Honeybees (Apis mellifera) learn to discriminate the smell of organic compounds from their respective deuterated isotopomers

The understanding of physiological and molecular processes underlying the sense of smell has made considerable progress during the past three decades, revealing the cascade of molecular steps that lead to the activation of olfactory receptor (OR) neurons. However, the mode of primary interaction of odorant molecules with the OR proteins within the sensory cells is still enigmatic. Two different concepts try to explain these interactions: the ‘odotope hypothesis’ suggests that OR proteins recognize structural aspects of the odorant molecule, whereas the ‘vibration hypothesis’ proposes that intra-molecular vibrations are the basis for the recognition of the odorant by the receptor protein. The vibration hypothesis predicts that OR proteins should be able to discriminate compounds containing deuterium from their common counterparts which contain hydrogen instead of deuterium. This study tests this prediction in honeybees (Apis mellifera) using the proboscis extension reflex learning in a differential conditioning paradigm. Rewarding one odour (e.g. a deuterated compound) with sucrose and not rewarding the respective analogue (e.g. hydrogen-based odorant) shows that honeybees readily learn to discriminate hydrogen-based odorants from their deuterated counterparts and supports the idea that intra-molecular vibrations may contribute to odour discrimination.     

Identification of a new contingency-based response in honey bees (Apis mellifera) through revision of the proboscis extension conditioning paradigm

The two experiments reported present new information in the area of classical conditioning experiments with honey bees. Experiment 1 establishes a single unconditioned stimulus (US) technique as a preferred technique for conditioning of the proboscis extension response. Experiment 1 further identifies a new head turn response which occurs when the standard compound US technique is used. Experiment 2 demonstrates that the newly identified head turn response is contingency-based and provides important new response to the repertoire of honey bee learning experiments.     

Evidence for instantaneous e-vector detection in the honeybee using an associative learning paradigm

Many insects use the polarization pattern of the sky for obtaining compass information during orientation or navigation. E-vector information is collected by a specialized area in the dorsal-most part of the compound eye, the dorsal rim area (DRA). We tested honeybees' capability of learning certain e-vector orientations by using a classical conditioning paradigm with the proboscis extension reflex. When one e-vector orientation (CS+) was associated with sugar water, while another orientation (CS-) was not rewarded, the honeybees could discriminate CS+ from CS-. Bees whose DRA was inactivated by painting did not learn CS+. When ultraviolet (UV) polarized light (350 nm) was used for CS, the bees discriminated CS+ from CS-, but no discrimination was observed in blue (442 nm) or green light (546 nm). Our data indicate that honeybees can learn and discriminate between different e-vector orientations, sensed by the UV receptors of the DRA, suggesting that bees can determine their flight direction from polarized UV skylight during foraging. Fixing the bees' heads during the experiments did not prevent learning, indicating that they use an 'instantaneous' algorithm of e-vector detection; that is, the bees do not need to actively scan the sky with their DRAs ('sequential' method) to determine e-vector orientation.     

ame-miR-124-3p对西方蜜蜂气味学习的影响

越来越多的证据表明miRNA在神经调控中起着重要的作用。本研究选取12日龄的西方蜜蜂Apis mellifera,探讨蜜蜂个体脑部ame-miR-124-3p对气味学习的功能研究。首先对11日龄的蜜蜂分别饲喂ame-miR-124-3P抑制剂及其无意义序列、ame-miR-124-3P的模拟物及其无意义序列。饲喂24 h后对4组蜜蜂进行气味联想性学习实验,随后通过伸吻反应(Proboscis extension response, PER)比较4组蜜蜂的嗅觉学习差异。最后采用实时荧光定量PCR(qRT-PCR)验证干扰结果。PER结果显示,饲喂ame-miR-124-3P抑制剂的蜜蜂学习比率显著降低,饲喂ame-miR-124-3P模拟物的蜜蜂学习比例显著升高。荧光定量PCR(qRT-PCR)检测结果显示ame-miR-124-3P的表达水平在饲喂ame-miR-124-3P抑制剂组蜜蜂脑部显著降低(P<0.05),饲喂ame-miR-124-3P模拟物组蜜蜂脑部显著升高(P<0.05)。研究发现,蜜蜂脑部ame-miR-124-3P受到抑制后,蜜蜂的气味学习能力显著性降低...     

微机控制的穿棱箱双向主动回避反应实验系统

目的：研究动物的学习与记忆功能。方法：研制一种微机控制的穿梭箱双向主动回避反应实验系统,自动分析和记录实验结果。结果：实验结果客观、准确。结论：本系统自动化程度高,操作简便,能满足实验需求     

Bumble Bees Show Asymmetrical Discrimination Between Two Odors in a Classical Conditioning Procedure

Olfactory processing of two odorants and their mixture was investigated in bumble bees Bombus terrestris using classical conditioning of the proboscis extension. In a standard procedure, workers were able to learn linalool, phenylacetaldehyde, and the mixture of these two components, with a similar level of response to these three stimuli. Thereafter, when we applied a differential conditioning procedure where one rewarded odorant was presented alternately against an unrewarded one, an asymmetrical discrimination between the two pure odors was found. Bumble bees performed well in the discriminative task when linalool was the rewarded stimulus and phenylacetaldehyde the unrewarded one, but they had difficulty learning phenylacetaldehyde if it was the rewarded odor in the symmetrical procedure. Indeed, unrewarded stimulations with linalool appeared to disrupt the learning of the alternative odor, possibly due to an innate biological meaning of linalool.     

5-羟甲基糠醛对意大利蜜蜂工蜂学习行为及头部学习相关基因表达的影响

5-羟甲基糠醛（HMF）是一种糖类物质加热变性后极易产生的物质,是特定糖类的降解产物,在蜂群饲养过程中若不规范操作则可能从蜜蜂饲料中产生这种物质。尽管其对哺乳动物的毒理效应已有明确研究,但关于HMF对蜜蜂神经毒性的探索仍十分有限,尤其在学习认知影响方面存在研究空白。本研究通过饲喂意大利蜜蜂Apis mellifera ligustica工蜂一定浓度的HMF并进行气味联想性学习测试受试蜜蜂的学习认知能力,目的在于探究HMF对蜜蜂的学习认知能力是否产生影响并在分子水平上进行验证,为此后实际养蜂生产和蜂群饲养管理中的HMF暴露提供风险评估。本试验从蜂群中抓取6日龄意大利蜜蜂并于恒温恒湿环境下笼养10 d。期间处理组工蜂自由取食含1 500 ng/μL HMF的30%（w/v）蔗糖溶液,对照组则自由取食30%蔗糖溶液。随后使用气味学习装置对16日龄工蜂进行伸吻反应（Proboscis extension response,PER）测试,并通过qRT-PCR对蜜蜂头部学习相关基因表达量进行差异分析。试验结果表明,处理组与对照组蜜蜂在10 d用药饲养过程中死亡率无显著差异（P>0.05）。PER结果则表明处理组蜜蜂伸吻率明显低于对照组,且差异显著（P<0.05）。处理组相较于对照组蜜蜂头部学习相关基因TpnC Ⅲa,OPN4,Obp 4和PRKACB显著下调表达;pkc则显著上调表达。综上结果表明,1 500 ng/μL HMF不会导致蜜蜂急性死亡,而蜜蜂学习能力会显著降低,同时影响蜜蜂头部学习相关基因的表达。     

亚致死剂量杀虫剂对意蜂工蜂嗅觉敏感性的影响

喙伸反应(PER)试验适用于评价杀虫剂对蜜蜂行为的影响。本实验利用喙伸反应研究了亚致死剂量(LD50/100～LD50/10)溴氰菊酯和吡虫啉对意蜂Apis mellifera ligustica L.工蜂嗅觉敏感性的影响。结果发现,经口饲喂溴氰菊酯5ng和10ng后,工蜂对0.1%的蔗糖溶液的敏感性显著下降(P<0.05),水应激指数降低,但对0.3%,1%,3%,10%和30%的蔗糖溶液的敏感性没有显著变化;而经口饲喂吡虫啉0.3ng和0.6ng后,工蜂对上述各浓度蔗糖溶液的敏感性变化不明显,但其水应激指数升高。     

Identification of neurons responsible for feeding behavior in the Drosophila brain

Drosophila melanogaster feeds mainly on rotten fruits,which contain many kinds of sugar.Thus,the sense of sweet taste has evolved to serve as a dominant regulator and driver of feeding behavior.Although several sugar receptors have been described,it remains poorly understood how the sensory input is transformed into an appetitive behavior.Here,we used a neural silencing approach to screen brain circuits,and identified neurons labeled by three Gal4 lines that modulate Drosophila feeding behavior.These three Gal4 lines labeled neurons mainly in the suboesophageal ganglia(SOG),which is considered to be the fly’s primary taste center.When we blocked the activity of these neurons,flies decreased their sugar consumption significantly.In contrast,activation of these neurons resulted in enhanced feeding behavior and increased food consumption not only towards sugar,but to an array of food sources.Moreover,upon neuronal activation,the flies demonstrated feeding behavior even in the absence of food,which suggests that neuronal activation can replace food as a stimulus for feeding behavior.These findings indicate that these Gal4-labeled neurons,which function downstream of sensory neurons and regulate feeding behavior towards different food sources is necessary in Drosophila feeding control.     

蜜蜂对复合气味中特征因素的选择性记忆巩固的研究

目的 蜜蜂天生具有丰富的嗅觉辨识能力,觅食、交配、导航以及社交活动均依赖其嗅觉系统,是研究嗅觉感知和学习记忆的行为及神经机制的理想模型。蜜蜂既能够将某个复合气味作为一个整体也可以将复合气味的各组成成分进行辨别和区分,但是在特征依赖的联合记忆中依据何种原则进行加工并存储到长期记忆还不清楚。方法 本文利用特征阳性（feature positive：AB+,B-）和特征阴性（feature negative：AB-,B+）的奖赏性嗅觉条件化,训练蜜蜂对复合气味和成分气味的辨别,并检测蜜蜂对复合气味（AB）、成分气味（B）以及特征气味（A）的中长时记忆（3 h）和长时记忆（24 h）。结果 在特征阳性的奖赏性嗅觉条件化中,蜜蜂对训练过的气味可以形成稳定的中长时和长时记忆,并且对复合气味中的特征气味的记忆与复合气味的记忆呈现高度相似。但在特征阴性的奖赏性嗅觉条件化中,蜜蜂虽能够在3 h和24 h对训练过的两种气味具有显著的伸喙反应差异,且对特征阴性的气味无显著反应,但对复合气味的反应随时间的推移而增加。结论 实验结果表明,蜜蜂选择性地将与奖赏信息联合出现的气味巩固到长时记忆中,但并未依据特征成分加工储存到长时记忆中。奖赏信息预示着食物源,与生存息息相关,表明对环境信息进行选择性的记忆巩固加工并储存可能是低等动物高效地编码生存相关信息的重要策略。     

Identification of feeding attractants in oak sap for adults of two nymphalid butterflies, Kaniska canace and Vanessa indica

The active compounds of oak‐sap odour in attracting adults of two butterflies, Kaniska canace (L.) and Vanessa indica (Herbst) (Lepidoptera: Nymphalidae), were identified by chemical analyses, electroantennogram (EAG) and two behavioural assays: proboscis extension reflex (PER) and attraction to artificial tree models. Fourteen compounds were identified from two sap samples collected in 1997 and 1998, of which the major volatiles were ethanol and acetic acid (≈ 900 p.p.m. and 500 p.p.m. in sap, respectively). However, the chemical composition of the minor volatiles varied considerably between the two samples. Among 13 chemicals tested, V. indica showed strong PER to five aliphatic acids (acetic, propionic, butyric, isobutyric and isovaleric), 2‐methylpropan‐1‐ol and 3‐hydroxybutan‐2‐one, whereas the PER‐active compounds for K. canace were these seven compounds and also ethanol, 3‐methylbutan‐1‐ol and 1‐hydroxypropan‐2‐one. In two‐choice behavioural bioassays, the model scented with a sap‐odour mimic, which was an aqueous mixture of the PER‐active compounds, was more attractive to the two butterflies than an unscented control. These results demonstrated that the sap odour stimulates foraging behaviour of the butterfly. Although EAG responses of both butterflies to 3‐methylbutan‐1‐ol and that of V. indica to 2‐methylpropan‐1‐ol were positively dose‐dependent, responses to other compounds were not strong and not dose‐dependent at 1–100 μg doses. These EAG responsiveness suggests that the olfactory receptors for these compounds might be few in the antenna and that the butterflies have enough olfactory sensitivity to the dose of 1 μg.     

桔小实蝇雄成虫联系性嗅觉学习

【目的】为了探究桔小实蝇 Bactrocera dorsalis (Hendel)雄成虫的嗅觉学习能力。【方法】本研究采用经典性嗅觉条件反射训练法（classical olfactory conditioning）在室内对固定的羽化后14-17日龄的桔小实蝇雄成虫进行气味与食物的联合学习训练, 即薄荷精油和10%蔗糖溶液联合的奖赏性训练（appetitive conditioning）以及甲基丁香酚(methyl eugenol, ME)和饱和盐溶液联合的惩罚性训练（aversive conditioning）,并以伸喙反射行为（proboscis extension reflex, PER）作为学习与否的判定标准。【结果】经过奖赏性训练后,桔小实蝇雄成虫对薄荷精油的伸喙反射率可从0%增加至68%;而经过惩罚性训练后,桔小实蝇对甲基丁香酚的伸喙反射率可从100%降低至36.54%,且这种伸喙反射率的变化是通过气味条件刺激（conditioned stimulus）和食物非条件刺激（unconditioned stimulus）的对称性联合而产生的。【结论】结果表明,桔小实蝇雄性成虫具有较强的联系性嗅觉学习能力,并且两种刺激的联合是形成学习记忆的必要条件。