Neurohypophyseal peptides in aging and Alzheimer's disease

The neurohypophyseal hormones arginine-vasopressin (AVP) and oxytocin (OT) are produced in the neurons of the hypothalamic supraoptic (SON) and paraventricular (PVN) nucleus and in the much smaller cells of the suprachiasmatic (SCN) nucleus. The SON is the main source of plasma AVP. Part of the AVP and OT neurons of the PVN join the hypothalamo-neurohypophyseal tract, whereas others send projections to the median eminence or various brain areas, where AVP and OT are involved in a number of central functions as neurotransmitters/neuromodulators. AVP and OT from the PVN can also regulate via the autonomous innervation endocrine glands and fat tissue. OT is produced for a major part in the PVN but some OT neurons are present in the SON. Moreover, both AVP and OT containing neurons are observed in the "accessory nuclei", i.e. islands situated between the SON and PVN. The SCN is the biological clock, and the number of AVP expressing neurons in the SCN shows both diurnal and seasonal rhythms. In addition to these hypothalamic areas, AVP and OT may be found to a lesser extent in some other brain areas, such as the bed nucleus of the stria terminalis, diagonal band of Broca, nucleus basalis of Meynert, lateral septal nucleus, globus pallidus and the anterior amygdaloid nucleus, as well as in the peripheral tissues. The AVP and OT containing neurons should not be considered as one system. Prominent functional differences exist between the different nuclei. The heterogeneity also becomes clear from the marked differences in the neurohypophyseal peptides containing neurons of the SON, PVN and SCN during aging, and in the most prevalent age-related neurodegenerative diseases, i.e. Alzheimer's disease (AD). For those reasons, we will discuss the SON, PVN and SCN separately.     

腹腔注射2-脱氧-D-葡萄糖可诱发大鼠视上核和室旁核神经元表达Fos

目的 探讨腹腔注射2-脱氧-D-葡萄糖（2-DG）能否激活大鼠下丘脑视上核（SON）和室旁核（PVN）神经元而表达Fos。方法 健康雄性SD大鼠12只,随机分为腹腔注射2-DG组（6只）、生理盐水对照组（3只）及正常对照组（3只）。各自处理后,应用免疫组织化学方法,观察各组下丘脑SON和PVN内Fos表达及其与催产素（OT）和加压素（VP）的双标情况,同时采用ELISA方法对血清中OT和VP的含量进行检测。 结果与生理盐水对照组和正常对照组相比,2-DG引发的特异性Fos免疫阳性产物主要集中分布于下丘脑外侧区和穹隆周区,在SON、PVN也有密集表达。SON和PVN内的Fos表达与该区的特异性神经活性物质OT和VP有共存。OT/Fos双标细胞率（双标细胞占OT阳性细胞的百分率）在SON和PVN分别为87.10%、90.57%,明显高于VP/Fos在这两个核团的双标率（双标细胞占VP阳性细胞的百分率,68.42%、76.92%）,两者比较差异有统计学意义（P<0.05）。ELISA检测结果显示,2-DG组动物血清中OT和VP水平与对照组相比无明显变化。 结论 腹腔注射2-DG可激活大鼠下丘脑SON和PVN内OT和VP神经元表达Fos,SON和PVN可能参与2-DG诱导的急性应激反应。     

Developmental changes of nitric oxide synthase expression in the rat hypothalamoneurohypophyseal system

The present study investigated the immunohistochemical localization of neuronal nitric oxide synthase (nNOS) in the hypothalamoneurohypophyseal system (HNS) of the developing rats on postnatal day 1 (PN1), 7 (PN7), 14 (PN14), 21 (PN21), and the adult rats. The nNOS-positive neurons were not discernable in the supraoptic nucleus (SON), the paraventricular nucleus (PVN), and the median eminence (ME) at PN1 and PN7. A few neurons positive for nNOS were first detected at PN14. At PN21, the nNOS-positive cells in SON and PVN rapidly increased in number. The pattern of nNOS expression at this stage approached that of the adult. Moreover, the increase of nNOS expression in the SON and PVN during the postnatal period was accompanied by the maturation of arginine vasopressin (AVP) and oxytocin (OT) neurons as indicated by the number and size of OT or AVP neurons in the SON and PVN. The patterns of AVP versus OT expression also reached that of the adult by the end of the third postnatal week. The time course of the change in nNOS expression coincided with the maturation of AVP and OT neurons in the HNS and suggested that NO synthesized by conversion of NOS is involved in the modulation of activity of neurons in the SON and PVN of the HNS.     

大鼠下丘脑弓状核神经元的神经支配

用ＨＲＰ逆行追踪和免疫细胞化学与电镜结合法研究了下丘脑弓状核神经元的神经支配。结果表明，室旁核向弓状核投射的神经元中，有一部分为催产素免疫反应阳性，一部分为后叶加压素免疫反应阳性，视上核也有部分ＯＴ免疫反应阳性神经元发出纤维投射至弓状核。将零乱毒素Ｂ亚单位结合ＨＲＰ注入第三脑室后，可见弓状核内有逆行标记细胞，电镜观察发现弓状核内的脑啡肽免疫反应阳性的树突接受ＨＲＰ反应阳性的触液神经元的轴突形成突触     

大鼠双侧海马伞离断对脂多糖诱导的下丘脑催产素能神经元功能活动的影响

为观察去海马传入对免疫应激时下丘脑室旁核、视上核催产素能神经元功能活动的影响 ,以探索海马调控外周免疫反应的脑内环路 ,本研究预先切断大鼠双侧海马伞 ,2 8d后腹腔内注射细菌内毒素脂多糖 (lipopolysaccharide,LPS,75 0μg/kg) ,用免疫组织化学双标记方法 ,以 F os蛋白作为神经元功能活动的标记物 ,观察下丘脑催产素能神经元中即刻早期基因的表达变化。结果显示 :双侧海马伞离断后 ,下丘脑室旁核中因腹腔脂多糖刺激而发生活化的催产素能神经元活化百分率显著下降 ;而视上核中催产素能神经元的活化未受明显影响。上述结果表明 ,海马对下丘脑 -垂体 -肾上腺轴以及免疫系统的抑制作用部分是通过其支配的下丘脑室旁核的催产素能神经元来完成的。     

Exercise training differentially affects intrinsic excitability of autonomic and neuroendocrine neurons in the hypothalamic paraventricular nucleus

Oxytocinergic and vasopressinergic brain stem projections have been shown to play an important role in mediating cardiovascular adjustments during exercise training (ET). The aim of the present work was to determine whether the intrinsic excitability of hypothalamic neurons giving rise to brain stem peptidergic projections is altered as a consequence of ET. Whole cell patch-clamp recordings were obtained from nucleus of the solitarii tract (NTS)-projecting paraventricular nucleus of the hypothalamus (PVN) neurons and from supraoptic nucleus (SON) and PVN magnocellular cells (MNCs), in hypothalamic slices obtained from sedentary (S) and ET rats. Our results indicate that intrinsic excitability of PVN neurons that innervate the NTS (PVN-NTS) is enhanced by ET, resulting in a more efficient input-output function (increase number of evoked actions potentials, steeper frequency/current relationships and slower decaying frequency/time relationships). Changes in input-output function were accompanied by smaller hyperpolarizing afterpotentials (HAPs) and afterhyperpolarizing potentials (AHPs), during and after trains of spikes, respectively. On the other hand, a decreased efficacy in the input-output function was observed in SON/PVN MNCs during ET. Altogether, our results indicate that ET differentially affects the intrinsic excitability of autonomic and neurosecretory SON and PVN neurons. Increased excitability in PVN-NTS neurons may contribute to enhanced release of OT and VP peptides in the dorsal brain stem, and cardiovascular fine-tuning during exercise training.     

不同禁水时间老龄大鼠下丘脑大细胞分泌神经元的免疫电镜观察

电镜观察了不同禁水时间老龄大鼠下丘脑视上核和室旁核分泌神经元，即加压素和催产素神经元及胶质细胞的超微结构变化，结果显示，禁水6与12h后，上述两核团中的分泌神经元胞体增大，胞质中粗面内质网（RER）排列紧密且规则，高尔基（Golgi)器的未成熟分泌颗粒及神经分泌颗粒增多，轴突内神经分泌颗粒少见，胶质细胞成份减少，突起回缩；相邻两神经元胞膜直接接触，质膜并列现象及突触增多，而在禁水24h后，神经元胞体内的神经分泌颗粒有减少，轴突中的神经分泌颗粒却增多并聚集成膨大区域，以上结果提示老龄大鼠视上核和室旁核的分泌神经元在禁水时其合成激素的功能是活跃的，而且催产素神经元的结构变化与加压素神经元结构变化是相似的，胶质细胞的结构变化为神经元质膜并列及突触的形式提供了有利条件。     

Synergistic activation of astrocytes by ATP and norepinephrine in the rat supraoptic nucleus

Supraoptic nucleus (SON) neurons receive a dense innervation from noradrenergic fibers, the activity of which stimulates vasopressin (VP) and oxytocin (OT) release, notably during homeostatic regulation of blood pressure and volume. This regulation is known to involve the co-release of norepinephrine (NE) and ATP, which act in synergy to stimulate Ca(2+) increase in SON neurons and to enhance release of VP and OT from hypothalamo-neurohypophysial explants. We here demonstrate that both ATP and NE also trigger transient intracellular Ca(2+) rise in rat SON astrocytes, the two agonists showing a synergistic action similarly to what has been reported in SON neurons. The responses to both agonists are not or are only moderately affected after blockade of neuronal activity by tetrodotoxin, or of neurotransmitter release by removal of extracellular Ca(2+), suggesting that the receptors involved are located on the astrocytes themselves. ATP acts via P2Y(1) receptors, as indicated by the pharmacological profile of Ca(2+) responses and the strong immunolabeling for this receptor in SON astrocytes. Responses to NE involve both alpha and beta adrenergic receptors, the latter showing a permissive role on the former. These results point to further implication of SON astrocytes in the regulation of VP and OT secretion, and suggest that they are potentially important elements participating in all regulatory processes of hypothalamo-neurohypophysial function that involve activation of noradrenergic pathways.     

大鼠下丘脑催产素及加压素样神经元与脑啡肽样传入纤维末梢的关系

用免疫组织化学ABC双标记法,显示大鼠下丘脑各大细胞神经分泌核团中催产素样免疫反应(OT-li)、加压素样免疫反应(VP-li)神经元、亮-脑啡肽样免疫反应(L-ENK-li)和甲-脑啡肽样免疫反应(M-ENK-li)纤维末梢,并对不同神经分泌核团内脑啡肽样传入纤维末梢与OT-li及VP-li神经元的关系加以分析。结果发现,L-ENK-li及M-ENK-li纤维末梢在下丘脑各神经分泌核团中,与OT-li及VP-li神经元均有一定程度的接触关系。L-ENK-li与M-ENK-li纤维末梢在第三脑室周、前连合核、背内侧和背外侧副核、穹窿前和后核的OT-li及VP-li神经元周围最密集,在视上核、室旁核次之,在血管周细胞群内的OT-li及VP-li神经元周围密度较低。这些神经分泌性核团内L-ENK-li纤维末梢,均较M-ENK-li纤维末梢密集。表明脑啡肽能传入纤维末梢,与视上核、室旁核及各神经分泌副核(除血管周细胞群外)中的OT-li及VP-li神经元,均有不同程度的接触关系。因此,脑啡肽能传入纤维末梢可能在大细胞神经分泌系统OT-li及VP-li神经元分泌活动的调节中,起较广泛的重要作用。     

Oxytocin,but not vassopressin,modulates nociceptive responses in dorsal horn neurons

Oxytocin (OT) and vasopressin (VP) are synthesized and secreted by the paraventricular hypothalamic nucleus (PVN), and both peptides have been implicated in the pain modulatory system. In the spinal cord, activation of OT-containing axons modulates nociceptive neuronal responses in dorsal horn neurons; however, it is not known whether the direct VPergic descending projection participates. Here, we show that both PVN electrical stimulation and topical application of OT in the vicinity of identified and recorded dorsal horn WDR selectively inhibit Aδ and C-fiber responses. In contrast, the topical administration of VP on the same neurons did not affect the nociceptive responses. In addition, the reduction in nociceptive responses caused by PVN stimulation or OT administration was blocked with a selective OT antagonist. The results suggest that the VP descending projection does not modulate the antinociceptive effects mediated by the PVN on dorsal horn neurons; instead, it is the hypothalamic-spinal OT projection that regulates nociceptive information.     

Effects of hyperosmotic stimulation and adrenalectomy on vasopressin mRNA levels in the paraventricular and supraoptic nuclei of the hypothalamus: in situ hybridization histochemical analysis using a synthetic oligonucleotide probe

The effects of salt loading and adrenalectomy on arginine vasopressin (AVP) mRNA levels in the paraventricular nucleus (PVN) and the supraoptic nucleus (SON) of the hypothalamus were studied by semiquantitative in situ hybridization histochemistry, using a synthetic oligonucleotide probe and a computer-assisted image analysis system. Salt loading (2% NaCl) for 7 days produced marked increases in AVP mRNA levels in the magnocellular neurons of the PVN, SON, and accessory nuclei. Adrenalectomy caused an increase in AVP mRNA expression in the magnocellular part of the PVN and the expansion of hybridization signals into its medial parvocellular region, where the cell bodies of corticotropin-releasing hormone (CRH) neurons are located. No apparent alteration of AVP mRNA levels was observed in the SON following adrenalectomy. These results indicate that hyperosmotic stimulation and the loss of circulating glucocorticoids had differential effects on AVP gene expression in the PVN and SON, and that the magnocellular PVN and SON neurons responded in different manners to the loss of feedback signals.     

Plastisity of secreteory neurons of hypothalamic supraoptic and paraventricular nuclei during light exposure

Changes of secreteory neurons of hypothalamic supraoptic (SON) and paraventricular nuclei (PVN) were analyzed using light and electron microscopy in 100 adult male rats from the first minutes up to 180 days after their round-the-clock 48 hours-long exposure to bright light. At the early stages after the exposure, SON and PVN cellular formula was changed in favor of functionally active neurons containing few secretory granules, large nucleus and nucleoli, low RNA content, sparse complexes of rough endoplasmic reticulum, vacuoles and lysosomes in their perikarya. After 24 hours in both SON and PVN and after 10 days in SON the number of cells accumulating secretion was greater than that in control group. Normalization of the cellular formula and of the structural organization of protein-synthesis apparatus in PVN secretory neurons took place at days 10-30, while those ones in SON--at days 30-180. The results obtained suggest that the range of plasticity in PVN neurons in animals exposed to continuous bright light is larger than that in SON neurons.     

后叶加压素在下丘脑的分布和释放途径及失水对正中隆起中后叶加压素的影响

我们用免疫细胞化学方法(ICC)对大白鼠下丘脑中加压素(VP)免疫反应阳性的神经元作了较详细研究。观察到VP阳性神经元存在于下丘脑室旁核(PVN)各亚核、视上核(SON)、交叉上核(SCN)、室周核(PN)及一些附属核团,包括环状核(CN)、交叉后核(RCN)、下丘脑外侧核(HLN)、穹窿周围核(PFN)。且发现一特殊的VP阳性胞体聚集区—很可能是多巴胺神经元聚集区A_(14)中的细胞,位于第三脑室侧壁中1/3段两侧,腹内侧核背侧。本文首次观察到在PVN和SON之间有VP阳性的神经纤维相联系。ICC和免疫电镜研究进一步证明在正中隆起外带存在VP阳性纤维,含大颗粒囊泡的VP末梢紧邻门脉毛细血管。将HRP注入第四脑室用HRP逆行追踪与ICC方法相结合,在PVN中观察到双标细胞,与直接用ICC法所见VP阳性轴突伸入第三脑室腔的结果一致。说明PVN中存在接触脑脊液神经元。干渴动物正中隆起的内、外带中的VP免疫反应明显减弱。     

Differential Activation of c‐fos Immunoreactivity After Hypophysectomy in Developing and Adult Rats

The aim of the present study is to compare c‐fos expression in identified hypothalamic vasopressin (AVP) and oxytocin (OT) neurons in developing (PN7 and PN14) and adult rats following hypophysectomy using dual‐labeled immunostaining. Our results showed that hypophysectomy induced c‐fos expression in supraoptic (SON) and paraventricular (PVN) nuclei in both the developing and adult rats. Few or no positive cells were observed in the same nuclei in sham‐operated animals. Quantitative analysis for c‐fos and either of the above named neuropeptides revealed that almost all AVP and OT neurons in the adult and PN14 groups expressed c‐fos in response to hypophysectomy. In PN7, hypophysectomy also induced all AVP neurons to express c‐fos in SON and PVN. However, few OT neurons in the SON and PVN produced c‐fos after hypophysectomy. In addition, the time course of c‐fos expression was different in the developing and adult rats after hypophysectomy. The c‐fos expression in the developing rats exhibited a more prolonged induction in which staining for c‐fos persisted for at least 3 days after hypophysectomy compared with that in the adult in which c‐fos immunoreactivity disappeared within 24 hr post‐lesion. This study demonstrates that c‐fos expression after hypophysectomy is regulated differently during development. Anat Rec, 2007. © 2007 Wiley‐Liss, Inc.     

胃电刺激对大鼠VMH胃扩张敏感神经元电活动及脑内催产素表达的影响

目的： 给予胃窦部以2组不同参数的电刺激，观察大鼠下丘脑腹内侧核（VMH）胃扩张（GD）敏感性神经元放电频率的变化及脑内有促进摄食作用的神经肽-催产素(OT) 表达的变化，为胃电刺激（GES）治疗肥胖的中枢作用机制及临床上治疗肥胖参数的选择提供理论依据。方法: ① 电生理实验：采用细胞外记录神经元单位放电方法，记录下丘脑腹内侧核神经元自发放电活动，根据神经元对胃扩张刺激反应的不同，分为胃扩张兴奋性神经元（GD-E）和胃扩张抑制性神经元（GD-I）， 并观察不同参数电刺激胃窦部，VMH内GD-E和GD-I放电频率的变化。②免疫组化实验：采用免疫荧光组织化学染色方法观察胃电刺激 2 h 对大鼠脑内OT阳性神经元表达的影响。结果: ① 电生理结果：GES1和GES2分别使60.4%和75.0%的GD敏感性神经元兴奋（P>0.05）。GES2和GES1分别可使GD-E神经元的放电频率平均增加343.59%±89.19%和97.44±33.67% (P<0.05)，GD-I神经元的放电频率平均增加366.30%±87.20%和112.00%±14.67% (P<0.05)。②免疫组化结果：GES1刺激胃窦部 2 h，室旁核（PVN）和视上核（SON）OT 免疫阳性神经元明显增加（P<0.05）。结论: GES可通过兴奋“饱中枢”-VMH内胃扩张敏感性神经元和增加脑内OT的表达来抑制摄食，且GES的作用效应与其强度有关。     

加压素能和宫缩素能神经元在新生儿下丘脑的分布

本文用ABC法对加压素样和宫缩素样神经元在新生儿下丘脑的分布进行了观察。结果发现,两者的神经元集中分布于视上核、室旁核及副大细胞分泌核。根据两者的分布特点,又将视上核、室旁核和副大细胞分泌核再分为若干分区,各分区均以加压素样神经元为主,约占总数的80～90％。本文结果提示加压素样和宫缩素样神经元在下丘脑的分布具有动物种属和年龄特征。     

Chronic morphine treatment inhibits oxytocin release from the supraoptic nucleus slices of rats

Effect of chronic morphine treatment on oxytocin (OT) release from the long term-cultured organotypic slice of the supraoptic nucleus (SON) was investigated using radioimmunoassay. The co-localization of oxytocin and mu-opioid receptor in neurons within the SON was observed with the double-labeled methods of in situ hybridization combined with immunohistochemistry. After exposure to morphine for 6days, the OT levels in culture media were significantly decreased. Naloxone caused much greater release of OT in chronic morphine treatment group than in controls. Naloxone has no effect after acute morphine treatment. 90% of OT-ir (immunoreactive) neurons expressed mu-opioid receptor mRNA in the SON and 45% of the neurons that expressed mu-opioid receptor mRNAs were OT-ir neurons. These results indicated that the neurons within SON could develop dependence on morphine in vitro, and these effects might be exerted via mu-opioid receptor in oxytocin neurons of the SON.     

Mechanisms underlying oxytocin-induced excitation of supraoptic neurons: prostaglandin mediation of actin polymerization

In nonneuronal tissues, activation of oxytocin receptors (OTRs), like other Galpha(q/11) type G-protein-coupled receptors (Galpha(q/11)/GPCRs), increase prostaglandin (PG) expression. This is not known for the OTRs expressed by central OT neurons. We examined mechanisms underlying OT's effects on supraoptic nucleus (SON) OT and vasopressin (VP) neurons in hypothalamic slices from lactating rats. OT application (10 pM, 10 min) significantly increased firing rates of OT and VP neurons, both of which expressed OTRs. Indomethacin, an inhibitor of PG synthetases, blocked these increases. OTR (but not a V1 receptor) antagonist blocked OT effects without blocking the excitatory effect of PGE2. Tetanus toxin blocked OT effects on fast synaptic inputs and firing activity of SON neurons but not OT-evoked depolarization, suggesting involvement of both pre- and postsynaptic neurons. Indomethacin also blocked the excitatory effects of phenylephrine, another Galpha(q/11)/GPCR activating agent but not those of PGE2, a non-Galpha(q/11)/GPCR activating agent in the SON. OT or phenylephrine, but not glutamate or KCl, enhanced cyclooxygenase 2 expression at cytosolic loci in SON neurons and nearby astrocytes, as revealed by immunocytochemistry. This OT effect was not blocked by TTX. Western blot analyses showed that OT significantly increased cyclooxygenase 2 but not actin expression. OT promoted the formation of filamentous actin (F-actin) networks at membrane subcortical areas of both OT and VP neurons. Indomethacin blocked enhancement of F-actin networks by OT but not by PGE2. These results indicate that PGs serve as a common mediator of Galpha(q/11)/GPCR-activating agents in neuronal function.     

The role of catecholaminergic neurons in the hypothalamus and medullary visceral zone in response to restraint water-immersion stress in rats

The activity of catecholaminergic neurons in the hypothalamus and the medullary visceral zone (MVZ) in rats in response to restraint water-immersion stress (RWIS) was measured by use of dual Fos and tyrosine hydroxylase (TH) immunohistochemistry. In RWIS rats Fos immunoreactive (Fos-IR) nuclei dramatically increased in the paraventricular nucleus (PVN), the supraoptic nucleus (SON), the dorsal motor nucleus of the vagus (DMV), the nucleus of the solitary tract (NTS), the area postrema (AP), and the ventrolateral medulla (VLM). A small number of TH-immunoreactive (TH-IR) and Fos/TH double-labeling neurons in the PVN, and their absence from the SON, were observed in both RWIS and nonstressed rats. More TH-IR neurons were observed in the MVZ of RWIS rats than in nonstressed rats. In RWIS and nonstressed rats, the percentage of Fos-IR nuclei in TH-IR neurons was 38.0 and 14.3% in the DMV, 34.4 and 9.7% in the NTS, 18.6 and 4.5% in the AP, and 45.7 and 18.9% in the VLM, respectively. In conclusion, catecholaminergic neurons in the MVZ are involved in the response to RWIS; although the PVN and SON also participate in the response to RWIS, the mechanism is not via catecholaminergic neurons.