<ol>
	<li>Papazian, D. M., Schwarz, T. L., Tempel, B. L., Jan, Y. N., Jan, L. Y. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cloning+of+genomic+and+complementary+DNA+from+Shaker,+a+putative+potassium+channel+gene+from+Drosophila.">Cloning of genomic and complementary DNA from Shaker, a putative potassium channel gene from Drosophila.</a> Science. 237, 749-753 (1987).</li><li>Tempel, B. L., Papazian, D. M., Schwarz, T. L., Jan, Y. N., Jan, L. Y. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Sequence+of+a+probable+potassium+channel+component+encoded+at+Shaker+locus+of+Drosophila.">Sequence of a probable potassium channel component encoded at Shaker locus of Drosophila.</a> Science. 237, 770-775 (1987).</li><li>Gu, H., O'Dowd, D. K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Whole+Cell+Recordings+from+Brain+of+Adult+Drosophila.">Whole Cell Recordings from Brain of Adult Drosophila.</a> J. Vis. Exp. (6), e248 (2007).</li><li>Gu, H., Jiang, S. A., Campusano, J. M., Iniguez, J., Su, H., Hoang, A. A., Lavian, M., Sun, X., O'Dowd, D. K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cav2-type+calcium+channels+encoded+by+cac+regulate+AP-independent+neurotransmitter+release+at+cholinergic+synapses+in+adult+Drosophila+brain.">Cav2-type calcium channels encoded by cac regulate AP-independent neurotransmitter release at cholinergic synapses in adult Drosophila brain.</a> J. Neurophysiol. 101, 42-53 (2009).</li><li>Baines, R. A., Bate, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Electrophysiological+development+of+central+neurons+in+the+Drosophila+embryo.">Electrophysiological development of central neurons in the Drosophila embryo.</a> J. Neurosci. 18, 4673-4683 (1998).</li><li>Lin, W. H., Wright, D. E., Muraro, N. I., Baines, R. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Alternative+splicing+in+the+voltage-gated+sodium+channel+DmNav+regulates+activation,+inactivation,+and+persistent+current.">Alternative splicing in the voltage-gated sodium channel DmNav regulates activation, inactivation, and persistent current.</a> J. Neurophysiol. 102, 1994-2006 (2009).</li><li>Worrell, J. W., Levine, R. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Characterization+of+voltage-dependent+Ca2++currents+in+identified+Drosophila+motoneurons+in+situ.">Characterization of voltage-dependent Ca2+ currents in identified Drosophila motoneurons in situ.</a> J. Neurophysiol. 100, 868-878 (2008).</li><li>Srinivasan, S., Lance, K., Levine, R. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Segmental+differences+in+firing+properties+and+potassium+currents+in+Drosophila+larval+motoneurons.">Segmental differences in firing properties and potassium currents in Drosophila larval motoneurons.</a> J. Neurophysiol. , (2011).</li><li>Choi, J. C., Park, D., Griffith, L. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Electrophysiological+and+morphological+characterization+of+identified+motor+neurons+in+the+Drosophila+third+instar+larva+central+nervous+system.">Electrophysiological and morphological characterization of identified motor neurons in the Drosophila third instar larva central nervous system.</a> J. Neurophysiol. 91, 2353-2365 (2004).</li><li>Pulver, S. R., Griffith, L. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Spike+integration+and+cellular+memory+in+a+rhythmic+network+from+Na+/K++pump+current+dynamics.">Spike integration and cellular memory in a rhythmic network from Na+/K+ pump current dynamics.</a> Nat. Neurosci. 13, 53-59 (2010).</li><li>Fayyazuddin, A., Zaheer, M. A., Hiesinger, P. R., Bellen, H. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+nicotinic+acetylcholine+receptor+Dalpha7+is+required+for+an+escape+behavior+in+Drosophila.">The nicotinic acetylcholine receptor Dalpha7 is required for an escape behavior in Drosophila.</a> PLoS Biol. 4, e63 (2006).</li><li>Duch, C., Vonhoff, F., Ryglewski, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Dendrite+elongation+and+dendritic+branching+are+affected+separately+by+different+forms+of+intrinsic+motoneuron+excitability.">Dendrite elongation and dendritic branching are affected separately by different forms of intrinsic motoneuron excitability.</a> J. Neurophysiol. 100, 2525-2536 (2008).</li><li>Ryglewski, S., Duch, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Shaker+and+Shal+mediate+transient+calcium-independent+potassium+current+in+a+Drosophila+flight+motoneuron.">Shaker and Shal mediate transient calcium-independent potassium current in a Drosophila flight motoneuron.</a> J. Neurophysiol. 102, 3673-3688 (2009).</li><li>Ryglewski, S., Lance, K., Levine, R. B., Duch, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cav2+Channels+Mediate+LVA+and+HVA+Calcium+Currents+in+Drosophila+Motoneurons.">Cav2 Channels Mediate LVA and HVA Calcium Currents in Drosophila Motoneurons.</a> J. Physiol. , (2011).</li><li>Ikeda, K., Koenig, J. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Morphological+identification+of+the+motor+neurons+innervating+the+dorsal+longitudinal+flight+muscle+of+Drosophila+melanogaster.">Morphological identification of the motor neurons innervating the dorsal longitudinal flight muscle of Drosophila melanogaster.</a> J. Comp. Neurol. 1273, 436-444 (1988).</li><li>Boerner, J., Godenschwege, T. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Whole+Mount+Preparation+of+the+Adult+Drosophila+Ventral+Nerve+Cord+for+Giant+Fiber+Dye+Injection.">Whole Mount Preparation of the Adult Drosophila Ventral Nerve Cord for Giant Fiber Dye Injection.</a> J. Vis. Exp. (52), e3080 (2011).</li></ol>

관심 없음 충돌이 선언 없습니다.

GFP 같은 형광 단백질과 세포를 시각화 할 때, 너무 많은 빛에 준비를 과잉 노출하지 않는 것이 중요합니다. 이 사진 손상 될 수 있습니다. 우리는 조명을위한 100W HBO 짧은 아크 수은 전구를 사용하며, 우리는 또한 0.8 (채도 ND 필터 0.3과 0.5)의 중간 밀도 (ND)을 ​​사용합니다. 청소 좋은 가시성의 품질을 판단 할 수 있도록하는 것은 매우 중요합니다. 따라서, ND 필터는 여러 번에 20 s의 짧은 기간 동?...

<table border="1"><tbody><tr><td> 에이전트 / 항목 </td><td> 회사 </td><td> 카탈로그 번호 </td></tr><tr><td> 단백 분해 효소 타입 XIV </td><td> 시그마 알드리치 USA </td><td> P5147 </td></tr><tr><td> Microfil 유연한 주입 바늘 </td><td> 세계 정밀 계측기 USA </td><td> MF28G-5 </td></tr><tr><td> 붕규산 유리 모세 혈관, OD 1.5 mm, ID 1.0 mm, 아니 필라멘트 </td><td> 세계 정밀 계측기 USA </td><td> PG52151-4 </td></tr><tr><td> DiI </td><td> Invitrogen USA </td><td> D3899 </td></tr><tr><td> Sylgard 엘라스토머 키트 184 (다우 코닝) </td><td> <a href="http://www.ellsworth.com" target="_blank">www.ellsworth.com</a> </td><td> 184 SIL ELAST KIT </td></tr><tr><td> ND 필터 세트 (언 마운트) </td><td> 크로마 </td><td> 22000b 시리즈 </td></tr><tr><td> 전극 홀더 1-HL-U </td><td> 모lecular 장치 </td><td> 1-HL-U </td></tr></tbody></table>

preparation of drosophila central neurons for in situ patch clamping

짧은 세대 시간 및 손쉬운 유전자 기술은 과일 파리 Drosophila melanogaster 기초 신경 과학 연구의 훌륭한 유전자 모델을 확인합니다. 그들은 neuronal 흥분을 중재 이후 이온 채널은 모든 행동의 기초입니다. 복제 제 1 전압 개폐 이온 채널은 Drosophila 전압 개폐 칼륨 채널 쉐이커 1,2했다. 신경계 기능에 대한 이온 채널과 막 흥분의 역할을 이해하는 방향으로는 현장 패치 클램프 녹음에서와 Drosophila에서 강력한 유전 도구를 사용할 수 결합 할 때 유용합니다. 수년 동안 이러한 녹음은 Drosophila의 CNS의 작은 크기에 방해되었습니다. 또한, glia 및 콜라겐으로 만든 강력한 집은 중앙 뉴런에 패치 피펫 액세스에 대한 장애물을 형성. 이 집의 제거는 성인 Drosophila의 CNS에서 뉴런의 패치 클램프 녹음에 필요한 전제 조건이다. 최근 몇 년과학자들은 성인 뇌 3,4 및 배아 5,6, 애벌레의 7,8,9,10, 그리고 성인 Drosophila 11,12,13,14의 복부 신경 코드를 뉴런에서 현장 패치 클램프 녹음에서 수행 할 수있었습니다. 안정적인 기가 - 씰은 좋은 패치의 주요 전제 조건이며, 누설 전류를 방지하기 위해 세포막과 패치 피펫의 깨끗한 접촉에 따라 달라집니다. 따라서 성인 Drosophila의 뉴런에서 현장 패치 클램프 녹음에서 전체 셀 철저하게 세척해야합니다. 첫 번째 단계에서, ganglionic 집은 효소 처리 할 수​​ 있으며 기계적으로 대상 셀에 액세스 할 수 있도록하기 위해 제거. 두 번째 단계에서, 세포막은 glia, 콜라겐 또는 기타 자료의 어떤 항목은 기가 - 씰 형성을 방해하지 수 있도록 연마되어야한다. 이 문서는 체세포의 전체 C의 Drosophila 복부 신경 코드, 비행 motoneuron 5 (MN5 15)에서 확인 중앙 신경 세포를 준비하는 방법에 대해 설명합니다예쁜 패치 클램프 녹음. 신경 세포의 식별 및 가시성이 MN5에서 GFP의 대상 표현에 의해 달성된다. 우리는 패치 클램프 기술 자체를 설명하는 것을 목표로하지 않습니다.

Watch this Scientific Journal Video about Preparation of Drosophila Central Neurons for in situ Patch Clamping at JoVE.com

Preparation of Drosophila Central Neurons for in situ Patch Clamping

현장 패치에 클램프 녹음은 그대로 회로의 뉴런의 electrophysiological 특성에 사용됩니다. CNS는 작고 강력한 피복에 둘러싸여 있기 때문에 Drosophila 유전자 모델 패치를 고정으로 어렵습니다. 이 문서는 이후의 패치 클램프 녹음의 집과 깨끗한 뉴런을 제거 할 수있는 절차에 대해 설명합니다.

의 작성<em&gt; Drosophila</em에 대한&gt; 중부 뉴런<em&gt; 현장에서</em&gt; 패치 클램핑

Short generation times and facile genetic techniques make the fruit fly Drosophila melanogaster an excellent genetic model in fundamental neuroscience ...

preparation-of-drosophila-central-neurons-for-in-situ-patch-clamping

Research

JoVE Journal

Neuroscience

Preparation of Drosophila Central Neurons for in situ Patch Clamping

14.0K 조회수.  Arizona State University. 현장 패치에 클램프 녹음은 그대로 회로의 뉴런의 electrophysiological 특성에 사용됩니다. CNS는 작고 강력한 피복에 둘러싸여 있기 때문에 Drosophila 유전자 모델 패치를 고정으로 어렵습니다. 이 문서는 이후의 패치 클램프 녹음의 집과 깨끗한 뉴런을 제거 할 수있는 절차에 대해 설명합니다.

비디오: Preparation of Drosophila Central Neurons for in situ Patch Clamping

Primary Neuronal Cultures from the Brains of Late Stage Drosophila Pupae

In this video, we demonstrate the preparation of primary neuronal cultures from the brains of late stage Drosophila pupae. The procedure begins with the removal of brains from animals at 70-78 hrs after puparium formation. The isolated brains are shown after brief incubation in papain followed by several washes in serum-free growth medium. The process of mechanical dissociation of each brain in a 5 ul drop of media on a coverslip is illustrated. The axons and dendrites of the post-mitotic neurons are sheered off near the soma during dissociation but the neurons begin to regenerate processes within a few hours of plating. Images show live cultures at 2 days. Neurons continue to elaborate processes during the first week in culture. Specific neuronal populations can be identified in culture using GAL4 lines to drive tissue specific expression of fluorescent markers such as  GFP or RFP. Whole cell recordings have demonstrated the cultured neurons form functional, spontaneously active cholinergic and GABAergic synapses. A short video segment illustrates calcium dynamics in the cultured neurons using Fura-2 as a calcium indicator dye to monitor spontaneous calcium transients and nicotine evoked calcium responses in a dish of cultured neurons. These pupal brain cultures are a useful model system in which genetic and pharmacological tools can be used to identify intrinsic and extrinsic factors that influence formation and function of central synapses.

This video demonstrates the preparation of primary neuronal cultures from the brains of late stage Drosophila pupae. Views of live cultures show neurite outgrowth and imaging of calcium levels using Fura-2.

늦은 단계 Drosophila Pupae의 두뇌에서 기본의 연결을 문화

In this video, we demonstrate the preparation of primary neuronal cultures from the brains of late stage Drosophila pupae. The procedure begins with the ...

연구

생물학

Whole Cell Recordings from Brain of Adult Drosophila

In this video, we demonstrate the procedure for isolating whole brains from adult Drosophila in preparation for recording from single neurons. We begin by describing the dissecting solution and capture of the adult females used in our studies. The procedure for removing the whole brain intact, including both optic lobes, is illustrated. Dissection of the overlying trachea is also shown.  The isolated brain is not only small but needs special care in handling at this stage to prevent damage to the neurons, many of which are close to the outer surface of the tissue. We show how a special holder we developed is used to stabilize the brain in the recording chamber. A standard electrophysiology set up is used for recording from single neurons or pairs of neurons. A fluorescent image, viewed through the recording microscope, from a GAL4 line driving GFP expression (GH146) illustrates how projection neurons (PNs) are identified in the live brain. A high power Nomarski image shows a view of a single neuron that is being targeted for whole cell recording. When the brain is successfully removed without damage, the majority of the neurons are spontaneously active, firing action potentials and/or exhibiting spontaneous synaptic input. This in situ preparation, in which whole cell recording of identified neurons in the whole brain can be combined with genetic and pharmacological manipulations, is a useful model for exploring cellular physiology and plasticity in the adult CNS.

This video demonstrates the procedure for isolating whole brains from adult Drosophila in preparation for recording from single neurons using standard whole cell technology. It includes images of GFP labeled cells and neurons viewed during recording.

성인 Drosophila의 머리에서 전체 셀 레코딩

In this video, we demonstrate the procedure for isolating whole brains from adult Drosophila in preparation for recording from single neurons. We begin by ...

Mapping and Application of Enhancer-trap Flippase Expression in Larval and Adult Drosophila CNS

The Gal4/ UAS binary method is powerful for gene and neural circuitry manipulation in Drosophila. For most neurobiological studies, however, Gal4 expression is rarely tissue-specific enough to allow for precise correlation of the circuit with behavioral readouts. To overcome this major hurdle, we recently developed the FINGR method to achieve a more restrictive Gal4 expression in the tissue of interest. The FINGR method has three components: 1) the traditional Gal4/UAS system; 2) a set of FLP/FRT-mediated Gal80 converting tools; and 3) enhancer-trap FLP (ET-FLP). Gal4 is used to define the primary neural circuitry of interest. Paring the Gal4 with a UAS-effector, such as UAS-MJD78Q or UAS-Shits, regulates the neuronal activity, which is in turn manifested by alterations in the fly behavior. With an additional UAS-reporter such as UAS-GFP, the neural circuit involved in the specific behavior can be simultaneously mapped for morphological analysis. For Gal4 lines with broad expression, Gal4 expression can be restricted by using two complementary Gal80-converting tools: tubP&gt;Gal80&gt; ('flip out') and tubP&gt;stop&gt;Gal80 ('flip in'). Finally, investigators can turn Gal80 on or off, respectively, with the help of tissue-specific ET-FLP. In the flip-in mode, Gal80 will repress Gal4 expression wherever Gal4 and ET-FLP intersect. In the flip-out mode, Gal80 will relieve Gal4 repression in cells in which Gal4 and FLP overlap. Both approaches enable the restriction of the number of cells in the Gal4-defined circuitry, but in an inverse pattern. The FINGR method is compatible with the vast collection of Gal4 lines in the fly community and highly versatile for traditional clonal analysis and for neural circuit mapping. In this protocol, we demonstrate the mapping of FLP expression patterns in select ET-FLPx2 lines and the effectiveness of the FINGR method in photoreceptor cells. The principle of the FINGR method should also be applicable to other genetic model organisms in which Gal4/UAS, Gal80, and FLP/FRT are used.

Mapping and Application of Enhancer-trap Flippase Expression in Larval and Adult Drosophila CNS

We describe a Flippase-induced intersectional Gal80/Gal4 repression (FINGR) method, allowing tissue-specific FLP to determine Gal80 expression patterns. Wherever Gal4 and FLP overlap, Gal4 expression is turned on (Gal80 flipped out) or off (Gal80 flipped in). The FINGR method is versatile for clonal analysis and neural circuit mapping.

매핑 및 애벌레와 성인의 확장기 - 트랩 Flippase의 표현의 응용 Drosophila CNS

The Gal4/ UAS binary method is powerful for gene and neural circuitry manipulation in Drosophila. For most neurobiological studies, however, Gal4 ...

신경과학

Electrophysiological Recording in the Drosophila Embryo

Drosophila is a premier genetic model for the study of both embryonic development and functional neuroscience. Traditionally, these fields are quite isolated from each other, with largely independent histories and scientific communities. However, the interface between these usually disparate fields is the developmental programs underlying acquisition of functional electrical signaling properties and differentiation of functional chemical synapses during the final phases of neural circuit formation. This interface is a critically important area for investigation. In Drosophila, these phases of functional development occur during a period of &lt;8 hours (at 25&deg;C) during the last third of embryogenesis. This late developmental period was long considered intractable to investigation owing to the deposition of a tough, impermeable epidermal cuticle. A breakthrough advance was the application of water-polymerizing surgical glue that can be locally applied to the cuticle to enable controlled dissection of late-stage embryos. With a dorsal longitudinal incision, the embryo can be laid flat, exposing the ventral nerve cord and body wall musculature to experimental investigation. Whole-cell patch-clamp techniques can then be employed to record from individually-identifiable neurons and somatic muscles. These recording configurations have been used to track the appearance and maturation of ionic currents and action potential propagation in both neurons and muscles. Genetic mutants affecting these electrical properties have been characterized to reveal the molecular composition of ion channels and associated signaling complexes, and to begin exploration of the molecular mechanisms of functional differentiation. A particular focus has been the assembly of synaptic connections, both in the central nervous system and periphery. The glutamatergic neuromuscular junction (NMJ) is most accessible to a combination of optical imaging and electrophysiological recording. A glass suction electrode is used to stimulate the peripheral nerve, with excitatory junction current (EJC) recordings made in the voltage-clamped muscle. This recording configuration has been used to chart the functional differentiation of the synapse, and track the appearance and maturation of presynaptic glutamate release properties. In addition, postsynaptic properties can be assayed independently via iontophoretic or pressure application of glutamate directly to the muscle surface, to measure the appearance and maturation of the glutamate receptor fields. Thus, both pre- and postsynaptic elements can be monitored separately or in combination during embryonic synaptogenesis. This system has been heavily used to isolate and characterize genetic mutants that impair embryonic synapse formation, and thus reveal the molecular mechanisms governing the specification and differentiation of synapse connections and functional synaptic signaling properties.

Electrophysiological Recording in the Drosophila Embryo

Electrophysiological recordings from Drosophila embryos allow analyses of developing muscle and neuron electrical properties, as well as characterization of functional synaptogenesis at the glutamatergic neuromuscular junction and central cholinergic and GABAergic synapses.

에 Electrophysiological 녹음 Drosophila 엠브료

Drosophila is a premier genetic model for the study of both embryonic development and functional neuroscience. Traditionally, these fields are quite ...

Electrophysiological Method for Whole-cell Voltage Clamp Recordings from Drosophila Photoreceptors

Whole-cell voltage clamp recordings from Drosophila melanogaster photoreceptors have revolutionized the field of invertebrate visual transduction, enabling the use of D. melanogaster molecular genetics to study inositol-lipid signaling and Transient Receptor Potential (TRP) channels at the single-molecule level. A handful of labs have mastered this powerful technique, which enables the analysis of the physiological responses to light under highly controlled conditions. This technique allows control over the intracellular and extracellular media; the membrane voltage; and the fast application of pharmacological compounds, such as a variety of ionic or pH indicators, to the intra- and extracellular media. With an exceptionally high signal-to-noise ratio, this method enables the measurement of dark spontaneous and light-induced unitary currents (i.e.&#160;spontaneous and quantum bumps) and macroscopic Light-induced Currents (LIC) from single D. melanogaster photoreceptors. This protocol outlines, in great detail, all the key steps necessary to perform this technique, which includes both electrophysiological and optical recordings. The fly retina dissection procedure for the attainment of intact and viable ex vivo isolated ommatidia in the bath chamber is described. The equipment needed to perform whole-cell and fluorescence imaging measurements are also detailed. Finally, the pitfalls in using this delicate preparation during extended experiments are explained.

Electrophysiological Method for Whole-cell Voltage Clamp Recordings from Drosophila Photoreceptors

Whole-cell recordings from Drosophila melanogaster photoreceptors enable the measurement of spontaneous dark bumps, quantum bumps, macroscopic responses to light, and current-voltage relationships under various conditions. In combination with D. melanogaster genetic manipulation tools, this method enables the study of the ubiquitous inositol-lipid signaling pathway and its target, the TRP channel.

전 세포 전압 클램프 기록을위한 전기 생리 학적 방법<em&gt; Drosophila</em&gt; 광 수용체

Whole-cell voltage clamp recordings from Drosophila melanogaster photoreceptors have revolutionized the field of invertebrate visual transduction, ...

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 ...

Examining Monosynaptic Connections in Drosophila Using Tetrodotoxin Resistant Sodium Channels

Here, a new technique termed Tetrotoxin (TTX) Engineered Resistance for Probing Synapses (TERPS) is applied to test for monosynaptic connections between target neurons. The method relies on co-expression of a transgenic activator with the tetrodotoxin-resistant sodium channel, NaChBac, in a specific presynaptic neuron. Connections with putative post-synaptic partners are determined by whole-cell recordings in the presence of TTX, which blocks electrical activity in neurons that do not express NaChBac. This approach can be modified to work with any activator or calcium imaging as a reporter of connections. TERPS adds to the growing set of tools available for determining connectivity within networks. However, TERPS is unique in that it also reliably reports bulk or volume transmission and spillover transmission.

Examining Monosynaptic Connections in Drosophila Using Tetrodotoxin Resistant Sodium Channels

This article features a method to test the monosynaptic connections between neurons by employing tetrodotoxin and the tetrodotoxin-resistant sodium channel, NaChBac.

Drosophila 테트로도톡신 저항 나트륨 채널을 사용 하 여 Monosynaptic 연결 검사

Here, a new technique termed Tetrotoxin (TTX) Engineered Resistance for Probing Synapses (TERPS) is applied to test for monosynaptic connections between ...

Quantitative Analysis of Neuronal Dendritic Arborization Complexity in Drosophila

Dendrites are the branched projections of a neuron, and dendritic morphology reflects synaptic organization during the development of the nervous system. Drosophila larval neuronal dendritic arborization (da) is an ideal model for studying morphogenesis of neural dendrites and gene function in the development of nervous system. There are four classes of da neurons. Class IV is the most complex with a branching pattern that covers almost the entire area of the larval body wall. We have previously characterized the effect of silencing the Drosophila ortholog of SOX5 on class IV neuronal dendritic arborization complexity (NDAC) using four parameters: the length of dendrites, the surface area of dendrite coverage, the total number of branches, and the branching structure. This protocol presents the workflow of NDAC quantitative analysis, consisting of larval dissection, confocal microscopy, and image analysis procedures using ImageJ software. Further insight into da neuronal development and its underlying mechanisms will improve the understanding of neuronal function and provide clues about the fundamental causes of neurological and neurodevelopmental disorders.

Quantitative Analysis of Neuronal Dendritic Arborization Complexity in Drosophila

This protocol focuses on quantitative analysis of neuronal dendritic arborization complexity (NDAC) in Drosophila, which can be used for studies of dendritic morphogenesis.

초파리 에서 신경 돌기 Arborization 복잡성의 정량 분석

Dendrites are the branched projections of a neuron, and dendritic morphology reflects synaptic organization during the development of the nervous system. ...

Inducing Long-Term Plasticity of Intrinsic Neuronal Excitability in Neurons of the Dorsal Lateral Geniculate Nucleus

The dorsal lateral geniculate nucleus (dLGN) has long been held to act as a basic relay for visual information traveling from the retina to cortical areas, but recent findings suggest largely underestimated functional plasticity of dLGN principal cells. However, the cellular mechanisms supporting these changes have not been fully explored. Here, we report a protocol to induce long-term potentiation of intrinsic neuronal excitability (LTP-IE) in dorsal dLGN relay cells from acute brain slices of young rats. Intrinsic plasticity is generally induced in parallel with synaptic plasticity. However, in dLGN neurons, LTP-IE is reliably induced by spiking activity at a frequency of 40 Hz for 10 min. LTP-IE in dLGN relay neurons is long-lasting as it can be followed up to 40 min after the induction protocol. In conclusion, the results of this study provide the first evidence for the induction of intrinsic plasticity in dLGN relay cells, thus further pointing to the role of thalamic neurons in activity-dependent visual plasticity.

Here, we describe a protocol for inducing long-term plasticity of neuronal intrinsic excitability in relay neurons from the dorsal lateral geniculate nucleus maintained in ex vivo brain slices.

The dorsal lateral geniculate nucleus (dLGN) has long been held to act as a basic relay for visual information traveling from the retina to cortical ...

Extracellular Recording of Electrical Activity in the Drosophila Central Nervous System

Source: Swale, D. R., et al. <a href="https://app.jove.com/v/58375">Electrophysiological Recording of The Central Nervous System Activity of Third-Instar Drosophila Melanogaster.</a> J. Vis. Exp. (2018)
The video demonstrates the extracellular recording of neuronal activity from the central nervous system (CNS) of Drosophila larvae. The transected CNS is positioned in a recording chamber, and the electrical activity from a descending nerve is recorded to establish the baseline firing frequency. A test agent is then introduced, and the change in firing frequency is recorded to determine the agent&#39;s excitatory or inhibitory effect.

초파리 중추신경계의 전기적 활동에 대한 세포외 기록

All procedures involving sample collection have been performed in accordance with the institute&#39;s IRB guidelines.
1. Dissect and Prepare the Larval ...

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