Over-expression of SK2 channel in PVN reduces renal sympathetic nerve activity in chronic heart failure rats

AIM：To investigate the effect of over-expression of small-conductance calcium-activated potassium channel subtype 2 (SK2) in the paraventricular nucleus (PVN) of hypothalamus on heart rate(HR), mean arterial pressure (MAP) and renal sympathetic nerve activity (RSNA) of the rats with chronic heart failure (CHF). METHODS：Adenovirus vector encoding SK2 (pDC316-mCMV-EGFP-rKCNN2) was constructed.The CHF model of the male Sprague Dawley (SD) rats was established by the ligation of anterior descending coronary artery. Echocardiogram was used at the 6th week after the operation to identify the CHF model. Adenovirus vector encoding SK2 (pDC316-mCMV-EGFP-rKCNN2) or control adenovirus vector (pDC316-mCMV-EGFP) were transfected into the PVN in vivo. The cardiac function was monitored by echocardiogram. The expression of SK2 at mRNA and protein levels was determined by RT-PCR, Western blotting and immunofluorescence. The HR, MAP and RSNA were measured by PowerLab 8/30 in anesthetized rats with bilateral PVN microinjection of SK channel blocker apamin. RESULTS：Treatment with pDC316-mCMV-EGFP-rKCNN2 significantly decreased the renal sympathetic nerve activity in the rat with CHF. Injection of pDC316-mCMV-EGFP did not change the renal sympathetic nerve activity in the rats in sham group. CONCLUSION：The expression and function of SK channels in PVN in the heart failure rats were decreased, suggesting a reduced negative regulation of sympathetic activity in central neural system. Increased expression of SK2 in the PVN leads to a reduction in sympathetic outflow under the condition of CHF, which may provide a new target for the treatment of heart failure.     

In vitro evidence that leptin suppresses melatonin secretion during long days and stimulates its secretion during short days in seasonal breeding ewes

Recent observations in seasonal-breeding mammals indicate that the hypothalamus is programmed to become leptin resistant during long days (LD) and leptin sensitive during short days (SD). These observations support the possibility that photoperiod mediates at least part of its effects on melatonin secretion through changes in leptin sensitivity. Herein we examined the interaction of season and recombinant ovine leptin (oleptin) on melatonin secretion by pineal explants in short-term culture. Glands were collected after sunset from eight ewes during LD (March, April, May, June) and from an additional eight ewes during SD (September, October, November, December). Glands were transected saggitally and coronally into quarters, with each equilibrated in 2.5ml of DMEM for 120min, followed by a 3h incubation in medium containing either 0 or 50ng/ml of oleptin. Treatment with oleptin reduced (P<0.001) melatonin secretion compared to controls during LD by approximately 22% at 2, 2.5 and 3h of culture. However, in cultures from glands collected during SD, oleptin stimulated (P<0.078) melatonin secretion approximately 50% compared to control. These effects were consistent throughout each respective season. We conclude that the secretion of melatonin from the ovine pineal gland is negatively responsive to leptin during LD, whereas leptin may stimulate melatonin secretion during SD.     

GnIH多肽对半滑舌鳎(Cynoglossus semilaevis)下丘脑生殖相关基因表达的影响

本研究首次通过下丘脑离体孵育的方法研究促性腺激素抑制激素(Gonadotropin-inhibitory hormone,GnIH)多肽对半滑舌鳎(Cynoglossus semilaevis)下丘脑中生殖相关基因的表达调控.研究结果显示,tsGnIH-1促进了gnrh2和gnih的表达,对gnrh3和kiss2的表达无影响;tsGnIH-2抑制了gnrh3的表达,对gnrh2、kiss2和gnih的表达无影响.GnIH多肽对生殖相关基因的不同调控表明同一前体蛋白编码的不同GnIH多肽在生殖调控中的作用不尽相同.本研究结果增加了对GnIH参与鱼类生殖调控机制的认识,为深入研究奠定了基础.     

Effects of "Bioactive" amino acids leucine, glutamate, arginine and tryptophan on feed intake and mRNA expression of relative neuropeptides in broiler chicks

ABSTRACT: Feed intake control is vital to ensuring optimal nutrition and achieving full potential for growth and development in poultry. The aim of the present study was to investigate the effects of L-leucine, L-glutamate, L-tryptophan and L-arginine on feed intake and the mRNA expression levels of hypothalamic Neuropeptide involved in feed intake regulation in broiler chicks. Leucine, glutamate, tryptophan or arginine was intra-cerebroventricularly (ICV) administrated to 4d-old broiler chicks respectively and the feed intake were recorded at various time points. Quantitative PCR was performed to determine the hypothalamic mRNA expression levels of Neuropeptide Y (NPY), agouti related protein (AgRP), pro-opiomelanocortin (POMC), melanocortin receptor 4 (MC4R) and corticotrophin releasing factor (CRF). Our results showed that ICV administration of L-leucine (0.15 or 1.5 mumol) significantly (P < 0.05) increased feed intake up to 2 h post-administration period and elevated both hypothalamic NPY and AgRP mRNA expression levels. In contrast, ICV administration of L-glutamate (1.6 mumol) significantly (P < 0.05) decreased feed intake 0.25, 0.5 and 2 h post-injection, and increased hypothalamic CRF and MC4R mRNA expression levels. Meanwhile, both L-tryptophan (10 or 100 mug) and L-arginine (20 or 200 mug) had no significant effect on feed intake. These findings suggested that L-leucine and L-glutamate could act within the hypothalamus to influence food intake, and that both orexigenic and anorexigenic Neuropeptide genes might contribute directly to these effects.     

Inhibition of acetylcholinesterase activity by rivastigmine decreases lipopolysaccharide-induced IL-1β expression in the hypothalamus of ewes

The present study was designed to determine the effect of subcutaneous rivastigmine treatment on IL-1β expression and IL-1 type I receptor (IL-1R1) gene expression in the hypothalamic structures (preoptic area [POA], anterior hypothalamus [AHA], and medial basal hypothalamus [MBH]) of ewes after lipopolysaccharide (LPS) treatment. Endotoxin treatment increased (P ≤ 0.01) both IL-1β and IL-1R1 gene expression in the POA, AHA, and MBH compared with the control group, whereas concomitant rivastigmine and LPS injection abolished this stimulatory effect. It was also found that LPS elevated (P ≤ 0.01) IL-1β concentration in the hypothalamus (71.0 ± 2.3 pg/mg) compared with controls (16.1 ± 3.6 pg/mg). The simultaneous injection of LPS and rivastigmine did not increase IL-1β concentration in the hypothalamus (24.6 ± 13.0 pg/mg). This central change in IL-1β synthesis seems to be an effect of acetylcholinesterase (AChE) inhibition by rivastigmine, which decreases (P ≤ 0.01) the activity of this enzyme from 78.5 ± 15.0 μmol · min⁻¹ · g⁻¹ of total protein in the control and 68.8 ± 9.8 μmol · min⁻¹ · g⁻¹ of total protein in LPS-treated animals to 45.2 ± 5.6 μmol · min⁻¹ · g⁻¹ of total protein in the rivastigmine and LPS-treated group. Our study showed that rivastigmine could effectively reverse the stimulatory effect of immune stress induced by LPS injection on IL-1β synthesis through a decrease in AChE activity in the hypothalamic area of sheep. Our results also proved that the cholinergic anti-inflammatory pathway could directly modulate the central response to endotoxin.     

下丘脑IFN-γR和ER免疫组化双标技术探索

[目的]选出一种提高下丘脑中雌激素受体(ER)和γ-干扰素受体(IFN-γR)免疫组化双标阳性检出率和染色强度的方法。[方法]用MaxVision法对大鼠下丘脑冰冻切片和石蜡切片分别进行IFN-γR和ER免疫组化双标染色,其中石蜡切片进行抗原微波修复。[结果]下丘脑石蜡切片抗原微波修复后IFN-γR和ER阳性检出率较冰冻切片有明显提高,且双标着色对比鲜明,易于镜下观察。[结论]下丘脑IFN-γR和ER免疫组化双标染色宜选用石蜡切片并进行微波修复,能提高双标阳性率,且显色强,值得推广。     

Effect of ghrelin in septal nucleus on gastric motility of diabetic gastroparesis rats and its potential mechanism regulated by hypothalamic arcuate nucleus

AIM：To study the effect of ghrelin in septal nucleus on the gastric motility of the rats with diabetes mellitus (DM) and to investigate the regulation of ghrelin pathway between arcuate and septal nucleus nuclei on gastric motility. METHODS：Streptozotocin was injected intraperitoneally to establish a DM rat model. Fluorescence immunohistochemistry and real-time PCR were used to detect the expression of ghrelin receptor GHS-R1a. The gastric motility was evaluated by implantation of a force transducer on the surface of rats stomachs and the motility index was also calculated. The neural connections between arcuate and septal nuclei were analyzed by the technique of fluorogold tracing. The neural control pathway of gastric motility was determined by central drug injection, nucleus lesion or nucleus electrical stimulation. RESULTS：The expression of GHS-R1a in the septal nucleus of DM rats was lower than that in normal rats (P<0.05). The amplitude and frequency of gastric motility in the DM rats were lower than those in the normal rats (P<0.05). The gastric motility of normal and DM rats were increased by injection of ghrelin into the septal nucleus in a dose-dependent manner. Seven days after injection of fluorogold into the septal nucleus, some neurons in arcuate nucleus were labeled by fluorogold and part of the labeled neurons were ghrelin immunopositive. No effect of nucleus lesion or nucleus electrical stimulation on the gastric motility in the normal rats was observed. In DM rats, the lesion of septal nucleus decreased the gastric motility (P<0.05). In the normal rats, the change of gastric motility caused by electrical stimulation in arcuate nucleus was not affected by the lesion of septal nucleus (P>0.05), while the change was attenuated in DM rats (P<0.05). The ghrelin receptor antagonist ［D-Lys-3］-GHRP-6 had no significant effect on the gastric motility induced by electrical stimulation in arcuate nucleus of the normal rats (P>0.05), but it reduced the change in the DM rats (P<0.05). CONCLUSION:Ghrelin in septal nucleus and the ghrelinergic pathway between arcuate and septal nuclei play an important role in the modulation of gastric motility in DM rats.     

Sexual maturation in hens is not associated with increases in serum leptin and the expression of leptin receptor mRNA in hypothalamus

Background

In mammals, leptin is an attractive candidate for mediating the metabolic signal and the reproductive function via the specific receptor in hypothalamus. However, till now, the role of leptin on reproduction in birds is less well established. This experiment was conducted to elucidate the role of leptin on the onset of reproduction in bird, as a first step, to detect the changes of peripheral leptin and leptin receptor mRNA expression in hypothalamus between mature and immature hens at the same age. 120 ISA brown pullets at D60 were allocated randomly into two groups, long light (LL) group being raised under artificial light regimes with incrementally increased light phase (from 8 L:16D to 14 L:12D) and short light (SL) group raised on consistent light (8 L:16D) for 12 wk.

Results

The results showed that pullets in LL group reached sexual maturation 15 d earlier than those in SL group. Serum E₂ showed a significant increase with age, but no difference was observed between two groups. Serum leptin concentration decreased significantly from D112 to D136 in LL, and was markedly higher in LL group than that in SL at D112, while there was no significant difference between two groups at D136. Leptin receptor and GnRH-I mRNA expression in hypothalamus were significantly increased with age, yet there was no significant difference between SL and LL chickens at the same age. The expression of FSH-β and LH-β mRNA in pituitary was increased with age but did not show significant difference between LL and SL group. GnRH-I mRNA expression was very rich in pineal gland, and decreased from D112 to D136 in LL but not in SL group, and there was no difference between two groups at the same age.

Conclusions

These results indicate that the earlier sexual maturation in hens induced by long-light regime is not accompanied with an increase in serum leptin or leptin receptor gene expression in hypothalamus, or genes expression in HPG axis.     

Sexual maturation in hens is not associated with increases in serum leptin and the expression of leptin receptor mRNA in hjpothalamus

Background： In mammals, leptin is an attractive candidate for mediating the metabolic signal and the reproductive function via the specific receptor in hypothalamus. However, till now, the role of leptin on reproduction in birds is less well established. This experiment was conducted to elucidate the role of leptin on the onset of reproduction in bird, as a first step, to detect the changes of peripheral leptin and leptin receptor mRNA expression in hypothalamus between mature and immature hens at the same age. 120 ISA brown pullets at D60 were allocated randomly into two groups, long light （LL） group being raised under artificial light regimes with incrementally increased light phase （from 8 L：]6D to 14 L：]2D） and short light （SL） group raised on consistent light （8 L：16D） for 12 wk. Results： The results showed that pullets in LL group reached sexual maturation 15 d earlier than those in SL group. Serum E2 showed a significant increase with age, but no difference was observed between two groups. Serum leptin concentration decreased significantly from D112 to D136 in LL, and was markedly higher in LL group than that in SL at D112, while there was no significant difference between two groups at D136. Leptin receptor and GnRH-I mRNA expression in hypothalamus were significantly increased with age, yet there was no significant difference between SL and LL chickens at the same age. The expression of FSH-13 and LH-13 mRNA in pituitary was increased with age but did not show significant difference between LL and gland, and decreased from D112 to D136 in LL but not groups at the same age. SL group. GnfiH-I mRNA expression was very rich in pinea n SL group, and there was no difference between two Conclusions： These results indicate that the earlier sexual maturation in hens induced by long-light regime is not accompanied with an increase in serum leptin or leptin receptor gene expression in hypothalamus, or genes expression in HPG axis.     

The Effect of Energy on Serotonin-Like Neurons in Duck Hypothalamus

In the present study, immunohistochemical method (SABC method) was used to examine the distribution of serotonin-like neurons in hypothalamus of Cherry Valley ducks bred with high energy diet and low energy diet respectively. All films were analysed by using a computer-assisted image analysis system. In high energy group, labelled neurons are widely distributed in paraventricular nucleus(PVN, 17.73± 1.41 neurons/19.46× 103 μm2) and ventromedial hypothalamus (VMH,15.8±1.71 neurons/19.46×103 μm2); in low energy group, labelled ceils are widely distributed in lateral hypothalamus (LH,11.25±1.53 neurons/19.46×103 μm2), yet there is no positive neuron in PVN. These results indicate that serotonin-like neurons in hypothalamus are involved in the regulation of food intake and energy metabolism.