Name: functional analysis of the larval feeding circuit in drosophila
Uploaded: 2013-11-19T16:45:00
Description: Watch this Scientific Journal Video about Functional Analysis of the Larval Feeding Circuit in Drosophila at JoVE.com

저자는 WSN에 수여 세인트 루이스 대학에서 대통령의 연구 기금을 인정하고 싶습니다

1. 인구 케이지의 유지 보수 <ol><li> 12 시간의 명암주기에 25 ° C에서 인구의 새장을 유지한다. 한 컨트롤과 실험군은 동일 조명 조건에 노출 될 때,이 기술은 표준 실험실 설정에서 수행 될 수있다. </li><li> 여성은 사과 주스 한천 플레이트에서 하룻밤 알을 낳을 수 있도록 허용합니다. </li><li> 24 시간 동안 25 ° C에서 새로 입금 계란 판을 유지하여 부화 된 유생을 수집합니다. 부화 유충을 유치하기 ...

<ol>
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S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+trophic+role+for+serotonin+in+the+development+of+a+simple+feeding+circuit.">A trophic role for serotonin in the development of a simple feeding circuit.</a> Dev. Neurosci. 32, 217-237 .</li><li>De Vry, J., Schreiber, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effects+of+selected+serotonin+5-HT+1+and+5-HT+2+receptor+agonists+on+feeding+behavior:+possible+mechanisms+of+action.">Effects of selected serotonin 5-HT 1 and 5-HT 2 receptor agonists on feeding behavior: possible mechanisms of action.</a> Neurosci. Biobehav. Rev. 24, 341-353 (2000).</li><li>Schoofs, A., Niederegger, S., van Ooyen, A., Heinzel, H., Spie&#223;, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+brain+can+eat:+Establishing+the+existence+of+a+central+pattern+generator+for+feeding+in+third+instar+larvae+of+Drosophila+virilis+and+Drosophila+melanogaster.">The brain can eat: Establishing the existence of a central pattern generator for feeding in third instar larvae of Drosophila virilis and Drosophila melanogaster.</a> J. 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Res. 24, 163-173 (1975).</li><li>Joshi, A., Mueller, L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Evolution+of+higher+feeding+rate+in+Drosophila+due+to+density-dependent+natural+selection.">Evolution of higher feeding rate in Drosophila due to density-dependent natural selection.</a> Evolution. 42, 1090-1093 (1988).</li><li>Budnik, V., Wu, C., White, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Altered+branching+of+serotonin-containing+neurons+in+Drosophila+mutants+unable+to+synthesize+serotonin+and+dopamine.">Altered branching of serotonin-containing neurons in Drosophila mutants unable to synthesize serotonin and dopamine.</a> J. 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D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Temperature+sensing+across+species.">Temperature sensing across species.</a> Pflugers Archives. 454, 777-791 (2007).</li></ol>

후반 배아 중 발생하는 세로토닌 stomatogastric 회로의 탈선 개발, 자사의 성숙한 기능에 영향을 미칠 것입니다. 신경 돌기 구조의 변화는 용기가 (효모 용액에 입 후크의 수축에 의해 측정) 비율을 공급하는 회로의 기능 출력 (그림 1)과 상관 관계가 될 수있다 지배하는. 초파리 UAS-인 Gal4 양자 시스템을 사용함으로써, 특히 특정 조직에 관련 증명서의 상향 또는 하향 조절 식을 대?...

초파리는 급속한 생성 시간, 저렴한 실험 비용 및 유전 적 및 환경 적 요인을 조작하고 제어하는 능력을 신경 회로의 개발을위한 연구 매우 강력한 모델 시스템이다. 신경, 신경 경로 찾기 및 시냅스는 인간과 초파리 사이에 보존되어 있으므로, 작성 유지하고 신경 회로를 수정의 메커니즘뿐만 아니라 보존됩니다. 세로토닌 (5 - 하이드 록시 트립 타민, 5-HT)와 같은 고전적인 신경 전달 물질, 성숙한 신경 회로 1-3 신호 분자로 자신의 역할을 채택하기 전에 성장 인자 역할을 할 수 있습니다 이전 연구는 성숙한 신경 세포 (4)의 연결을 변경 배아 동안 5-HT의 수준을 교란하는 것으로 나타났습니다. 기타 교양 Helisoma 신경 세포에 5-HT의 자궁외 응용 프로그램이 신경 돌기 가지뿐만 아니라 시냅스 5-7을 억제하는 것으로 나타났습니다. D에rosophila는 발달 5-HT 수준은 반비례 CNS (8)로부터 foregut에 돌출 신경 돌기의 길이를 따라 정맥류의 수와 크기뿐만 아니라 aborization의 정도와 관련된다. 세로토닌 신경 전달 초파리 8-9를 포함, 다양한 종류의 먹이 행동을 조절하는 것으로 알려져있다. 초파리 급전 회로 foregut에 뇌로부터 축삭 돌기의 발달의 변화와 기능적 출력 (급전)를 상관시키는 모델로서 이용 될 수있는 비교적 간단한 회로이다. Schoofs 등. 초파리 유충의 먹이가 근육 (10)에 영향을 미치는 중앙 패턴 발생기에 의해 조절되는 것으로 나타났습니다. 특정 근육 해부학이 완전히 이해되지 않지만, 그것은 더듬이 신경, 상악 신경, 그리고 prothoracic 액세서리 신경에 관련된 근육의 대상에 대해 책임이 있음을 보였다동작을 먹이. 무척추 동물 먹이의 근육과 신경 해부학을 포함하는 대부분의 데이터는 Calliphora 애벌레로 제한됩니다. 두 번째 령 유충의 먹이 속도는 cephalopharyngeal 경피 (입 후크)의 수축에 의해 평가하고, 재현성 및 높은 처리량입니다 수 있습니다. cephalopharyngeal 플레이트는 정면 신경을 통해 중앙의 5-HT의 신경 세포에서 섬유에 의해 신경 지배된다. proventriculus, 또는 foregut은 중장에서 총생 (그림 1) 11 ~ 12 foregut의 수축에 대한 책임은 세로토닌 섬유 (신경을 recurrens)에 의해 신경 지배한다. 축삭 분기의 변경, 및 신경 돌기 길이를 따라 정맥류의 개수 및 크기는, 면역 조직 화학적 기법을 사용하여 정량 할 수있다. 직접 또는 간접적으로 개발 중에 신경 5-HT를 조작하는 것은 morpholo 변화로 평가되고 상관 될 수있는이 급전 회로의 출력 기능을 변경할 수신경 돌기 구조의 GY.

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			<td height="40" style="height:40px;width:217px;">Eclipse E-800 Microscope</td>
			<td style="width:175px;">Nikon Instruments</td>
			<td style="width:115px;"></td>
			<td style="width:251px;"></td>
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		<tr height="60">
			<td height="60" style="height:60px;width:217px;">Neuroleucida</td>
			<td style="width:175px;">MBF Biosciences</td>
			<td style="width:115px;">NL-15</td>
			<td style="width:251px;">Used to analyze gut fiber architecture, not necessary to have</td>
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		<tr height="20">
			<td height="20" style="height:20px;width:217px;">Northern Eclipse</td>
			<td style="width:175px;">Empix Inc</td>
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			<td style="width:251px;">Imaging software</td>
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		<tr height="60">
			<td height="60" style="height:60px;width:217px;">G-2E/C TRITC EX 528-553</td>
			<td style="width:175px;">Nikon Instruments</td>
			<td style="width:115px;">96312</td>
			<td style="width:251px;">Filter for specific secondary antibody</td>
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			<td height="80" style="height:80px;width:217px;">N.A. 0.75; W.D. 0.72 mm; DIC Prism: 40xI, 40x I-C; Spring loaded</td>
			<td style="width:175px;">Nikon Instruments</td>
			<td style="width:115px;">MRH00400</td>
			<td style="width:251px;">Objective used for imaging</td>
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			<td height="41" style="height:41px;width:217px;">Simple Neurite Tracer</td>
			<td style="width:175px;">NIH Image J</td>
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			<td style="width:251px;"><a href="http://fiji.sc/Simple_Neurite_Tracer">http://fiji.sc/Simple_Neurite_Tracer</a></td>
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functional analysis of the larval feeding circuit in drosophila

초파리 유충에서 세로토닌 공급 회로는 회로의 개발 과정에서 매우 중요한 신경 기판을 조사하는데 사용될 수있다. 회로의 출력 기능을 이용하여, 수유, stomatogastric 시스템의 신경 구조의 변화는 가시화 될 수있다. 먹이는 행동은 뇌에서 신경 지배를받는 입 후크의 후퇴 속도를 관찰하여 기록 할 수 있습니다. 애벌레 한천 기판을 가로 질러 통과 할 자신의 입 후크를 사용하기 때문에 전위의 동작은, 먹이에 대한 생리 학적 제어로 사용됩니다. 먹이는 행동의 변화는 신경 돌기의 축삭 구조와 상관 관계 될 수있는 창자를 지배하는. 면역 조직 화학을 사용하여 이들 변경을 가시화하고 정량 할 수있다. 그들이 조작에 매우 민감로 행동 패러다임 동안 유충을 잘못 취급하면 데이터를 변경할 수 있습니다. 신경 돌기 아키텍처 지배하는 적절한 영상창자는 정확한 정맥류의 수와 크기의 정량뿐만 아니라 분기 노드의 범위에 대한 중요합니다. 대부분의 회로의 분석에만 신경 돌기 아키텍처 또는 행동 효과의 가시화를 허용하지만,이 모델은 하나의 연결 구조에 장애가있는 회로의 기능적인 출력을 상관시킬 수있다.

초파리 유충에서 세로토닌 공급 회로는 신경계 발달에 특정 요인의 영향을 관찰 할 수있는 매우 효과적인 모델이 될 수 있습니다. 공급 비율을 정량함으로써, 그 기능의 출력 (도 1)와 급전 회로의 축삭 아키텍처를 연결하는 것이 가능하다. 유충은 한천의 표면을 가로 질러 자신을 추진하기 위해 입 후크를 사용하기 때문에 전위의 분석은, 입 후크의 철회에 대한 생리 학적 제어...

Watch this Scientific Journal Video about Functional Analysis of the Larval Feeding Circuit in Drosophila at JoVE.com

Functional Analysis of the Larval Feeding Circuit in Drosophila

초파리 유충의 먹이 회로는 공급 속도의 변화가 stomatogastric 신경 회로의 변화와 상관 관계가 될 수있는 간단하면서도 강력한 모델을 제공합니다. 이 회로는 입 후크 돌기뿐만 아니라 foregut 보내 중앙 세로토닌 성 신경 세포로 구성된다.

의 애벌레 먹이 회로의 기능적 분석 초파리</i

The serotonergic  feeding circuit in Drosophila  melanogaster larvae can be used to investigate neuronal substrates of  critical importance during the ...

functional-analysis-of-the-larval-feeding-circuit-in-drosophila

Research

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Neuroscience

Functional Analysis of the Larval Feeding Circuit in Drosophila

Visualization of Larval Segmental Nerves in 3rd Instar Drosophila Larval Preparations

Drosophila melanogaster is emerging as a powerful model system for studying the development and function of the nervous system, particularly because of its convenient genetics and fully sequenced genome. Additionally, the larval nervous system is an ideal model system to study mechanisms of axonal transport as the larval segmental nerves contain bundles of axons with their cell bodies located within the brain and their nerve terminals ending along the length of the body. Here we describe the procedure for visualization of synaptic vesicle proteins within larval segmental nerves. If done correctly, all components of the nervous system, along with associated tissues such as muscles and NMJs, remain intact, undamaged, and ready to be visualized. 3rd instar larvae carrying various mutations are dissected, fixed, incubated with synaptic vesicle antibodies, visualized and compared to wild type larvae. This procedure can be adapted for several different synaptic or neuronal antibodies and changes in the distribution of a variety of proteins can be easily observed within larval segmental nerves.

Visualization of Larval Segmental Nerves in 3rd Instar Drosophila Larval Preparations

Drosophila melanogaster larvae provide an ideal model system to investigate the mechanisms of axonal transport within larval segmental nerves. Using this procedure, 3rd instar larvae carrying various mutations can be compared to wild type larvae.

3 애벌레 Segmental 신경의 시각화 차 Instar Drosophila 애벌레 준비

Drosophila melanogaster is emerging as a powerful model system for studying the development and function of the nervous system, particularly because of ...

연구

신경과학

In vivo Visualization of Synaptic Vesicles Within Drosophila Larval Segmental Axons

Elucidating the mechanisms of axonal transport has shown to be very important in determining how defects in long distance transport affect different neurological diseases. Defects in this essential process can have detrimental effects on neuronal functioning and development. We have developed a dissection protocol that is designed to expose the Drosophila larval segmental nerves to view axonal transport in real time. We have adapted this protocol for live imaging from the one published by Hurd and Saxton (1996) used for immunolocalizatin of larval segmental nerves. Careful dissection and proper buffer conditions are critical for maximizing the lifespan of the dissected larvae. When properly done, dissected larvae have shown robust vesicle transport for 2-3 hours under physiological conditions. We use the UAS-GAL4 method 1 to express GFP-tagged APP or synaptotagmin vesicles within a single axon or many axons in larval segmental nerves by using different neuronal GAL4 drivers. Other fluorescently tagged markers, for example mitochrondria (MitoTracker) or lysosomes (LysoTracker), can be also applied to the larvae before viewing. GFP-vesicle movement and particle movement can be viewed simultaneously using separate wavelengths.

In vivo Visualization of Synaptic Vesicles Within Drosophila Larval Segmental Axons

This protocol discusses the live dissection of Drosophila larvae for the purpose of imaging the movement of GFP tagged axonal vesicles on microtubule tracks.

Drosophila 애벌레 Segmental Axons 이내 시냅틱 Vesicles의 생체내 시각화에

Elucidating the mechanisms of axonal transport has shown to be very important in determining how defects in long distance transport affect different ...

Morphological Analysis of Drosophila Larval Peripheral Sensory Neuron Dendrites and Axons Using Genetic Mosaics

Nervous system development requires the correct specification of neuron position and identity, followed by accurate neuron class-specific dendritic development and axonal wiring. Recently the dendritic arborization (DA) sensory neurons of the Drosophila larval peripheral nervous system (PNS) have become powerful genetic models in which to elucidate both general and class-specific mechanisms of neuron differentiation.
There are four main DA neuron classes (I-IV)1. They are named in order of increasing dendrite arbor complexity, and have class-specific differences in the genetic control of their differentiation2-10. The DA sensory system is a practical model to investigate the molecular mechanisms behind the control of dendritic morphology11-13 because: 1) it can take advantage of the powerful genetic tools available in the fruit fly, 2) the DA neuron dendrite arbor spreads out in only 2 dimensions beneath an optically clear larval cuticle making it easy to visualize with high resolution in vivo, 3) the class-specific diversity in dendritic morphology facilitates a comparative analysis to find key elements controlling the formation of simple vs. highly branched dendritic trees, and 4) dendritic arbor stereotypical shapes of different DA neurons facilitate morphometric statistical analyses.
DA neuron activity modifies the output of a larval locomotion central pattern generator14-16. The different DA neuron classes have distinct sensory modalities, and their activation elicits different behavioral responses14,16-20. Furthermore different classes send axonal projections stereotypically into the Drosophila larval central nervous system in the ventral nerve cord (VNC)21. These projections terminate with topographic representations of both DA neuron sensory modality and the position in the body wall of the dendritic field7,22,23. Hence examination of DA axonal projections can be used to elucidate mechanisms underlying topographic mapping7,22,23, as well as the wiring of a simple circuit modulating larval locomotion14-17.
We present here a practical guide to generate and analyze genetic mosaics24 marking DA neurons via MARCM (Mosaic Analysis with a Repressible Cell Marker)1,10,25 and Flp-out22,26,27 techniques (summarized in Fig. 1).

Morphological Analysis of Drosophila Larval Peripheral Sensory Neuron Dendrites and Axons Using Genetic Mosaics

The dendritic arborization sensory neurons of the Drosophila larval peripheral nervous system are useful models to elucidate both general and neuron class-specific mechanisms of neuron differentiation. We present a practical guide to generate and analyze dendritic arborization neuron genetic mosaics.

의 형태학의 분석 Drosophila</em유전 모자이크 사용하기> 애벌레 주변 감각 신경 세포의 Dendrites 및 Axons

Nervous system development requires the correct specification of neuron position and identity, followed by accurate neuron class-specific dendritic ...

Visualization of Proprioceptors in Drosophila Larvae and Pupae

Proprioception is the ability to sense the motion, or position, of body parts by responding to stimuli arising within the body. In fruitflies and other insects proprioception is provided by specialized sensory organs termed chordotonal organs (ChOs) 2. Like many other organs in Drosophila, ChOs develop twice during the life cycle of the fly. First, the larval ChOs develop during embryogenesis. Then, the adult ChOs start to develop in the larval imaginal discs and continue to differentiate during metamorphosis.
The development of larval ChOs during embryogenesis has been studied extensively 10,11,13,15,16. The centerpiece of each ChO is a sensory unit composed of a neuron and a scolopale cell. The sensory unit is stretched between two types of accessory cells that attach to the cuticle via specialized epidermal attachment cells 1,9,14. When a fly larva moves, the relative displacement of the epidermal attachment cells leads to stretching of the sensory unit and consequent opening of specific transient receptor potential vanilloid (TRPV) channels at the outer segment of the dendrite 8,12. The elicited signal is then transferred to the locomotor central pattern generator circuit in the central nervous system.
Multiple ChOs have been described in the adult fly 7. These are located near the joints of the adult fly appendages (legs, wings and halters) and in the thorax and abdomen. In addition, several hundreds of ChOs collectively form the Johnston's organ in the adult antenna that transduce acoustic to mechanical energy 3,5,17,4.
In contrast to the extensive knowledge about the development of ChOs in embryonic stages, very little is known about the morphology of these organs during larval stages. Moreover, with the exception of femoral ChOs 18 and Johnston's organ, our knowledge about the development and structure of ChOs in the adult fly is very fragmentary.
Here we describe a method for staining and visualizing ChOs in third instar larvae and pupae. This method can be applied together with genetic tools to better characterize the morphology and understand the development of the various ChOs in the fly.

Visualization of Proprioceptors in Drosophila Larvae and Pupae

A method to immunostain and visualize chordotonal organs in larvae and pupae of Drosophila melanogaster is described.

의 Proprioceptors의 시각화<em&gt; Drosophila</em&gt; 애벌레와 Pupae

Proprioception is the ability to sense the motion, or position, of body parts by responding to stimuli arising within the body. In fruitflies and other ...

Whole Mount Immunolabeling of Olfactory Receptor Neurons in the Drosophila Antenna

Odorant molecules bind to their target receptors in a precise and coordinated manner. Each receptor recognizes a specific signal and relays this information to the brain. As such, determining how olfactory information is transferred to the brain, modifying both perception and behavior, merits investigation. Interestingly, there is emerging evidence that cellular transduction and transcriptional factors are involved in the diversification of olfactory receptor neuron. Here we provide a robust whole mount immunological labeling method to assay in vivo olfactory receptor neuron organization. Using this method, we identified all olfactory receptor neurons with anti-ELAV antibody, a known pan-neural marker and Or49a-mCD8::GFP, an olfactory receptor neuron specifically expressed in Nba neuron using anti-GFP antibody.

Whole Mount Immunolabeling of Olfactory Receptor Neurons in the Drosophila Antenna

Herein we describe the process of whole mount immunostaining of Drosophila antennae, which enables us to better understand the molecular mechanisms involved in the diversification of olfactory receptor neurons (ORN)s.

에있는 후각 수용체 뉴런의 전체 마운트 Immunolabeling<em&gt; 초파리</em&gt; 안테나

Odorant molecules bind to their target receptors in a precise and coordinated manner. Each receptor recognizes a specific signal and relays this ...

Live Imaging of Drosophila Larval Neuroblasts

Stem cells divide asymmetrically to generate two progeny cells with unequal fate potential: a self-renewing stem cell and a differentiating cell. Given their relevance to development and disease, understanding the mechanisms that govern asymmetric stem cell division has been a robust area of study. Because they are genetically tractable and undergo successive rounds of cell division about once every hour, the stem cells of the Drosophila central nervous system, or neuroblasts, are indispensable models for the study of stem cell division. About 100 neural stem cells are located near the surface of each of the two larval brain lobes, making this model system particularly useful for live imaging microscopy studies. In this work, we review several approaches widely used to visualize stem cell divisions, and we address the relative advantages and disadvantages of those techniques that employ dissociated versus intact brain tissues. We also detail our simplified protocol used to explant whole brains from third instar larvae for live cell imaging and fixed analysis applications.

Live Imaging of Drosophila Larval Neuroblasts

This protocol details a streamlined method used to conduct live cell imaging in the context of an intact larval brain. Live cell imaging approaches are invaluable for the study of asymmetric neural stem cell divisions as well as other neurogenic and developmental processes, consistently uncovering mechanisms that were previously overlooked.

의 라이브 영상<em&gt; 초파리</em&gt; 애벌레 Neuroblasts

Stem cells divide asymmetrically to generate two progeny cells with unequal fate potential: a self-renewing stem cell and a differentiating cell. Given ...

Why Quantification Matters: Characterization of Phenotypes at the Drosophila Larval Neuromuscular Junction

Most studies on morphogenesis rely on qualitative descriptions of how anatomical traits are affected by the disruption of specific genes and genetic pathways. Quantitative descriptions are rarely performed, although genetic manipulations produce a range of phenotypic effects and variations are observed even among individuals within control groups.&#160;Emerging evidence shows that morphology, size and location of organelles play a previously underappreciated, yet fundamental role in cell function and survival. Here we provide step-by-step instructions for performing quantitative analyses of phenotypes at the Drosophila larval neuromuscular junction (NMJ). We use several reliable immuno-histochemical markers combined with bio-imaging techniques and morphometric analyses to examine the effects of genetic mutations on specific cellular processes. In particular, we focus on the quantitative analysis of phenotypes affecting morphology, size and position of nuclei within the striated muscles of Drosophila larvae. The Drosophila larval NMJ is a valuable experimental model to investigate the molecular mechanisms underlying the structure and the function of the neuromuscular system, both in health and disease. However, the methodologies we describe here can be extended to other systems as well.

Why Quantification Matters: Characterization of Phenotypes at the Drosophila Larval Neuromuscular Junction

Morphology, size and location of intracellular organelles are evolutionarily conserved and appear to directly affect their function. Understanding the molecular mechanisms underlying these processes has become an important goal of modern biology. Here we show how these studies can be facilitated by the application of quantitative techniques.

상기 표현형의 특성을 : 정량이 중요한 이유<em&gt; 초파리</em&gt; 애벌레 신경 근육 학 분기점

Most studies on morphogenesis rely on qualitative descriptions of how anatomical traits are affected by the disruption of specific genes and genetic ...

Removal of Drosophila Muscle Tissue from Larval Fillets for Immunofluorescence Analysis of Sensory Neurons and Epidermal Cells

Drosophila larval dendritic arborization (da) neurons are a popular model for investigating mechanisms of neuronal morphogenesis. Da neurons develop in communication with the epidermal cells they innervate and thus their analysis benefits from in situ visualization of both neuronally and epidermally expressed proteins by immunofluorescence. Traditional methods of preparing larval fillets for immunofluorescence experiments leave intact the muscle tissue that covers most of the body wall, presenting several challenges to imaging neuronal and epidermal proteins. Here we describe a method for removing muscle tissue from Drosophila larval fillets. This protocol enables imaging of proteins that are otherwise obscured by muscle tissue, improves signal to noise ratio, and facilitates the use of super-resolution microscopy to study da neuron development.

Removal of Drosophila Muscle Tissue from Larval Fillets for Immunofluorescence Analysis of Sensory Neurons and Epidermal Cells

Studies of neuronal morphogenesis using Drosophila larval dendritic arborization (da) neurons benefit from in situ visualization of neuronal and epidermal proteins by immunofluorescence. We describe a procedure that improves immunofluorescence analysis of da neurons and surrounding epidermal cells by removing muscle tissue from the larval body wall.

제거<em&gt; 초파리</em감각 뉴런과 표피 세포의 면역 형광 분석을위한 애벌레 필렛에서&gt; 근육 조직

Drosophila larval dendritic arborization (da) neurons are a popular model for investigating mechanisms of neuronal morphogenesis. Da neurons develop in ...

Using Linear Agarose Channels to Study Drosophila Larval Crawling Behavior

Drosophila larval crawling is emerging as a powerful model to study neural control of sensorimotor behavior. However, larval crawling behavior on flat open surfaces is complex,&#160;including: pausing, turning, and meandering. This complexity in the repertoire of movement hinders detailed analysis of the events occurring during a single crawl stride cycle. To overcome this obstacle, linear agarose channels were made that constrain larval behavior to straight, sustained, rhythmic crawling. In principle, because agarose channels and the Drosophila larval body are both optically clear, the movement of larval structures labeled by genetically-encoded fluorescent probes can be monitored in intact, freely-moving larvae. In the past, larvae were placed in linear channels and crawling at the level of whole organism, segment, and muscle were analyzed1. In the future, larvae crawling in channels can be used for calcium imaging to monitor neuronal activity. Moreover, these methods can be used with larvae of any genotype and with any researcher-designed channel. Thus the protocol presented below is widely applicable for studies using the Drosophila larva as a model to understand motor control.

Using Linear Agarose Channels to Study Drosophila Larval Crawling Behavior

The Drosophila larva is a powerful model system to study neural control of behavior. This publication describes the use of linear agarose channels to elicit sustained bouts of linear crawling and methods to quantify the dynamics of larval structures during repetitive crawling behavior.

선형 아가로 오스 채널을 사용하여 공부하기<em&gt; 초파리</em&gt; 애벌레 크롤링 동작

Drosophila larval crawling is emerging as a powerful model to study neural control of sensorimotor behavior. However, larval crawling behavior on flat ...

Focal Macropatch Recordings of Synaptic Currents from the Drosophila Larval Neuromuscular Junction

Drosophila neuromuscular junction (NMJ) is an excellent model system to study glutamatergic synaptic transmission. We describe the technique of focal macropatch recordings of synaptic currents from visualized boutons at the Drosophila larval NMJ. This technique requires customized fabrication of recording micropipettes, as well as a compound microscope equipped with a high magnification, long-distance water immersion objective, differential interference contrast (DIC) optics, and a fluorescent attachment. The recording electrode is positioned on the top of a selected synaptic bouton visualized with DIC optics, epi-fluorescence, or both. The advantage of this technique is that it allows monitoring the synaptic activity of a limited number of sites of release. The recording electrode has&#160;a diameter of several microns, and the release sites positioned outside of the electrode rim do&#160;not significantly affect the recorded currents. The recorded synaptic currents have fast kinetics and can be readily resolved. These advantages are especially important for the studies of mutant fly lines with enhanced spontaneous or asynchronous synaptic activity.

Focal Macropatch Recordings of Synaptic Currents from the Drosophila Larval Neuromuscular Junction

Synaptic currents can be recorded focally from visualized synaptic boutons at the Drosophila third instar larvae neuromuscular junction. This technique enables monitoring the activity of a single synaptic bouton.

초파리 애벌레 신경 근육 학 교차로에서 시 냅 스 전류의 초점 Macropatch 녹음

Drosophila neuromuscular junction (NMJ) is an excellent model system to study glutamatergic synaptic transmission. We describe the technique of focal ...