Calcium imaging in the insect nervous system using an innovative dye loading technique
使用创新的染料加载技术对昆虫神经系统进行钙成像
基本信息
- 批准号:BB/T002085/1
- 负责人:
- 金额:$ 61.07万
- 依托单位:
- 依托单位国家:英国
- 项目类别:Research Grant
- 财政年份:2020
- 资助国家:英国
- 起止时间:2020 至 无数据
- 项目状态:已结题
- 来源:
- 关键词:
项目摘要
Neuroscience aims at understanding the organisation and function of nervous systems focussing on vertebrate and invertebrate model animals. Recording the activity of neurons is classically achieved with extra- or intracellular electrodes which measure the electrical spike and synaptic activity with high temporal and amplitude resolution. Over the last decades optical recordings have been developed, based e.g. on calcium sensitive dyes, which are loaded into neurons or populations of neurons. Upon binding to Calcium ions these dyes change their fluorescence intensity or wavelength and generate optical signals that can be monitored with sensitive cooled CCD cameras or confocal microscopes. As calcium is an essential molecule for neuronal functioning and synaptic processing the optical signals report the excitatory activity of the labelled neurons - although not with the precision of electrophysiological measurements. Optical imaging however, allows to analyse the spatial dimensions of neuronal processing at the cellular and network level and has substantially contributed to our understanding of nervous systems. In some model systems, like the fruit fly Drosophila or the zebrafish, even genetically encoded calcium indicators are available, which can be expressed in specific cell lines of the nervous system. The corresponding molecular-genetic tools are not (yet) available in other organisms, which nonetheless are model systems to study a specific behaviour. One of these system are acoustically communicating insects like crickets or bush-crickets, which for decades have been in the focus of neuroscience research aiming to understand the neuronal mechanisms underlying hearing, auditory pattern recognition and sound pattern generation. Progress in analysing these systems was mainly based on electrophysiological and neuroanatomical techniques, as loading central or afferent neurons with calcium sensitive dyes, is difficult and required loading these neurons with intracellular dye injection via microelectrodes. Inspired by the iontophoretic application of drugs through the intact human skin, we recently developed a dye delivery method through the intact sheath of nerves or ganglia. Glass capillaries with 40-80 micro-meter tip diameter are filled with the calcium sensitive tracer and are attached to the neuronal sheath e.g. of an insect auditory nerve. Initially they are used as recording electrodes, and once a good signal has been established the circuit is switched into iontophoretic dye delivery mode. The calcium sensitive dye moves into the axons of the nerve and then travels in both directions from the injection site, labelling the peripheral sensory organ and the central afferent axonal arborisations in the nervous system. No other labelling method can achieve this. The method can also be used to specifically label auditory neuropil regions in the brain of crickets. Our preliminary imaging experiments prove the principle and demonstrate that sound evoked optical signals report the specific activity of the labelled auditory afferents and central neurons. We now aim to further take advantage of this technique to systematically study (1.) the spatial organisation of auditory processing in the cricket brain; (2) the activation of auditory afferents in the hearing organ and (3) the spatiotemporal activity patterns underlying the singing motor activity in male crickets. Besides using our sensitive CCD camera system, we will use more advanced confocal and two-photon imaging systems to enhance the resolution of the data. The chosen objectives represent central questions in insect neuroscience and should contribute to further our understanding of insect acoustic communication. Moreover the planned experiments will demonstrate the versatility of our method as a new tool in insect neuroscience and will be relevant to the wider community of researchers.
神经科学旨在了解神经系统的组织和功能,重点是脊椎动物和无脊椎动物模型动物。记录神经元的活动通常是通过细胞外或细胞内电极来实现的,这些电极以高的时间和幅度分辨率测量电尖峰和突触活动。在过去的几十年中,已经开发了光学记录,例如基于钙敏感染料,其被加载到神经元或神经元群中。在与钙离子结合后,这些染料改变其荧光强度或波长,并产生可以用灵敏的冷却CCD相机或共聚焦显微镜监测的光学信号。由于钙是神经元功能和突触处理的重要分子,因此光学信号报告标记神经元的兴奋性活动-尽管不具有电生理测量的精度。然而,光学成像允许在细胞和网络水平上分析神经元处理的空间维度,并大大有助于我们对神经系统的理解。在一些模型系统中,如果蝇或斑马鱼,甚至可以获得遗传编码的钙指示剂,其可以在神经系统的特定细胞系中表达。相应的分子遗传学工具在其他生物体中还不可用,尽管如此,这些生物体是研究特定行为的模型系统。这些系统之一是声学通信昆虫,如蟋蟀或蟋蟀,几十年来一直是神经科学研究的焦点,旨在了解听觉,听觉模式识别和声音模式生成的神经机制。分析这些系统的进展主要基于电生理学和神经解剖学技术,因为用钙敏感染料加载中枢或传入神经元是困难的,并且需要通过微电极用细胞内染料注射加载这些神经元。受药物通过完整人体皮肤的离子电渗应用的启发,我们最近开发了一种通过神经或神经节的完整鞘的染料递送方法。具有40-80微米尖端直径的玻璃毛细管填充有钙敏感示踪剂,并附着于例如昆虫听神经的神经鞘。最初,它们被用作记录电极,一旦建立了良好的信号,电路就被切换到离子电渗染料输送模式。钙敏感染料移动到神经轴突中,然后从注射部位向两个方向行进,标记神经系统中的外周感觉器官和中枢传入轴突分支。没有其他的标签方法可以做到这一点。该方法还可以用于特异性标记蟋蟀大脑中的听觉神经元区域。我们初步的成像实验证明了这一原理,并表明,声音诱发的光信号报告标记的听觉传入和中枢神经元的特定活动。我们现在的目标是进一步利用这种技术来系统地研究(1)。蟋蟀大脑中听觉处理的空间组织;(2)听觉器官中听觉传入的激活;(3)雄性蟋蟀歌唱运动活动的时空活动模式。除了使用灵敏的CCD相机系统外,我们还将使用更先进的共聚焦和双光子成像系统来提高数据的分辨率。所选择的目标代表昆虫神经科学的中心问题,并应有助于进一步我们了解昆虫的声音通信。此外,计划中的实验将证明我们的方法作为昆虫神经科学中的新工具的多功能性,并将与更广泛的研究人员社区相关。
项目成果
期刊论文数量(2)
专著数量(0)
科研奖励数量(0)
会议论文数量(0)
专利数量(0)
Response properties of spiking and non-spiking brain neurons mirror pulse interval selectivity.
- DOI:10.3389/fncel.2022.1010740
- 发表时间:2022
- 期刊:
- 影响因子:5.3
- 作者:
- 通讯作者:
Sound processing in the cricket brain: evidence for a pulse duration filter.
- DOI:10.1152/jn.00252.2023
- 发表时间:2023-10-01
- 期刊:
- 影响因子:2.5
- 作者:
- 通讯作者:
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Berthold Hedwig其他文献
Evidence for a command neuron controlling calling song in the cricket emGryllus assimilis/em
蟋蟀(Gryllus assimilis)中控制鸣叫声的命令神经元的证据
- DOI:
10.1016/j.jinsphys.2025.104798 - 发表时间:
2025-04-01 - 期刊:
- 影响因子:2.300
- 作者:
Chu-Cheng Lin;Berthold Hedwig - 通讯作者:
Berthold Hedwig
Pulses, patterns and paths: neurobiology of acoustic behaviour in crickets
- DOI:
10.1007/s00359-006-0115-8 - 发表时间:
2006-03-08 - 期刊:
- 影响因子:2.200
- 作者:
Berthold Hedwig - 通讯作者:
Berthold Hedwig
Tonic signaling from O2 sensors sets neural circuit activity and behavioral state
来自氧气传感器的强化信号设定神经回路活动和行为状态
- DOI:
10.1038/nn.3061 - 发表时间:
2012-03-04 - 期刊:
- 影响因子:20.000
- 作者:
Karl Emanuel Busch;Patrick Laurent;Zoltan Soltesz;Robin Joseph Murphy;Olivier Faivre;Berthold Hedwig;Martin Thomas;Heather L Smith;Mario de Bono - 通讯作者:
Mario de Bono
El grillo è buon cantore: for Franz Huber on the occasion of his 90th birthday
- DOI:
10.1007/s00359-016-1074-3 - 发表时间:
2016-02-15 - 期刊:
- 影响因子:2.200
- 作者:
Berthold Hedwig - 通讯作者:
Berthold Hedwig
Phonotaxis of male field crickets, emGryllus bimaculatus/em, to conspecific calling song
雄性斗蟋(Gryllus bimaculatus)对同种鸣叫的趋音性行为
- DOI:
10.1016/j.anbehav.2023.08.010 - 发表时间:
2023-11-01 - 期刊:
- 影响因子:2.100
- 作者:
Adam Bent;Berthold Hedwig - 通讯作者:
Berthold Hedwig
Berthold Hedwig的其他文献
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{{ truncateString('Berthold Hedwig', 18)}}的其他基金
Auditory processing: The cellular and synaptic mechanisms of a delay-line and coincidence-detector circuit
听觉处理:延迟线和重合检测器电路的细胞和突触机制
- 批准号:
BB/P022111/1 - 财政年份:2017
- 资助金额:
$ 61.07万 - 项目类别:
Research Grant
RESUBMISSION: Neural processing underlying auditory pattern recognition in an insect brain
重新提交:昆虫大脑中听觉模式识别的神经处理
- 批准号:
BB/J01835X/1 - 财政年份:2013
- 资助金额:
$ 61.07万 - 项目类别:
Research Grant
Motor control of auditory steering in crickets
蟋蟀听觉转向的运动控制
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BB/G018723/1 - 财政年份:2009
- 资助金额:
$ 61.07万 - 项目类别:
Research Grant
Functional organisation of a corollary discharge mechanism
必然放电机制的功能组织
- 批准号:
BB/F008783/1 - 财政年份:2008
- 资助金额:
$ 61.07万 - 项目类别:
Research Grant
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