我们感谢约阿希姆·埃伯向我们介绍的触觉学习模式，蜜蜂。这项工作是由群集支持的卓越277 CITEC，资助在德国卓越计划的框架。

1。准备蜜蜂<ol><li>花蜜或花粉觅食，被发现在该领域无论是从蔗糖进纸器或直接从蜂巢入口，而返回的觅食之旅。每一个单一的蜜蜂被捕获到的玻璃小瓶，用泡沫插件进入实验室作进一步处理，并立即采取关闭。 </li><li>在实验室中，捕获的蜜蜂进行了简要冷却冰箱在4°C不动，直到他们首先显示的迹象。 </li><li>每个单一的固定化蜜蜂被安装在一个小的金属管与粘合带之间的头部和胸部和腹部上。长鼻和天线是可自由移动的，应小心。 </li><li>油漆的固定用白色油漆的蜜蜂（ 例如无溶剂TIPP-EX），以堵塞视觉的复眼和单眼。 </li><li>加一小滴融化的蜡头后面的蜜蜂来解决它在录音磁带头部和胸部之间，以防止头部运动。 </li><li>一些在磁带上，更好地识别标记每个单蜜蜂和将管子固定的蜜蜂到一个潮湿的环境，以防止脱水。 </li><li>互相喂单一的蜜蜂，持续5秒的30％的蔗糖溶液，用注射器的液滴，并让所有蜜蜂恢复与触觉的调理协议开始之前30分钟。 </li></ol> 2。轻触式空调<ol><li>在调节之前，每个单个的蜜蜂将测试其长鼻扩展响应（PER）到30％蔗糖的刺激施加到的触角。由此，将吸管尖端具有交叉的虚拟线之间的打开的下颌骨。丢弃所有的蜜蜂不响应的PER蔗糖刺激。 </li><li>对于触觉空调使用的黄铜多维数据集（ 例如，3×5毫米）与平稳或刻图案， 例如 ，水平或垂直槽形成的光栅150微米波长，作为条件刺激（CS）。 F或非条件刺激（US）使用了30％的蔗糖溶液（户糖，在水中稀释）。 </li><li>黄铜立方体（CS）被放置到一个保持器上显微操作（ 例如 MM33Märzhäuser）的，以确保在调节过程中的精确定位。美国提出了填充用30％的蔗糖溶液，用注射器的蜜蜂。 </li><li>的调理过程包括五个审间5分钟的时间间隔（ITI）的触觉刺激（CS）和蔗糖溶液（US）的配对。将前面的显微操作器与所安装的触觉刺激（CS）中的一个单一的蜜蜂。位置CS缓慢，使得触觉刺激的表面是平行的蜜蜂的头部（ 图1中A和 B）。动物和触觉对象之间的距离应该是在测试蜜蜂触角的工作半径的范围内， 即在一个舒适的位置w的蜜蜂应该是能够扫描的触觉刺激第i个两个天线。在图1中所示的例子中，距离是3毫米。让蜜蜂扫描触觉刺激（CS），持续5秒。后的第3秒内，呈现出30％的蔗糖溶液（US）用注射器下长鼻液滴。使用的注射器的前端轻轻抬起长鼻。蔗糖刺激下的长鼻将引起每15个非条件。让蜜蜂舔蔗糖的奖励。使用秒表与一个报警信号，以维持精确的时间间隔期间空调。 </li><li>空调PER的用作蜜蜂学习成功的量度。蜜蜂开始后的第一个奖励配对，以应对CS演示文稿延长他们的长鼻，这表明他们期待即将出台的奖励。完全伸出长鼻过程中随时观察到3秒的时间窗口之前的触觉刺激呈现和蔗糖介绍，取得了积极的响应。无响应计数负面的。在OCC的PER的urrence必须注意由实验者。每次试验的比例绘制蜜蜂的市盈率在CS介绍。 </li></ol> 3。运动录音<ol><li>一个单一的驾驭蜜蜂触角变动记录有数字视频相机透镜（ 如巴斯勒A602f-2配备技术规格VZM 200的操作，在50 fps的通过一个防火电线连接）与一个合适的宏。是相机处于上方的动物中的自顶向下的视图（ 图1 A）。在图1中，在录音的空间分辨率为0.02毫米/像素。 </li><li>校准摄像头，通过记录一个10×10毫米的棋盘与从不同方位下的摄像头的目标的边缘的长度为1毫米的单的照片。校准可以用的摄像机标定工具箱Matlab的16。 </li><li>下面的相机镜头放置一个固定的蜜蜂。目前的触觉刺激，固定显微操作，给动物。继续以同样的方式所描述的触觉空调，和记录触角运动而蜜蜂被扫描对象。充分触角的工作范围和触觉的刺激，这是很重要的是可见的。刺激模式的选择，测试和，试验数字取决于实验的设计。 </li></ol> 4。数据分析<ol><li>在Matlab计算图像的背景。首先，随着时间的推移的平均灰度值，要对每一个像素计算。静态对象，如固定头的蜜蜂和触觉刺激，将构成图像的背景。移动的物体，如两个天线，不会成为背景的一部分。 </li><li>所记录的视频的每个帧的顺序加载，和当前帧的图像的背景之间的差异具有以被计算。相减的结果，强调的移动从帧到帧的图像中的那些部分。编号两坳一隆一斜坡，触角的蜜蜂的非零值（ 图1C）的唯一区域。 </li><li>进行进一步的处理，两个最大的非零象素值的区域被假定为与天线。要产生含有对每一个像素的值1，如果像素属于天线和值，否则为0，对于这两种天线的二进制掩码。这个面膜是为本土化触角提示后，以此为基础。要获得初步的掩模对整个图像的差分图像的各像素的灰度值进行比较，以一个预先定义的阈值。因为我们照顾的噪音阈值选择是相当低的。初步面膜之后，还进行两种类型的错误:第一，由于图像噪声小区域仍有部分的面膜。其次，属于天线的领域，可能不一定完全连接。后者多发生，如果背景具有大致相同的发光作为天线。为了消除这些文物，图像处理形态学操作应用， 即图像的侵蚀和扩张相结合，依赖于标准的Matlab的功能imerode和imdilate（见17页158-205作进一步的解释）。经过去噪的过程中，天线的假设仍然包含在相同的面具。因此，作为下一个步骤的二进制掩码集中不相交的区域，使用标准的Matlab:功能bwlabel（的图像分割算法的进一步详情，请参阅17页40-48）。 </li><li>在每个群集的像素的数目进行计数，并选择两个最大的群集。重心计算区分之间的左和右的天线（ 图1D）。 </li><li>群集的近端到远端的方向（参见图1D）中的最高值的像素在一个可以被定义为为每个天线的触角小费。 </li></ol>

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没有利益冲突的声明。

蜜蜂制备:收集和固定的蜜蜂应该迅速完成，以保持动物的低应力水平。应激反应的影响，因此，可能有间接影响的学习表现在蜜蜂19，20。的应力水平可以降低由放置玻璃小瓶，与直接在冰上的蜜蜂采集后立即迅速固定它们。必须加以考虑，蜜蜂需要更多的时间来恢复麻醉期间之前的时间越长。 视觉触觉学习有负面影响，在蜜蜂18。因此，重要的...

<table border="1"><tbody><tr><td> 的试剂/材料名称 </td><td> 公司 </td><td> 目录编号 </td><td> 评论 </td></tr><tr><td>花粉的蜜蜂觅食，L. </td><td></td><td></td><td>私人荨麻疹</td></tr><tr><td>玻璃小瓶中（22×45毫米） </td><td>卡尔·罗斯GMBH＆CO </td><td> X655.1 </td><td></td></tr><tr><td>铜管的，10×30毫米</td><td></td><td></td><td>自产</td></tr><tr><td>铜立方体，3 x 5毫米</td><td></td><td></td><td>自产</td></tr><tr><td> TIPP-EX（无溶剂） </td><td> TIPP-EX GMBH＆CO KG </td><td></td><td></td></tr><tr><td>蔗糖溶液（30％） </td><td></td><td></td><td>家居糖用水稀释</td></tr><tr><td>乙醇（70％） </td><td>卡尔·罗斯GMBH＆CO </td><td> 9065.3 &lt;/ TD&gt; <td></td></tr><tr><td>注射器（1ml）和针（0.6×25） </td><td>卡尔·罗斯GMBH＆CO </td><td> 59.1 </td><td></td></tr><tr><td>秒表</td><td>卡尔·罗斯GMBH＆CO </td><td> L423.1 </td><td></td></tr><tr><td>微操作机器人</td><td> MärzhäuserMM33 </td><td> 00-42-101-0000 </td><td></td></tr><tr><td>数码摄像机</td><td>巴斯勒A602f-2 </td><td></td><td></td></tr><tr><td>微距镜头相机</td><td>技术规格VZM 200 </td><td></td><td></td></tr><tr><td> MATLAB R2009b </td><td> The MathWorks公司</td><td></td><td></td></tr></tbody></table>

tactile conditioning and movement analysis of antennal sampling strategies in honey bees (apis mellifera l.)

蜜蜂（ 蜜蜂 L.）是典型的社会性昆虫，其复杂的劳动分工和联想学习能力1，2众所周知的。工蜂花上半年在昏暗的蜂巢，他们的生活，他们是护士的幼虫或建设正六边形梳子食品（ 如花粉或花蜜）和育雏3。非凡的多感官触角，触角和各种触觉介导的任务，包括蜂巢5号楼和模式识别6中发挥了举足轻重的作用。以后在生活中，每个单一蜜蜂离开蜂巢饲料，用于食品。然后，一只蜜蜂学习歧视有利可图的食物来源，记住它们的位置，并将其传达到它的巢队友7。蜜蜂用不同的花卉信号，如颜色或气味，7，8，但也有触觉线索，从花瓣表面形成多感官记忆的食物来源。在实验室的合作的条件，蜜蜂可以训练，学习模式食欲，区分触觉的物体的特征，如边缘或沟槽，其触角10日，11日，12日，13。这种学习模式是密切相关的经典嗅觉空调的长鼻延伸反应（PER）的利用，蜜蜂14。在实验室中的触觉学习范式的优点是将与各种生理测量，包括触角的运动模式的分析，对学习行为实验相结合的可能性。

在下面的实验中，我们研究了，触觉学习如何在蜜蜂的行为影响触角采样。对于这一点，我们监测的天真和空调蜜蜂触角尖端之前和期间提交的触觉刺激的运动。 首先，自发触角花粉觅食一组（N = 42）的运动记录，持续1分钟。蜜蜂的二分之一（N = 21），然后，空调的配对5次用30％的蔗糖的奖励的触觉刺激。这是空调基。触觉刺激是30×50毫米的黄铜表面的立方体，一个刻有的?...

Watch this Scientific Journal Video about Tactile Conditioning And Movement Analysis Of Antennal Sampling Strategies In Honey Bees Apis mellifera L. at JoVE.com

Tactile Conditioning And Movement Analysis Of Antennal Sampling Strategies In Honey Bees Apis mellifera L.

在这个协议中，我们展示了如何驾驭的条件蜜蜂的触觉刺激和引进的2D动作捕捉技术的运动学分析精细尺度的触角采样模式。

触觉空调和取样策略在蜜蜂触角的运动分析（<em&gt;蜜蜂</em&gt; L.）

Honey bees (Apis mellifera L.) are eusocial insects and well known for their complex division of labor and associative learning capability1, 2. The worker ...

tactile-conditioning-movement-analysis-antennal-sampling-strategies

Research

JoVE Journal

Neuroscience

Tactile Conditioning And Movement Analysis Of Antennal Sampling Strategies In Honey Bees (Apis mellifera L.)

10.5K 次观看.  Bielefeld University. 在这个协议中，我们展示了如何驾驭的条件蜜蜂的触觉刺激和引进的2D动作捕捉技术的运动学分析精细尺度的触角采样模式。

视频: Tactile Conditioning And Movement Analysis Of Antennal Sampling Strategies In Honey Bees Apis mellifera L.

Eye Movement Monitoring of Memory

Explicit (often verbal) reports are typically used to investigate memory (e.g. "Tell me what you remember about the person you saw at the bank yesterday."), however such reports can often be unreliable or sensitive to response bias 1, and may be unobtainable in some participant populations. Furthermore, explicit reports only reveal when information has reached consciousness and cannot comment on when memories were accessed during processing, regardless of whether the information is subsequently accessed in a conscious manner. Eye movement monitoring (eye tracking) provides a tool by which memory can be probed without asking participants to comment on the contents of their memories, and access of such memories can be revealed on-line 2,3. Video-based eye trackers (either head-mounted or remote) use a system of cameras and infrared markers to examine the pupil and corneal reflection in each eye as the participant views a display monitor. For head-mounted eye trackers, infrared markers are also used to determine head position to allow for head movement and more precise localization of eye position. Here, we demonstrate the use of a head-mounted eye tracking system to investigate memory performance in neurologically-intact and neurologically-impaired adults. Eye movement monitoring procedures begin with the placement of the eye tracker on the participant, and setup of the head and eye cameras. Calibration and validation procedures are conducted to ensure accuracy of eye position recording. Real-time recordings of X,Y-coordinate positions on the display monitor are then converted and used to describe periods of time in which the eye is static (i.e. fixations) versus in motion (i.e., saccades). Fixations and saccades are time-locked with respect to the onset/offset of a visual display or another external event (e.g. button press). Experimental manipulations are constructed to examine how and when patterns of fixations and saccades are altered through different types of prior experience. The influence of memory is revealed in the extent to which scanning patterns to new images differ from scanning patterns to images that have been previously studied 2, 4-5. Memory can also be interrogated for its specificity; for instance, eye movement patterns that differ between an identical and an altered version of a previously studied image reveal the storage of the altered detail in memory 2-3, 6-8. These indices of memory can be compared across participant populations, thereby providing a powerful tool by which to examine the organization of memory in healthy individuals, and the specific changes that occur to memory with neurological insult or decline 2-3, 8-10.

Eye movement monitoring (or eye tracking) reveals where in space the eyes linger, when and for how long. Here, we demonstrate how eye tracking can be used to investigate the integrity of memory in multiple participant populations, without requiring verbal, or otherwise explicit, reports.

眼运动监测内存

Explicit (often verbal) reports are typically used to investigate memory (e.g. "Tell me what you remember about the person you saw at the bank ...

科研

神经科学

Behavioural Pharmacology in Classical Conditioning of the Proboscis Extension Response in Honeybees (Apis mellifera)

Honeybees (Apis mellifera) are well known for their communication and orientation skills and for their impressive learning capability1,2. Because the survival of a honeybee colony depends on the exploitation of food sources, forager bees learn and memorize variable flower sites as well as their profitability. Forager bees can be easily trained in natural settings where they forage at a feeding site and learn the related signals such as odor or color. Appetitive associative learning can also be studied under controlled conditions in the laboratory by conditioning the proboscis extension response (PER) of individually harnessed honeybees3,4. This learning paradigm enables the study of the neuronal and molecular mechanisms that underlie learning and memory formation in a simple and highly reliable way5-12. A behavioral pharmacology approach is used to study molecular mechanisms. Drugs are injected systemically to interfere with the function of specific molecules during or after learning and memory formation13-16.
Here we demonstrate how to train harnessed honeybees in PER conditioning and how to apply drugs systemically by injection into the bee flight muscle.

Behavioural Pharmacology in Classical Conditioning of the Proboscis Extension Response in Honeybees Apis mellifera

We demonstrate how to implement a behavioral pharmacology method in an appetitive olfactory conditioning paradigm in honeybees (Apis mellifera) by systemic application of drugs. This method allows investigation of the mechanisms underlying learning and memory formation in a simple and reliable way.

在蜜蜂的长鼻扩展响应的经典条件反射行为药理学（蜜蜂）

Honeybees (Apis mellifera) are well known for their communication and orientation skills and for their impressive learning capability1,2. Because the ...

Automated Sholl Analysis of Digitized Neuronal Morphology at Multiple Scales

Neuronal morphology plays a significant role in determining how neurons function and communicate1-3. Specifically, it affects the ability of neurons to receive inputs from other cells2 and contributes to the propagation of action potentials4,5. The morphology of the neurites also affects how information is processed. The diversity of dendrite morphologies facilitate local and long range signaling and allow individual neurons or groups of neurons to carry out specialized functions within the neuronal network6,7. Alterations in dendrite morphology, including fragmentation of dendrites and changes in branching patterns, have been observed in a number of disease states, including Alzheimer's disease8, schizophrenia9,10, and mental retardation11. The ability to both understand the factors that shape dendrite morphologies and to identify changes in dendrite morphologies is essential in the understanding of nervous system function and dysfunction.
Neurite morphology is often analyzed by Sholl analysis and by counting the number of neurites and the number of branch tips. This analysis is generally applied to dendrites, but it can also be applied to axons. Performing this analysis by hand is both time consuming and inevitably introduces variability due to experimenter bias and inconsistency. The Bonfire program is a semi-automated approach to the analysis of dendrite and axon morphology that builds upon available open-source morphological analysis tools. Our program enables the detection of local changes in dendrite and axon branching behaviors by performing Sholl analysis on subregions of the neuritic arbor. For example, Sholl analysis is performed on both the neuron as a whole as well as on each subset of processes (primary, secondary, terminal, root, etc.) Dendrite and axon patterning is influenced by a number of intracellular and extracellular factors, many acting locally. Thus, the resulting arbor morphology is a result of specific processes acting on specific neurites, making it necessary to perform morphological analysis on a smaller scale in order to observe these local variations12.
The Bonfire program requires the use of two open-source analysis tools, the NeuronJ plugin to ImageJ and NeuronStudio. Neurons are traced in ImageJ, and NeuronStudio is used to define the connectivity between neurites. Bonfire contains a number of custom scripts written in MATLAB (MathWorks) that are used to convert the data into the appropriate format for further analysis, check for user errors, and ultimately perform Sholl analysis. Finally, data are exported into Excel for statistical analysis. A flow chart of the Bonfire program is shown in Figure 1.

We have developed a computer program to analyze neuronal morphology. In combination with two existing open source analysis tools, our program performs Sholl analysis and determines the number of neurites, branch points, and neurite tips. The analyses are performed so that local changes in neurite morphology can be observed.

肖尔自动数字化的神经元形态分析，多尺度

Neuronal morphology plays a significant role in determining how neurons function and communicate1-3.  Specifically, it affects the ability of neurons to ...

Coherence between Brain Cortical Function and Neurocognitive Performance during Changed Gravity Conditions

Previous studies of cognitive, mental and/or motor processes during short-, medium- and long-term weightlessness have only been descriptive in nature, and focused on psychological aspects. Until now, objective observation of neurophysiological parameters has not been carried out - undoubtedly because the technical and methodological means have not been available -, investigations into the neurophysiological effects of weightlessness are in their infancy (Schneider et al. 2008).
While imaging techniques such as positron emission tomography (PET) and magnetic resonance imaging (MRI) would be hardly applicable in space, the non-invasive near-infrared spectroscopy (NIRS) technique represents a method of mapping hemodynamic processes in the brain in real time that is both relatively inexpensive and that can be employed even under extreme conditions. The combination with electroencephalography (EEG) opens up the possibility of following the electrocortical processes under changing gravity conditions with a finer temporal resolution as well as with deeper localization, for instance with electrotomography (LORETA).
Previous studies showed an increase of beta frequency activity under normal gravity conditions and a decrease under weightlessness conditions during a parabolic flight (Schneider et al. 2008a+b). Tilt studies revealed different changes in brain function, which let suggest, that changes in parabolic flight might reflect emotional processes rather than hemodynamic changes. However, it is still unclear whether these are effects of changed gravity or hemodynamic changes within the brain. Combining EEG/LORETA and NIRS should for the first time make it possible to map the effect of weightlessness and reduced gravity on both hemodynamic and electrophysiological processes in the brain. Initially, this is to be done as part of a feasibility study during a parabolic flight. Afterwards, it is also planned to use both techniques during medium- and long-term space flight.
It can be assumed that the long-term redistribution of the blood volume and the associated increase in the supply of oxygen to the brain will lead to changes in the central nervous system that are also responsible for anaemic processes, and which can in turn reduce performance (De Santo et al. 2005), which means that they could be crucial for the success and safety of a mission (Genik et al. 2005, Ellis 2000).
Depending on these results, it will be necessary to develop and employ extensive countermeasures. Initial results for the MARS500 study suggest that, in addition to their significance in the context of the cardiovascular and locomotor systems, sport and physical activity can play a part in improving neurocognitive parameters. Before this can be fully established, however, it seems necessary to learn more about the influence of changing gravity conditions on neurophysiological processes and associated neurocognitive impairment.

The effect of weightlessness and hypergravity on both hemodynamic and electrophysiological processes in the brain is going to be followed during parabolic flight by EEG and NIRS techniques. 
A feasibility study of a more complex experiment, which is planned to carry out during medium- and long-term space flight.

大脑皮质功能和神经在更改重力条件下的性能之间的一致性

Previous studies of cognitive, mental and/or motor processes during short-, medium- and long-term weightlessness have only been descriptive in nature, and ...

Calcium Imaging of Odor-evoked Responses in the Drosophila Antennal Lobe

The antennal lobe is the primary olfactory center in the insect brain and represents the anatomical and functional equivalent of the vertebrate olfactory bulb1-5. Olfactory information in the external world is transmitted to the antennal lobe by olfactory sensory neurons (OSNs), which segregate to distinct regions of neuropil called glomeruli according to the specific olfactory receptor they express. Here, OSN axons synapse with both local interneurons (LNs), whose processes can innervate many different glomeruli, and projection neurons (PNs), which convey olfactory information to higher olfactory brain regions.
Optical imaging of the activity of OSNs, LNs and PNs in the antennal lobe - traditionally using synthetic calcium indicators (e.g. calcium green, FURA-2) or voltage-sensitive dyes (e.g. RH414) - has long been an important technique to understand how olfactory stimuli are represented as spatial and temporal patterns of glomerular activity in many species of insects6-10. Development of genetically-encoded neural activity reporters, such as the fluorescent calcium indicators G-CaMP11,12 and Cameleon13, the bioluminescent calcium indicator GFP-aequorin14,15, or a reporter of synaptic transmission, synapto-pHluorin16 has made the olfactory system of the fruitfly, Drosophila melanogaster, particularly accessible to neurophysiological imaging, complementing its comprehensively-described molecular, electrophysiological and neuroanatomical properties2,4,17. These reporters can be selectively expressed via binary transcriptional control systems (e.g. GAL4/UAS18, LexA/LexAop19,20, Q system21) in defined populations of neurons within the olfactory circuitry to dissect with high spatial and temporal resolution how odor-evoked neural activity is represented, modulated and transformed22-24.
Here we describe the preparation and analysis methods to measure odor-evoked responses in the Drosophila antennal lobe using G-CaMP25-27. The animal preparation is minimally invasive and can be adapted to imaging using wide-field fluorescence, confocal and two-photon microscopes.

Calcium Imaging of Odor-evoked Responses in the Drosophila Antennal Lobe

We describe an established technique to measure and analyze odor-evoked calcium responses in the antennal lobe of living Drosophila melanogaster.

在气味诱发反应的钙成像果蝇触角叶

The antennal lobe is the primary olfactory center in the insect brain and represents the anatomical and functional equivalent of the vertebrate olfactory ...

Electrophysiological Measurements and Analysis of Nociception in Human Infants

Pain is an unpleasant sensory and emotional experience. Since infants cannot verbally report their experiences, current methods of pain assessment are based on behavioural and physiological body reactions, such as crying, body movements or changes in facial expression. While these measures demonstrate that infants mount a response following noxious stimulation, they are limited: they are based on activation of subcortical somatic and autonomic motor pathways that may not be reliably linked to central sensory processing in the brain. Knowledge of how the central nervous system responds to noxious events could provide an insight to how nociceptive information and pain is processed in newborns.
The heel lancing procedure used to extract blood from hospitalised infants offers a unique opportunity to study pain in infancy. In this video we describe how electroencephalography (EEG) and electromyography (EMG) time-locked to this procedure can be used to investigate nociceptive activity in the brain and spinal cord.
This integrative approach to the measurement of infant pain has the potential to pave the way for an effective and sensitive clinical measurement tool.

The assessment and treatment of pain in infants is difficult because infants cannot verbally report their experience. In this video we describe quantitative electrophysiological methods and analysis techniques that can be used to measure the response to noxious events from the infant nervous system.

在人类婴儿的伤害性电生理测量和分析

Pain is an unpleasant sensory and emotional experience. Since infants cannot verbally report their experiences, current methods of pain assessment are ...

Rapid Determination of the Thermal Nociceptive Threshold in Diabetic Rats

Painful diabetic neuropathy (PDN) is characterized by hyperalgesia i.e., increased sensitivity to noxious stimulus, and allodynia i.e., hypersensitivity to normally innocuous stimuli1. Hyperalgesia and allodynia have been studied in many different rodent models of diabetes mellitus2. However, as stated by B&ouml;lcskei et al, determination of "pain" in animal models is challenging due to its subjective nature3. Moreover, the traditional methods used to determine behavioral responses to noxious thermal stimuli usually lack reproducibility and pharmacological sensitivity3. For instance, by using the hot-plate method of Ankier4, flinch, withdrawal and/or licking of either hind- and/or fore-paws is quantified as reflex latencies at constant high thermal stimuli (52-55 &deg;C). However, animals that are hyperalgesic to thermal stimulus do not reproducibly show differences in reflex latencies using those supra-threshold temperatures3,5. As the recently described method of B&ouml;lcskei et al.6, the procedures described here allows for the rapid, sensitive and reproducible determination of thermal nociceptive thresholds (TNTs) in mice and rats. The method uses slowly increasing thermal stimulus applied mostly to the skin of mouse/rat plantar surface. The method is particularly sensitive to study anti-nociception during hyperalgesic states such as PDN. The procedures described bellow are based on the ones published in detail by Alm&aacute;si et al 5 and B&ouml;lcskei et al 3. The procedures described here have been approved the Laboratory Animal Care and Use Committee (LACUC), Wright State University.

Here, we describe a rapid reliable and simple procedure to determine the lowest temperature at which rats or mice show nocifensive behavior, i.e. the thermal nociceptive threshold (TNT). This method applies a slowly increasing thermal stimulus allowing precise and reproducible estimation of TNTs with minimum, if any, stress to the animals.

糖尿病大鼠的热痛觉阈值的快速测定

Painful diabetic neuropathy (PDN) is characterized by hyperalgesia i.e., increased sensitivity to noxious stimulus, and allodynia i.e., hypersensitivity ...

RNAi-mediated Double Gene Knockdown and Gustatory Perception Measurement in Honey Bees (Apis mellifera)

This video demonstrates novel techniques of RNA interference (RNAi) which downregulate two genes simultaneously in honey bees using double-stranded RNA (dsRNA) injections. It also presents a protocol of proboscis extension response (PER) assay for measuring gustatory perception.	
	RNAi-mediated gene knockdown is an effective technique downregulating target gene expression. This technique is usually used for single gene manipulation, but it has limitations to detect interactions and joint effects between genes. In the first part of this video, we present two strategies to simultaneously knock down two genes (called double gene knockdown). We show both strategies are able to effectively suppress two genes, vitellogenin (vg) and ultraspiracle (usp), which are in a regulatory feedback loop. This double gene knockdown approach can be used to dissect interrelationships between genes and can be readily applied in different insect species.	
	The second part of this video is a demonstration of proboscis extension response (PER) assay in honey bees after the treatment of double gene knockdown. The PER assay is a standard test for measuring gustatory perception in honey bees, which is a key predictor for how fast a honey bee's behavioral maturation is. Greater gustatory perception of nest bees indicates increased behavioral development which is often associated with an earlier age at onset of foraging and foraging specialization in pollen. In addition, PER assay can be applied to identify metabolic states of satiation or hunger in honey bees. Finally, PER assay combined with pairing different odor stimuli for conditioning the bees is also widely used for learning and memory studies in honey bees.

RNAi-mediated Double Gene Knockdown and Gustatory Perception Measurement in Honey Bees Apis mellifera

In this protocol, we describe two strategies that simultaneously suppress two genes (double gene knockdown) in honey bees. Then we present how to use the proboscis extension response (PER) assay to study the effect of double gene knockdown on honey bee gustatory perception.

RNAi介导的双基因敲除和味觉感知测量蜜蜂（<em&gt;蜜蜂</em&gt;）

This video demonstrates novel techniques of RNA  interference (RNAi) which downregulate two genes simultaneously in honey bees  using double-stranded RNA ...

Neuropharmacological Manipulation of Restrained and Free-flying Honey Bees, Apis mellifera

Honey bees demonstrate astonishing learning abilities and advanced social behavior and communication. In addition, their brain is small, easy to visualize and to study. Therefore, bees have long been a favored model amongst neurobiologists and neuroethologists for studying the neural basis of social and natural behavior. It is important, however, that the experimental techniques used to study bees do not interfere with the behaviors being studied. Because of this, it has been necessary to develop a range of techniques for pharmacological manipulation of honey bees. In this paper we demonstrate methods for treating restrained or free-flying honey bees with a wide range of pharmacological agents. These include both noninvasive methods such as oral and topical treatments, as well as more invasive methods that allow for precise drug delivery in either systemic or localized fashion. Finally, we discuss the advantages and disadvantages of each method and&#160;describe common hurdles and how to best overcome them. We conclude with a discussion on the importance of adapting the experimental method to the biological questions rather than the other way around.

Neuropharmacological Manipulation of Restrained and Free-flying Honey Bees, Apis mellifera

This manuscript describes several protocols for administering pharmacological agents to honey bees, including simple noninvasive methods for free-flying bees, as well as more invasive variants that allow precise localized treatment of restrained bees.

约束和自由飞行，蜜蜂的神经药理学操纵，<em&gt;蜜蜂</em

Honey bees demonstrate astonishing learning abilities and advanced social behavior and communication. In addition, their brain is small, easy to visualize ...

The Use of Trace Eyeblink Classical Conditioning to Assess Hippocampal Dysfunction in a Rat Model of Fetal Alcohol Spectrum Disorders

Neonatal rats were administered a relatively high concentration of ethyl alcohol (11.9% v/v) during postnatal days 4-9, a time when the fetal brain undergoes rapid organizational change and is similar to accelerated brain changes that occur during the third trimester in humans. This model of fetal alcohol spectrum disorders (FASDs) produces severe brain damage, mimicking the amount and pattern of binge-drinking that occurs in some pregnant alcoholic mothers. We describe the use of trace eyeblink classical conditioning (ECC), a higher-order variant of associative learning, to assess long-term hippocampal dysfunction that is typically seen in alcohol-exposed adult offspring. At 90 days of age, rodents were surgically prepared with recording and stimulating electrodes, which measured electromyographic (EMG) blink activity from the left eyelid muscle and delivered mild shock posterior to the left eye, respectively. After a 5 day recovery period, they underwent 6 sessions of trace ECC to determine associative learning differences between alcohol-exposed and control rats. Trace ECC is one of many possible ECC procedures that can be easily modified using the same equipment and software, so that different neural systems can be assessed. ECC procedures in general, can be used as diagnostic tools for detecting neural pathology in different brain systems and different conditions that insult the brain.

Trace eyeblink classical conditioning (ECC) was used to assess hippocampal-dependent associative learning in adult rats that were administered a high concentration (11.9% v/v) of alcohol during early neonatal brain development. In general, ECC procedures are sound diagnostic tools for detecting brain dysfunction across many psychological and biomedical settings.

使用的跟踪经典条件反射，评估胎儿酒精谱系障碍大鼠模型海马功能障碍

Neonatal rats were administered a relatively high concentration of ethyl alcohol (11.9% v/v) during postnatal days 4-9, a time when the fetal brain ...