Cloning,Tissue Distribution,and Transmembrane Orientation of the Olfactory Co-Receptor Orco from Two Important Lepidopteran Rice Pests,the Leaffolder(Cnaphalocrocis medinalis) and the Striped Stem Borer(Chilo suppressalis)

In insects,the sense of smell is mainly mediated by olfactory receptors（Ors）.Olfactory co-receptor（Orco）,which is coexpressed with the Ors in almost all olfactory receptor neurons（ORNs）,is demonstrated to be an essential component in the insect olfactory system.It can be potential target for developing novel olfactory-disruption strategy to control insect pests.In this study,two full-length cDNA sequences encoding Orcos（CmedOrco and ChsupOrco） were cloned from two Lepidopteran rice pests,the rice leaffolder,Cnaphalocrocis medinalis and the rice striped stem borer,Chilo suppressalis.The amino acid sequences of CmedOrco and ChsupOrco showed high similarity to the previously identified Orcos from other insect species. Bioinformatic prediction and cellular immunofluorescence indicated that CmedOrco and ChsupOrco were both seven-transmembrane proteins with intracellular N-termini and extracellular C-termini.mRNA expression levels of the two Orcos were much higher in male and female antennae than those in non-olfactory tissues,and the ChsupOrco transcripts reached a peak level in adults compared to other life stages.Our results provide a foundation from which it will be possible to elucidate the roles of Orco in moth olfaction and for the development of environment-friendly management strategies of these two rice insect pests.

Mating behavior induces changes of expression of Fos protein,plasma testosterone and androgen receptors in the accessory olfactory bulb (AOB) of the male mandarin vole Microtus mandarinus

In order to investigate the neuroendocrine mechanism of the mating behavior in the adult male mandarin voles Microtus mandarinus,the radioimmunoassay(RIA)and immunohistochemistry methods were used to investigate the differences in plasma testosterone(T)concentrations and distribution of T immunoreactive neurons(T-IRs),androgen receptor immunoreactive neurons(AR-IRs)and Fos protein immunoreactive neurons(Fos-IRs)in the accessory olfactory bulb(AOB)and the main olfactory bulb(MOB)following exposure to clean hard-wood shavings(control group),soiled bedding(exposure group)or contact with an estrous female(mating group).Results showed that plasma T concentration was significantly higher in the mating group than that in the exposure group,and both the mating group and the exposure group displayed significantly higher plasma T concentration than the control group.T-IRs,AR-IRs and Fos-IRs were investigated with the immunohistochemistry method in granule cell(GC)and mitral cell(MC)of the MOB and the AOB in the three groups.There were significantly more T-IRs,AR-IRs and Fos-IRs in MC and GC of the AOB in the mating group than that in the exposure group or the control group.T-IRs,AR-IRs and Fos-IRs did not show significant differences between the exposure group and the control group.Furthermore,obvious differences in MC and GC of the MOB were not found among the three groups.The results confirm that both changes of T and AR in the AOB might be underlying mating behavior in the adult male mandarin voles.

Expression of estrogen receptor (ER) -α and -βtranscripts in the neonatal and adult rat cerebral cortex, cerebellum, and olfactory bulb

In the present study expression of estrogen receptor subtype -alpha (ERalpha) and -beta (ERbeta) in the cerebral cortex, cerebellum, and olfactory bulb was investigated and compared between neonatal (1 to approximately 3-days-old) and adult (250 to approximately 350 g) rats, using reverse transcription-polymerase chain reaction (RT-PCR). No ERalpha transcripts were detectable in the adult cerebellum and olfactory bulb, whereas very weak expression of ERalpha was present in the adult cerebral cortex. No significant difference in ERbeta transcripts was detectable between the neonatal and adult rats. While transcripts for both ER subtypes were co-expressed in these brain areas of neonatal rats, although ERalpha expression was significantly weaker than ERbeta. Even in the cerebral cortex known to contain both ER subtypes in adult rats, ERalpha transcripts in neonatal rats were much higher than in adult. These observations provide evidence for the existence of different expression patterns of ERalpha/ERbeta transcripts in these three brain areas between the neonatal and adult rats, suggesting that each ER subtype may play a distinct role in the regulation of differentiation, development, and functions of the brain by estrogen.

The Olfactory Receptor Pseudo-pseudogene: A Potential Therapeutic Target in Human Diseases

Recently, Prieto-Godino et al.[1] found that the olfactory receptor 75a （Ir75a） gene is a functional pseudo-pseudogene in Drosophila sechellia. For a long time, Ir75a has been regarded as an acetic acid receptor that detects acetic acid and induces obvious olfactory responses in olfactory sensory neurons （OSNs）f2J. Nonetheless, Prieto-Godino et al. confirmed that Ir75a lost its sensitivity to acetic acid in D. sechellia. Thus, the D. sechelfia Ir75a gene is generally recognized as a pseudogene in OSNs.

Comparison of the fraction of olfactory receptor pseudogenes in wolf (Canis lupus) with domestic dog (Canis familiaris)

Olfactory receptors（ORs）,the first dedicated molecules with which odorants physically interact to arouse an olfactory sensation,constitute the largest gene family in vertebrates.Dogs and wolves,like many other mammals,have a highly developed capability to detect and identify odorant molecules,even at minimum concentrations.In this study,the olfactory receptor repertoire from domestic dog and its closest relative,the wolf,were sequenced to estimate the fraction of pseudogenes in each subspecies.The fraction of disrupted olfactory receptor genes in dog was 17.78%,whereas,that in wolf was 12.08%.As expected the dog was less dependent on olfaction than the wolf,and the dog had more olfactory receptor pseudogenes.However,the observed difference between the two subspecies was not at the significant level（χ2 = 1.388,p = 0.239 0.05）.The values indicated that although domestication might play a role in the reduction of OR genes,it could not be concluded that the living environment provided by domestication lead to a significant reduction of the functional olfactory receptor repertoire.Furthermore,the purpose of domestication may also have influence on the ratio of functional olfactory receptor genes reduction.

Rivastigmine Restores 5-HT_1AReceptor Levels in the Hippocampus of Olfactory Bulbectomized Mice

Rivastigmine, a dual acetylcholinesterase and butyrylcholinesterase inhibitor, is used for symptomatic treatment of patients with mild to moderately severe dementia in Alzheimer’s disease (AD) patients. In the present study, we found that 5-HT1A receptor (5-HT1AR) is downregulated, whereas 5-HT2A receptor (5-HT2AR) is upregulated in the hippocampal dentate gyrus (DG) and CA1 region by olfactory bulbectomy (OBX) in mice. Furthermore, chronic treatment with rivastigmine (1.0 mg/kg) for 2 weeks starting 2 weeks after OBX operation restored the decreased 5-HT1AR and the increased 5-HT2AR levels. To determine whether cholinergic receptor stimulation by rivastigmine is involved in the rivastigmine-induced regulation of 5-HTR levels, we treated the mice with mecamylamine (2.5 mg/kg), or atropine (5.0 mg/kg) with rivastigmine (1.0 mg/kg) once a day for 2 weeks. Notably, the rivastigmine-induced 5-HT1AR upregulation was eliminated by mecamylamine but not by atropine treatments. On the other hand, the restored 5-HT2AR level by rivastigmine was not affected by either mecamylamine or atropine. Treatment with 8-OH-DPAT, a selective 5-HT1AR agonist improved the decreased 5-HT1AR and the increased 5-HT2AR levels in OBX mice. On the other hand, treatment with TCB-2, a potent 5-HT2AR agonist had no effects on the 5-HT1AR and 5-HT2AR dysregulation in OBX mice. Taken together, nicotinic acetylcholine receptor (nAChR) stimulation mediates rivastigmine-induced upregulation of 5-HT1AR. Therefore, we speculate that the increased ACh levels by rivastigmine can stimulate nAChR located on serotonergic nerve terminals and stimulate 5-HT1AR by the enhanced 5-HT release in the hippocampus. The 5-HT1AR stimulation likely mediates the improvement of 5-HT1AR levels as auto-receptor in OBX hippocampus.

Lateral olfactory tract usher substance(LOTUS) protein,an endogenous Nogo receptor antagonist,converts a non-permissive to permissive brain environment for axonal regrowth

It is well known that primates,including humans,hardly recover motor function after spinal cord injury（SCI）when compared with non-primate mammals such as rodents.This limited functional recovery is in part due to a non-permissive environment of the central nervous system（CNS）inhibiting axonal regrowth.

Progress in Research on Insect Olfactory Perception of Habitat Odor Molecules

A highly sensitive olfactory system allows insects to precisely identify and position volatile compounds from different sources in their habitats,and plays a crucial role in their foraging,mating,and oviposition activities.During evolution,insects have successfully developed a large and complex olfactory system to adapt to heterogeneous environments,enabling the maintenance of inset population.A comprehensive examination of the olfactory system of insects may therefore yield novel insights into the development of innovative pest control and prevention strategies,as well as the study of olfactory mechanisms in vertebrates and even humans.This paper outlines the current state of research into the signal transduction mechanism by which insects perceive the olfactory molecules of their habitats.The aim of this review is to provide a reference point for future studies into the olfactory perception mechanism and its potential applications in pest management.

Molecular Cloning and cDNA Sequence Analysis of Two New Lepidopteran OR83b Orthologue Chemoreceptors

The aim of this study was to isolate the full-length cDNA sequences of an unusually highly conserved olfactory receptor （orthologue to the Drosophila melanogaster DOR83b） from the antennae of Spodoptera exigua and S. litura by using reverse transcription-polymerase chain reaction （RT-PCR） and rapid amplification of cDNA ends （RACE） methods. Bioinformatics methods were used to further analyze the cDNA sequences and putative amino acid sequences. The two full-length cDNA sequences from olfactory receptor （OR） of male S. exigua and S. litura were named as SexiOR2 and SlitOR2, respectively. SexiOR2 and SlitOR2 consisted of nucleotide sequence of 1 906 and 2 483 bp, respectively, and both with deduced amino acid sequences of 473 residues. The sequence analysis indicated that the deduced amino acid sequences of the cDNA shared the high homologies with OR83b orthologue chemoreceptor sequences from previously reported moths, implying that the cDNA sequences were of OR83b orthologue chemoreceptor genes.

Factors that modulate olfactory dysfunction

The olfactory system is one of a few areas in the nervous system which is capable of regeneration throughout the life.Olfactory sensory neurons reside in the nasal cavity are continuously replenished with new neurons arising from stem cells.Some factors such as aging,neurodegenerative diseases,head trauma,brain tumor extraction and infection cause olfactory dysfunction which significantly influences physical wellbeing,quality of life,mental health,nutritional status,memory processes,identifying danger and is associated with increased mortality.Therefore,finding a treatment to improve olfactory dysfunction is needed.Recent research efforts in the field have shown some very promising new approaches to treat olfactory dysfunction.This review explores the current studies that have addressed therapeutic approaches to improve olfactory neuron regeneration based on cell transplantation therapy,modulation of physiological olfactory dysfunction and drug treatments.

Microencapsulation improves inhibitory effects of transplanted olfactory ensheathing cells on pain after sciatic nerve injury

Olfactory bulb tissue transplantation inhibits P2X2/3 receptor-mediated neuropathic pain. However, the olfactory bulb has a complex cellular composition, and the mechanism underlying the action of purified transplanted olfactory ensheathing cells（OECs） remains unclear. In the present study, we microencapsulated OECs in alginic acid, and transplanted free and microencapsulated OECs into the region surrounding the injured sciatic nerve in rat models of chronic constriction injury. We assessed mechanical nociception in the rat models 7 and 14 days after surgery by measuring paw withdrawal threshold, and examined P2X2/3 receptor expression in L4–5 dorsal root ganglia using immunohistochemistry. Rats that received free and microencapsulated OEC transplants showed greater withdrawal thresholds than untreated model rats, and weaker P2X2/3 receptor immunoreactivity in dorsal root ganglia. At 14 days, paw withdrawal threshold was much higher in the microencapsulated OEC-treated animals. Our results confirm that microencapsulated OEC transplantation suppresses P2X2/3 receptor expression in L4–5 dorsal root ganglia in rat models of neuropathic pain and reduces allodynia, and also suggest that transplantation of microencapsulated OECs is more effective than transplantation of free OECs for the treatment of neuropathic pain.

Related Factors Affecting Olfactory Ability of Dogs

Dog has a very sensitive olfactory sensory system. Observation showed that there were great differences in olfactory ability of different breeds of dogs, and even the same dog presented different olfactory abilities under different physiological conditions. Thus, the author discussed several factors influencing olfaction of dog from the aspects of gene, environment, anatomy and physiology.

Roles of the main olfactory and vomeronasal systems in the detection of androstenone in inbred strains of mice

We investigated the role of the main olfactory and accessory olfactory systems （MOS and AOS respectively） in the detection of androstenone. We used the following experimental approaches： behavioral, surgical removal of the vomeronasal organ （VNX） followed by histochemical verification and Fos immunohistochemistry. Using a Y-maze paradigm we estimated sensitivity of NZB/B1NJ and CBA/J mice to androstenone. CBA mice were 2,000-fold more sensitive to androstenone than NZB mice. VNX caused a 4-tol6-fold decrease in sensitivity to androstenone in highly-sensitive CBA mice, but did not affect thresholds in NZB mice. Results indicate the involvement of the MOS and AOS in the detection of androstenone. We observed a specific pattern of Fos-positive cells in the main olfactory bulb of CBA mice but not in NZB mice subsequent to exposure of mice to androstenone; the compound activated cells in the accessory olfactory bulb in both strains of mice, indicating the involvement of the vomeronasal organ. Patterns of Fos-positive cells in the vomeronasal organ were recorded subsequent to exposure to androstenone. Fos-positive receptor cells in the vomeronasal organ of CBA and NZB mice were different, in CBA mice Fos-positive cells were noted in both the basal and apical zones, however, in NZB mice activation was observed only in the apical zone [Current Zoology 56 （6）： 813-818, 2010].

基于嗅觉受体激活关系模拟的气味感知预测

气味分子与嗅觉受体相互作用是引起气味感知的重要环节,对于揭示气味感知机制具有重要意义。然而,获得气味分子与人类嗅觉受体激活关系的实验性结果耗时耗力,且目前可用的激活关系数据数量不足以支持智能气味感知研究。因此,本研究构建了嗅觉受体蛋白质关系网络,并提取特征来训练气味分子-嗅觉受体激活关系预测模型。在气味感知预测中综合考虑气味分子特征和嗅觉受体蛋白激活模拟关系,实现了对人类气味感知的高精度回归预测。实验结果表明,融合气味分子-嗅觉受体激活关系的人类气味感知预测相关度指标为0.94,明显优于现有的气味感知预测模型。此外,研究还在预测基础上总结了气味分子-嗅觉受体激活-气味感知模式。本研究为气味感知预测引入了可观测的嗅觉受体激活机制特征,为深入探索和理解气味感知机制提供了新思路。

The olfactory system of Pieris brassicae caterpllars:from receptors to glomeruli

Tolfactory system of adult lepidopterans is among the best described neuronal circuits.However,comparatively little is known about the organization of the olfactory system in the larval stage of these insects.Here,we explore the expression of olfactory receptors and the organization of olfactory sensory neurons in caterpillars of Pieris brassicae,a significant pest species in Europe and a well-studied species for its chemical ecology.To describe the larval olfactory system in this species,we first analyzed the head transcriptome of third-instar larvae(L3)and identified 16 odorant receptors(ORs)including the OR coreceptor(Orco),13 ionotropic receptors(IRs),and 8 gustatory receptors(GRs).We then quantified the expression of these 16 ORs in different life stages,using qPCR,and found that the majority of ORs had significantly higher expression in the L4 stage than in the L3 and L5 stages,indicating that the larval olfactory system is not static throughout caterpillar development.Using an Orco-specific antibody,we identified all olfactory receptor neurons(ORNs)expressing the Orco protein in L3,L4,and L5 caterpillars and found a total of 34 Orco-positive ORNs,distributed among three sensilla on the antenna.The number of Orco-positive ORNs did not differ among the three larval instars.Finally,we used retrograde axon tracing of the antennal nerve and identified a mean of 15 glomeruli in the larval antennal center(LAC),suggesting that the caterpillar olfactory system follows a similar design as the adult olfactory system,although with a lower numerical redundancy.Taken together,our results provide a detailed analysis of the larval olfactory neurons in P brassicae,highlighting both the differences as well as the commonalities with the adult olfactory system.These findings contribute to a better understanding of the development of the olfactory system in insects and its life-stage-specific adaptations.

团头鲂嗅觉器官的发育及代表性嗅觉受体基因的表达模式

为了研究团头鲂嗅觉器官的发育过程和嗅觉受体基因(ORs)在嗅囊发育不同时期的组织表达模式,实验基于组织学、形态学及实时荧光定量PCR技术(qRT-PCR)的方法进行了探究。组织形态学结果显示,团头鲂嗅觉器官位于后背部和两侧眼睛前方,嗅囊紧贴于嗅腔底部,形状近似椭圆形,嗅基板沿着头尾方向平行排列。嗅囊在团头鲂刚孵化时出现嗅窝外围凹进的痕迹,随后凹进处长出嗅窝,并在中央出现长条形的嗅基板。随着仔鱼的生长发育,单侧嗅囊初级嗅基板由疏松逐渐排列紧密,嗅基板隆起高度、数量及嗅囊总体表面积逐渐增加,在成鱼阶段趋于稳定。qRT-PCR结果显示,代表性ORs在团头鲂发育不同时期的组织中表达模式各异,其中Beta-2、9、10、11在胚胎发育期的囊胚期、胚孔闭合期及嗅板期高表达;Beta-2、9、10、11和Epsilon-7在仔稚鱼期的3~15 dpf中微弱表达,在30~60 dpf时期高表达;Beta-2、10、11及Epsilon-6、7、10、13在幼鱼期至成鱼期嗅囊中高表达,Beta-2、10、9、11及Epsilon-7、10、13在3月龄团头鲂嗅球中高表达;Beta-2、10在幼鱼期至成鱼期脑中低表达。研究表明,团头鲂嗅觉器官的发育进程与ORs表达相关,不同发育时期ORs主要表达部位均在嗅囊中。本研究对进一步探究ORs的功能提供理论支持,也为后续深入研究鱼类嗅觉识别机制奠定了基础。

异位嗅觉受体的功能及其作为药物靶点的研究进展

嗅觉受体(olfactory receptors,ORs)是一类主要在鼻上皮嗅觉感觉神经元中分布的跨膜蛋白,介导气味向大脑传递实时感觉信号而产生嗅觉。近年来研究发现,ORs也可在鼻腔外组织或器官中表达,且与多种生物过程密切相关,如精子趋化性、伤口愈合、糖脂代谢及肠道分泌等。此外,ORs还与前列腺癌、乳腺癌及结直肠癌等多种恶性肿瘤关系密切,通过调节细胞增殖、凋亡、迁移及侵袭等过程影响肿瘤的发生发展。本文概述了异位ORs对人体组织器官功能的影响,并评估它们作为治疗疾病的药物靶点的潜在价值。

蜜蜂气味结合蛋白与外源性化合物互作机制研究进展

嗅觉是蜜蜂赖以生存和繁殖的主要感官方式,可为蜜蜂外出觅食、躲避天敌、交配等行为提供重要信息,而气味结合蛋白(odorant-binding proteins,OBPs)在其嗅觉感知中发挥着关键作用。探究外界环境中各种化学物质与蜜蜂OBPs之间的相互作用,对于揭示蜜蜂OBPs对不同外源性化合物的结合特性和结合机制具有重要意义。外源性化合物主要包括化学信息物质(信息素和蜜源开花植物挥发物)以及杀虫剂等农用化学品。一方面,化学信息物质与蜜蜂OBPs结合,在维持蜂群稳定、繁殖、觅食、授粉等生理功能中发挥着重要作用;另一方面,杀虫剂与蜜蜂OBPs结合则可能危害蜜蜂嗅觉系统,影响或干扰蜜蜂对环境气味分子的识别。本文综述了蜜蜂OBPs的种类、功能及其与外源性化合物互作机制的研究进展,以期为深入探究蜜蜂OBPs的生理功能、保护蜜蜂免受杀虫剂等化学物质的危害提供参考。

荒漠草地白刺粗角叶甲非典型气味受体基因克隆及组织表达谱

白刺粗角叶甲是我国西北荒漠草原危害防风固沙先锋植物白刺的重要害虫之一,开发和利用基于性信息素或寄主挥发物介导的昆虫化学通讯手段是防治该害虫的新途径。本研究以白刺粗角叶甲为对象,通过分子克隆技术获得白刺粗角叶甲成虫触角的非典型气味受体(atypical odorant receptor co-receptor,Orco)基因序列;利用同源建模方法预测蛋白三级结构,并构建系统发育树;借助荧光定量PCR技术检测DrybOrco基因在叶甲成虫各组织中的表达量差异。结果显示:克隆获得的白刺粗角叶甲非典型气味受体基因DrybOrco的cDNA全长为1918 bp,其中开放阅读框为1440 bp,编码479个氨基酸,蛋白分子量为53.94 kD,含有7个跨膜结构域,为疏水性膜蛋白。系统发育表明,Orco基因在6目68种昆虫间保守性较高(相似度大于68%),聚类分为3个分支,相同目的昆虫聚集到同一支。白刺粗角叶甲与其他鞘翅目昆虫聚为一支,其中与玉米根萤叶甲的Orco基因亲缘关系最近,核酸相似度高达92.28%。荧光定量表明,白刺粗角叶甲DrybOrco基因在成虫触角中特异性表达,且雄虫显著高于雌虫,表达量比值为2.27;此外,该基因在雄虫足和雌虫翅中少量表达,其他组织中几乎不表达。本研究明确了白刺粗角叶甲非典型气味受体基因DrybOrco的序列特征、蛋白结构和组织表达情况,这为阐明DrybOrco在该害虫化学通讯过程中的生理功能,以及探究白刺粗角叶甲寄主专化的嗅觉分子机制研究提供重要依据。

靶向嗅觉蛋白的植物精油源潜在昆虫行为控制剂研究进展

昆虫具有敏锐且复杂的嗅觉系统,这一系统的运行离不开多种嗅觉蛋白的参与。嗅觉蛋白可以通过对外界气味分子的捕获、运输、识别和降解,进而调控昆虫的取食、交配、繁殖、报警等一系列生理行为。本文综述了昆虫嗅觉蛋白中的气味结合蛋白(OBPs)和嗅觉受体(ORs)在调控昆虫行为上的功能及其晶体结构研究进展,并根据已报道的气味结合蛋白或嗅觉受体与配体小分子的共晶结构,重点分析了二者的互作模式。同时以植物精油为代表,介绍了其在调控昆虫行为活性方面的研究进展,期望为以昆虫气味结合蛋白或嗅觉受体为潜在靶标,聚焦新颖的天然植物精油源先导结构,开发新型的昆虫行为控制剂提供参考。