, as well as cumulative water and food consumption and body weight gain over a 7-day period. CRH-OE mice showed increased amounts of CRH peptide and mRNA only in the central nervous system. Despite the presence of the same CRH transgene in their genome, only in one of the two established lines of CRH-OE mice (line 2122, but not 2123) was overexpression of CRH associated with increased levels of bioactive CRH in the hypothalamus, increased body temperature and heart rate (predominantly during the light (inactive) phase of the diurnal cycle), decreased heart rate variability during the dark (active) phase, and increased food and water consumption, when compared with littermate wildtype mice. Because line 2122 of the CRH transgenic mice showed chronic stress-like neuroendocrine and autonomic changes, these mice appear to represent a valid animal model for chronic stress and might be valuable in the research on the consequences of CRH excess in situations of chronic stress.