The role of nuclear factor-κB in the effect of angiotensin II in the paraventricular nucleus in protecting the gastric mucosa from ischemia-reperfusion injury in rats

BACKGROUND: The purpose of this study was to elucidate the role of nuclear factor kappaB (NF-kappaB) in the development of gastric ischemia-reperfusion (GI-R) injury and in mediating the effects of angiotensin II (Ang II) in the paraventricular nucleus (PVN) on GI-R injury. METHODS: GI-R injury was induced in rats by clamping the celiac artery for 30 min and then reperfusing for 1 h. A cannula was inserted into the unilateral PVN for microinjection of Ang II. The expressions and levels of NF-kappaB (p65), IkappaB-alpha, and phosphorylated IkappaB-alpha in rat gastric mucosa were examined by Western blotting and immunohistochemistry. A laser Doppler flowmeter was used to assess gastric blood flow (GBF). Malondialdehyde (MDA) was determined using the thiobarbituric acid (TBA) method, and superoxide dismutase (SOD) activity was determined by the xanthine/xanthine oxidase method. RESULTS: Microinjection of Ang II (3, 30, and 300 ng) into the PVN dose-dependently inhibited GI-R injury. The levels and expressions of NF-kappaB (p65) and phosphospecific IkappaB-alpha protein increased 1 h after GI-R and were markedly reduced by microinjection of Ang II into the PVN. In contrast, the level and expression of IkappaB-alpha protein decreased 1 h after ischemia-reperfusion and recovered to the normal level by microinjection of Ang II into the PVN. The effects of Ang II were prevented by pretreatment with the Ang II AT1 receptor antagonist losartan (5 microg) microinjected into the lateral cerebral ventricle. Inhibition of NF-kappaB activity by pyrrolidine dithiocarbamate (PDTC, 200 mg/kg) produced similar effects in rats subjected to ischemia-reperfusion with or without microinjection of Ang II into the PVN. Administration of PDTC attenuated gastric mucosal injury and suppressed the activation of NF-kappaB (p65). Ang II microinjection into the PVN increased GBF and decreased the MDA content but did not alter SOD activity in the gastric mucosa following ischemia-reperfusion. CONCLUSIONS: NF-kappaB plays a role in PVN Ang II-mediated protection against GI-R injury. These central effects of Ang II are mediated by AT1 receptors.     

Intrauterine position affects estrogen receptor α expression in the ventromedial nucleus of the hypothalamus via promoter DNA methylation

There is well-established evidence in many mammalian species for effects of the intrauterine position (IUP) (the sex-specific positioning of the embryo) on postnatal brain function and behavior. We found that the IUP affects estrogen receptor (ER)α expression in adult female rats in the ventrolateral region of the hypothalamic ventromedial nucleus (vlVMH), which is associated with sexual behavior. The ERα expression level in the vlVMH was higher in females that developed in utero between two male siblings (2M females) than in those that developed between female siblings (2F females). We also found that the cytosine methylation status across the ERα promoter in the vlVMH was affected by the IUP, with greater methylation in 2F females. These findings showed a negative correlation between ERα expression levels in the vlVMH and methylation frequency in the ERα promoter. This suggests that genomic methylation sustains the effect of the fetal IUP on ERα expression in the vlVMH.     

Reversal of morphine-induced cell-type–specific synaptic plasticity in the nucleus accumbens shell blocks reinstatement

Drug-evoked plasticity at excitatory synapses on medium spiny neurons (MSNs) of the nucleus accumbens (NAc) drives behavioral adaptations in addiction. MSNs expressing dopamine D1 (D1R-MSN) vs. D2 receptors (D2R-MSN) can exert antagonistic effects in drug-related behaviors, and display distinct alterations in glutamate signaling following repeated exposure to psychostimulants; however, little is known of cell-type–specific plasticity induced by opiates. Here, we find that repeated morphine potentiates excitatory transmission and increases GluA2-lacking AMPA receptor expression in D1R-MSNs, while reducing signaling in D2-MSNs following 10–14 d of forced abstinence. In vivo reversal of this pathophysiology with optogenetic stimulation of infralimbic cortex-accumbens shell (ILC-NAc shell) inputs or treatment with the antibiotic, ceftriaxone, blocked reinstatement of morphine-evoked conditioned place preference. These findings confirm the presence of overlapping and distinct plasticity produced by classes of abused drugs within subpopulations of MSNs that may provide targetable molecular mechanisms for future pharmacotherapies.Opioid-based drugs are mainstays for pain management (1). However, side effects such as euphoria and the development of tolerance and dependence contribute to an increasing diversion of these readily available compounds for nontherapeutic use (2). Opioid agonist-based treatments are known to reduce some aspects of opioid addiction. On the other hand, these therapies often lead to high relapse rates when discontinued because they fail to eliminate key aspects of addiction such as conditioned associations that can trigger intense drug craving (2). Currently, development of alternative treatments for opioid addiction is hampered by a distinct lack of knowledge of the cellular plasticity that underlies persistent opioid-induced changes in behavior.The nucleus accumbens (NAc) region of the ventral striatum is involved in attribution of salience to drug-paired cues that can in turn motivate reward-related behavior (3, 4). Medium spiny neurons (MSNs), the principal cells of the NAc, are GABAergic projection neurons that receive coordinated glutamatergic afferents arising from several cortical and limbic brain regions (5, 6). MSNs are divided into two subpopulations based on expression of the dopamine receptor 1 (D1R-MSN) or dopamine receptor 2 (D2-MSN), with a small fraction (∼6–17%) expressing both receptors (7). Importantly, these subpopulations have divergent projection targets and exert antagonistic effects in reward-related behaviors (8).Long-lasting alterations in excitatory synaptic strength and glutamate release at MSNs produced by repeated exposure to drugs of abuse is a driving factor behind drug seeking and relapse (9–11). Numerous studies have examined effects of repeated psychostimulant exposure on synaptic strength and AMPA receptor (AMPAR)-mediated transmission in MSN subpopulations, with a majority of adaptations observed in D1R-MSN (12–14). Although opiate-induced changes in extracellular glutamate and glutamate receptor subunit expression have been reported, very little is known about opiate-induced changes in the efficacy of synaptic strength and function in the NAc, the degree to which this plasticity is cell-type specific, and the potential role for this plasticity in opiate-induced changes in behavior (15). To address these questions, we measured effects of repeated morphine on glutamatergic synaptic transmission in the NAc MSN subpopulations and used optogenetic and pharmacological approaches to determine the role of this pathophysiology in reward-seeking behavior.     

Gsα deficiency in the paraventricular nucleus of the hypothalamus partially contributes to obesity associated with gsα mutations

The G protein α-subunit G(s)α mediates receptor-stimulated cAMP generation. Heterozygous inactivating G(s)α mutations on the maternal allele result in obesity primarily due to reduced energy expenditure in Albright hereditary osteodystrophy patients and in mice. We previously showed that mice with central nervous system (CNS)-specific G(s)α deletion on the maternal allele (mBrGs KO) also develop severe obesity with reduced energy expenditure and that G(s)α is primarily expressed from the maternal allele in the paraventricular nucleus (PVN) of the hypothalamus, an important site of energy balance regulation. We now generated mice with PVN-specific G(s)α deficiency by mating Single-minded 1-cre and G(s)α-floxed mice. Homozygous G(s)α deletion produced early lethality. Heterozygotes with maternal G(s)α deletion (mPVNGsKO) also developed obesity and had small reductions in energy expenditure. However, this effect was much milder than that found in mBrGsKO mice and was more prominent in males. We previously showed mBrGsKO mice to have significant reductions in melanocortin receptor agonist-stimulated energy expenditure and now show that mBrGsKO mice have impaired cold-induced brown adipose tissue stimulation. In contrast, these effects were absent in mPVNGsKO mice. mPVNGsKO mice also had minimal effects on glucose metabolism as compared with mBrGsKO mice. Consistent with the presence of G(s)α imprinting, paternal heterozygotes showed no changes in energy or glucose metabolism. These results indicate that although G(s)α deficiency in PVN partially contributes to the metabolic phenotype resulting from maternal G(s)α mutations, G(s)α imprinting in other CNS regions is also important in mediating the CNS effects of G(s)α mutations on energy and glucose metabolism.     

MC4R在肥胖及糖代谢中的作用

黑皮质素受体4(MC4R)可通过作用于中枢阿片-促黑素细胞皮质素原(POMC)神经元、交感节前神经元、以及与其激动剂的相互作用,调控食物摄入和能量消耗,进而改善肥胖.此外,MC4R还可参与胰岛素敏感性及葡萄糖稳态的调控,为肥胖及糖尿病的治疗提供新靶点.     

Continuous illumination through larval development suppresses dopamine synthesis in the suprachiasmatic nucleus, causing activation of α-MSH synthesis in the pituitary and abnormal metamorphic skin pigmentation in flounder

In order to better understand the endocrine aberrations related to abnormal metamorphic pigmentation that appear in flounder larvae reared in tanks, this study examined the effects of continuous 24-h illumination (LL) through larval development on the expression of tyrosine hydroxylase-1 (th1), proopiomelanocortin (pomc), α-melanophore-stimulating hormone (α-MSH) and melanin concentrating hormone (MCH), which are known to participate in the control of background adaptation of body color. We observed two conspicuous deviations in the endocrine system under LL when compared with natural light conditions (LD). First, LL severely suppressed th1 expression in the dopaminergic neurons in the anterior diencephalon, including the suprachiasmatic nucleus (SCN). Second, pomc and α-MSH expression in the pars intermedia melanotrophs was enhanced by LL. Skin color was paler under LL than LD before metamorphic pigmentation, and abnormal metamorphic pigmentation occurred at a higher ratio in LL. We therefore hypothesize that continuous LL inhibited dopamine synthesis in the SCN, which resulted in up-regulation of pomc mRNA expression in the melanotrophs. In spite of the up-regulation of pomc in the melanotrophs, larval skin was adjusted to be pale by MCH which was not affected by LL. Accumulation of α-MSH in the melanotrophs is caused by uncoupling of α-MSH synthesis and secretion due to inhibitory role of MCH on α-MSH secretion, which results in abnormal metamorphic pigmentation by affecting differentiation of adult-type melanophores. Our data demonstrate that continuous illumination at the post-embryonic stage has negative effects on the neuroendocrine system and pituitary in flounder.     

Neuronal loss in the brainstem and cerebellum--part of the normal aging process? A morphometric study of the vermis cerebelli and inferior olivary nucleus.

Based on the known age-related loss of Purkinje cells (PC) in the cerebellum, this study focuses on whether a marked loss of PC occurs in individuals of very high age. The inferior olive, which is intimately connected with the cerebellum anatomically as well as functionally, was also studied. The study group included 15 nondemented and basically healthy cases aged 32-104 years. Linear neuronal density was expressed as number of PC per millimeter tissue measured in the vermis and as neuronal numbers per square millimeter tissue in the inferior olive. The linear PC density clearly decreased with increasing age, R2 = 0.82 and p < .001. The inferior olive showed a small but insignificant age-related neuronal loss. We conclude that aging results in reduced PC density in the vermis cerebelli, further accentuated in the very late stages of life.     

Adrenocorticotropin hormone (ACTH) effects on MAPK phosphorylation in human fasciculata cells and in embryonic kidney 293 cells expressing human melanocortin 2 receptor (MC2R) and MC2R accessory protein (MRAP)β

Adrenocorticotropin hormone (ACTH) exerts trophic effects on adrenocortical cells. We studied the phosphorylation of mitogen-activated proteins kinases (MAPKs) in human embryonic kidney cells stably expressing the ACTH receptor, MC2R, and its accessory protein MRAPβ and in primary cultures of human adrenal fasciculata cells. ACTH induced a maximal increase in p44/p42(mapk) and of p38 MAPK phosphorylation after 5min. Neither the overexpression of wild-type arrestin2, arrestin3 or their respective dominant negative forms affected p44/p42(mapk) phosphorylation. However, preincubation with the recycling inhibitors brefeldin A and monensin attenuated both cAMP accumulation and p44/p42(mapk) phosphorylation proportionally. Cyclic AMP-related PKA inhibitors (H89, KI(6-22)) and Rp-cAMPS decreased p44/p42(mapk) phosphorylation but not ACTH-mediated cAMP production. The selective Epac1/2 activator, 8-pCPT-2'-O-MecAMP, did not modify the effect of ACTH. Thus, cAMP/PKA, but not cAMP/Epac1/2 pathways, or arrestin-coupled internalization of MC2R is involved in ACTH-induced p44/p42(mapk) phosphorylation by human MC2R. Together, ACTH binding to MC2R stimulates PKA-dependent p44/p42(mapk) phosphorylation.     

Leptin transiently antagonizes ghrelin and long-lastingly orexin in regulation of Ca2＋ signaling in neuropeptide Y neurons of the arcuate nucleus

AIM： To explore the mechanism for interactions of leptin with ghrelin and orexin in the arcuate nucleus （ARC） activating neuropeptide Y （NPY） neurons during physiological regulation of feeding, METHODS： Single neurons from ARC of adult rats with matured feeding function were isolated. [Ca2＋]i was measured to monitore their activities. The time course of leptin effects on ghrelin-induced versus orexin-induced [Ca2＋]i increases in NPY neurons was studied. RESULTS： Administration of ghrelin or orexin-A at 101~ mol/L increased cytosolic Ca2~ concentration （[Ca2＋]~） in NPY neurons isolated from the ARC of adult rats. Upon administration of leptin at 10^-14-10^-12 mol/L, ghrelin-induced [Ca2＋]i increases were initially （〈 10 min） inhibited but later restored, exhibiting a transient pattern of inhibition. In contrast, orexin-induced [Ca2＋]i increases were inhibited by leptin in a long- lasting manner. Furthermore, a prior administration of leptin inhibited orexin action but not ghrelin action to increase ICa 2＋li, CONCLUSION： Leptin counteracted ghrelin effects transiently and orexin effects long-lastingly in NPY neurons. The transient property with which leptin counteracts ghrelin action in NPY neurons may allow the fasting-associated increase in ghrelin levels to activate NPY neurons in the presence of physiological leptin and to stimulate feeding.     

Fos expression incatecholaminergic medullary neu-rons induced by chemical stimulation of stomach projecting to the paraventricular nucleus of hypothalamus in rats

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肥胖MC4R重组腺病毒表达载体的构建及鉴定

目的构建黑素皮质素受体-4（MC4R）重组腺病毒表达载体,为肥胖症的基因治疗研究奠定基础。方法 PCR扩增犬MC4R目的基因后与T载体连接,双酶切鉴定正确的T-A载体获得具有黏性末端的MC4R目的基因;将其亚克隆至腺病毒穿梭载体pShuttle-CMV中,构建穿梭载体pShuttle-CMV/MC4R;PmeⅠ单酶切pshuttle-CMV/犬MC4R后,转化到含pAdeasy-1的BJ5183感受态细胞,采用细菌内同源重组法构建腺病毒载体pAdeasy-1/pshuttle-CMV/MC4R;筛选阳性克隆子;PacⅠ酶切鉴定。结果线性化的pShuttle-CMV/MC4R转化含pAdeasy-1的BJ5183感受态细胞,16 h后获得阳性克隆子,经PacⅠ酶切鉴定出一大于20 kb的大片段和3.0 kb的特征性片段,通过BglⅡ、XhoⅠ双酶切切下约1000 bp的片段,含MC4R目的基因的重组腺病毒载体构建成功。结论细菌内同源重组法可高效、快捷的制备含犬MC4R的重组腺病毒载体;该载体为人类肥胖症的基因治疗研究奠定了基础。     

Use of R ™ stent in the percutaneous coronary intervention of coronary bifurcation lesions

BACKGROUND: Percutaneous Coronary Intervention (PCI) of coronary bifurcation lesion is technically quite demanding. It has been associated with a lower procedural success, higher rates of complication and restenosis. Side-branch occlusion and plaque shifting or 'snow plow' effect are not uncommon. Stenting of the main vessel may cause 'stent jail' of the side-branch. Modern stent design may allow passage of a balloon or stent into the side-branch through the struts of the stent placed in the main vessel. A newly developed 316 stainless steel tubular stent, the R? stent is uniquely designed to provide flexibility, radial strength on deployment and conformability. Its large cell size facilitates PCI of bifurcation lesion. AIM: To assess the feasibility of R ? stent in the treatment of symptomatic patients with bifurcation coronary lesions. The main objective was to assess the ease of deployment, side-branch access and overall success of the R ? stent in this group of patients without any major adverse events. METHODS: Between December 1998 and September 2000 the R ? stent was used as a main stent in 28 consecutive patients with coronary bifurcation lesions, 46% of which had unstable angina. The mean age was 59 &#45 10 and 89% were male. Adjunctive medical therapy included clopidrogel, aspirin and intraprocedure heparin. Abciximab (ReoPro) was given to 9 patients. RESULTS: Successful stent deployment was achieved in all patients. Thirty-four R Stents and 16 other stents were used. Two patients had post-procedure rise in cardiac enzymes. There were no major adverse events at 30 days. LAD/D1 with LAD/diagonal was the target lesion in the majority of patients. Stenting of the side-branch was done in 18 and balloon dilatation in 9 patients. At 3-23 months (mean 11.8) follow-up, repeat angiography was done in 18 patients with restenosis in 4, two of them had repeat PCI and one had coronary artery bypass graft (CABG). CONCLUSION: Coronary bifurcation lesions are not uncommon. Current advances in stent technology offer a safe and effective revascularisation strategy for such complex lesions. The R ? stent appears to be a suitable device that provides good wall coverage, radial strength, conformability and easy side-branch access.     

Ambient temperature and 17β-estradiol modify Fos immunoreactivity in the median preoptic nucleus, a putative regulator of skin vasomotion

Estrogen has pronounced effects on thermoregulation, but the anatomic sites of integration between the reproductive and thermoregulatory axes are unknown. In this study, we tested whether estradiol-17β (E(2)) treatment would alter the activity of thermoregulatory brain regions responding to mild changes in ambient temperature (T(AMBIENT)). Core and tail skin temperatures were recorded at the ambient temperatures of 20, 24, or 31 C in ovariectomized (OVX) rats with and without E(2). Neuronal activity was evaluated by counting the number of Fos-immunoreactive cells in the brains of rats killed 90 min after exposure to one of the three ambient temperatures. Of 14 brain areas examined, the median preoptic nucleus (MnPO) was the only site that exhibited increased Fos immunoreactivity at the high T(AMBIENT) of 31 C. At 24 C, OVX rats exhibited increased numbers of MnPO Fos-immunoreactive cells, compared with OVX + E(2) rats. Interestingly, tail skin vasomotion and MnPO Fos expression were affected in a similar manner by T(AMBIENT) and E(2) treatment. In the arcuate nucleus and anteroventral periventricular nucleus (AVPV), Fos immunoreactivity was highest at the low T(AMBIENT) of 20 C, with inhibitory (arcuate nucleus) and stimulatory (AVPV) effects of E(2). No other areas responded to both T(AMBIENT) and E(2) treatment. These results implicate the MnPO, the arcuate nucleus, and the AVPV as sites of integration between the reproductive and thermoregulatory axes. Combined with studies showing the importance of MnPO neurons in heat-defense pathways, the MnPO emerges as a likely site for E(2) modulation of thermoregulatory vasomotion.