עבודה זו נתמכה על ידי מענק מDFG לGMT

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M., Seifert, E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Axon+growth+from+limb+motorneurons+in+the+locust+embryo:+the+effect+of+target+limb+removal+on+the+pattern+of+axon+branching+in+the+periphery.">Axon growth from limb motorneurons in the locust embryo: the effect of target limb removal on the pattern of axon branching in the periphery.</a> Developmental Biology. 106, 438-449 (1984).</li><li>Whitington, P. M., Leach, D., Sandeman, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Evolutionary+change+in+neural+development+within+the+arthropods:+axonogenesis+in+the+embryos+of+two+crustaceans.">Evolutionary change in neural development within the arthropods: axonogenesis in the embryos of two crustaceans.</a> Development. 118, 449-461 (1993).</li><li>Williams, D. W., Tyrer, M., Shepherd, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Tau+and+tau+reporters+disrupt+central+projections+of+sensory+neurons+in+Drosophila.">Tau and tau reporters disrupt central projections of sensory neurons in Drosophila.</a> J. Comp. Neurol. 428, 630-640 (2000).</li></ol>

החוקרים מצהירים כי אין להם אינטרסים כלכליים מתחרים.

אחד יתרונות עיקריים של דרוזופילה כמודל למערכת הוא שהיא מאפשרת ניתוח של התפתחות ותפקוד ברמה של תאים בודדים. זה שימושי במיוחד לגבי מערכת עצבים, שם המגוון של סוגי תאים הוא גבוה במיוחד והתפקוד והמורפולוגיה של תאי שכנים יכולים להיות שונים לחלוטין. <p class="jove_content" style=...

ידע של מורפולוגיה עצבית ברמה של תאים בודדים הוא תנאי מפתח להבנת קישוריות עצבית ותפקוד מערכת עצבים מרכזיים. כך, החל מהימים הראשונים של מדעי המוח, החוקרים בקשו לפתח טכניקות תיוג תא בודדות (ראה 7 לטיפול הסטורי של בעיה זו). שיטות קלסיות כגון צביעת Golgi לספק רזולוציה מצוינת של מורפולוגיה עצבית אך אינן מתאימים אם אחד מבקש לתייג סוג מסוים של נוירון באופן מכוון, כמכתים מתרחשת באופן אקראי. הפיתוח של שיטות להכתמת תאים בודדים על ידי הזרקה תאית או juxtacellular של צבעים מmicroelectrode התייחס לדרישה של תיוג ספציפי. היישום של זריקה אחת נוירון לצבוע כדי תסיסנית הציג אתגר גדול, בגלל גודלו הקטן של שניהם האורגניזם ותאי העצב שלה. עם זאת, כתמי נוירון בודדים של עוברית דרוזופילה </emהנוירונים&gt; הושגו באמצע 1980 במעבדה של הקורים 15 גודמן. בעוד השיטה הייתה בהחלט שימושית וספקה כמה תובנה חשובות על מנגנונים להתפתחות עצבית בדרוזופילה במשך 20-30 השנים האחרונות (לדוגמה, 2, 10), עובדים רבים נרתעו ממנו, בעיקר בגלל הדרישות הטכניות שלה. הזמינה בשנים האחרונות יותר של טכניקות גנטיות לתיוג עצבי בדרוזופילה גם תרם לפופולריות של הזרקת צבע נוירון בודדה. ביטוי GAL4 ביים של מבני GFP קרום ממוקדים יכול לספק פתרון מצוין למורפולוגיה עצבית 1, 20. עם זאת, לשיטה זו יש מגבלות מסוימות: הביטוי לעתים קרובות בלתי הנמנע של ה-GFP בתאים מרובים יכול לטשטש את המבנה של תאי עצב בודד וקו נהג GAL4 לא יהיה זמין לנהוג ביטוי בתא עצב מסוים של ריבית. MARCM (ניתוח פסיפס עם ציטוטמרקר סלולרי ressible) שיטה 8 יכול לספק תיוג של מהות כל נוירון ברמת התא הבודדה, אך לא ניתן להשתמש בהצלחה בעובר והזחל מוקדם בגלל התחלופה האיטית של חלבון GAL80. בהתחשב במגבלות אלה של תיוג גנטי, אנו מאמינים כי הזרקת צבע נוירון הבודדה בעובר דרוזופילה נשארת טכניקה חשובה וראויה ליישום רחב יותר. כדי לקדם מטרה זו, אנו מספקים כאן תיאור מפורט של השיטה. המחשת כוחה מסופקת על ידי החשבון האחרון שלנו על המורפולוגיה של הסט השלם של interneurons בneuromeres בטן של העובר מאוחר תסיסנית 14. מחקר זה, שחשף גם את השונות מורפולוגית של סוגי תאים עצביים בודדים ועקרונות של ארגון neuromere במערכת העצבים המרכזיים של העובר, לא היה אפשרי עם כל שיטת תיוג זמינה אחרת.

<table border="1" style=";text-align:right;direction:rtl"><tbody><tr><td> שם מגיב / חומרים </td><td> חברה </td><td> מספר קטלוגים </td><td> תגובות </td></tr><tr><td></td><td></td><td></td><td> ריאגנטים </td></tr><tr><td> DAB </td><td> סיגמה אולדריץ </td><td> <a href="http://www.sigmaaldrich.com/catalog/product/sigma/d5905" target="_blank">D-5905</a> </td><td> 2-3 מ&quot;ג / מ&quot;ל ​​ב100 מ&quot;מימ TRIS-HCL, pH 7.4 </td></tr><tr><td> DiI </td><td> Invitrogen בדיקות / מולקולריות </td><td> <a href="http://products.invitrogen.com/ivgn/product/D282" target="_blank">D-282</a> </td><td> 1 מ&quot;ג / מ&quot;ל ​​באתנול </td></tr><tr><td> <a href="http://www.merckmillipore.com/germany/chemicals/formaldehydloesung-min-37-prozent/MDA_CHEM-104002/p_Wt6b.s1L6pAAAAEWpOEfVhTl" target="_blank">פורמלדהיד</a> </td><td> מרק Millipore </td><td></td><td> 7,4% ב PBS </td> &lt;/ P&gt; <tr><td> גליצרול </td><td> <a href="http://www.carlroth.com/catalogue/" target="_blank">רוט</a> </td><td> 3783 </td><td> 70% ב PBS </td></tr><tr><td> Heptane דבק </td><td> Beiersdorf AG </td><td> צ&#39;לו 31-39-30 * </td><td> לדלל CA. 1-1 עם n-heptane </td></tr><tr><td> PBS </td><td></td><td></td><td> 1x </td></tr><tr><td> TRIS </td><td> <a href="http://www.carlroth.com/catalogue/" target="_blank">רוט</a> </td><td> 4855 </td><td></td></tr><tr><td> Vectashield </td><td> מעבדות וקטור </td><td> <a href="http://www.vectorlabs.com/catalog.aspx?prodID=427" target="_blank">H1000</a> </td><td></td></tr><tr><td></td><td></td><td></td><td> ציוד </td></tr><tr><td> מיקרוסקופ confocal </td><td> ליקה </td><td> TCS SP2 </td><td> כדי להציג ולתעד labelings בפלואורסצנטי </td></tr><tr><td> DC -mplifier </td><td> דרגן חברה </td><td> אבן הפינה ION-100 </td><td> כדי לבצע את labelings </td></tr><tr><td> Coverslips </td><td> <a href="http://www.menzel.de/" target="_blank">מנזל</a> </td><td> BB018018A1 </td><td> 18 מ&quot;מ x 18 </td></tr><tr><td> Coverslips </td><td> <a href="http://www.menzel.de/" target="_blank">מנזל</a> </td><td> BB024060A1 </td><td> 24 מ&quot;מ x 60 </td></tr><tr><td> ביתור מיקרוסקופ </td><td> ליקה </td><td> MZ8 </td><td> להכין וdisect עוברים </td></tr><tr><td> נימים שטוחות </td><td> <a href="http://www.hilgenberg-gmbh.de/" target="_blank">Hilgenberg</a> </td><td></td><td> מ&quot;מ קוטר חיצוני 1; עובי 0.1 מ&quot;מ glas </td></tr><tr><td> הזרקת נימים </td><td> מוצרי מדע </td><td> 100 GB TF 8P </td><td></td></tr><tr><td> R micromanipulator </td><td> ליקה </td><td></td><td> כדי לבצע את labelings </td></tr><tr><td> דגם P-97 </td><td> סאטר מכשירים </td><td></td><td> למשוך נימים </td></tr><tr><td> שקופיות אובייקטים </td><td> <a href="http://www.marienfeld-superior.com/" target="_blank">Marienfeld Superior</a> </td><td> 1000000 </td><td></td></tr><tr><td> מיקרוסקופ המדעי </td><td> Zeiss </td><td> Axioskop 2 mot </td><td> כדי להציג labelings לאחר photoconversion </td></tr><tr><td> מיקרוסקופ המדעי </td><td> אולימפוס </td><td> BX50 </td><td> כדי לבצע את labelings </td></tr><tr><td> מצלמת וידאו הסוני MC3255 </td><td> סוני / AVT הורן </td><td> סוני MC3255 </td><td> כדי להקליט labelings לאחר photoconversion </td></tr></tbody></table> טבלת 1.

labeling of single cells in the central nervous system of drosophila melanogaster

במאמר זה יתאר כיצד בנפרד לתייג תאי עצב במערכת העצבים המרכזיים העוברית של דרוזופילה melanogaster על ידי ההזרקה של juxtacellular DiI סמן lipophilic ניאון הממברנה. שיטה זו מאפשרת הדמיה של מורפולוגיה של תאים עצבית בפירוט רב. אפשר לתייג את כל תאים במערכת העצבים מרכזיים: גופי תא של הנוירונים היעד הם דמיינו תחת אופטיקה DIC או על ידי ביטוי של סמן גנטי ניאון כגון ה-GFP. לאחר תיוג, DiI יכול להפוך לחום קבוע כתם ידי photoconversion כדי לאפשר הדמיה של מורפולוגיה תא עם אור מועבר ואופטיקה DIC. לחלופין, את התאים שכותרת DiI-ניתן לצפות ישירות עם מיקרוסקופיה confocal, המאפשרים חלבוני כתב ניאון הציגו גנטי להיות colocalised. הטכניקה יכולה לשמש בכל בעל חיים, ללא קשר לגנוטיפ, כך שניתן לנתח פנוטיפים מוטציה ברזולוצית תא בודדה.

איור 4 ממחיש תוצאות טיפוסיות של הטכניקה, שאנו מתארים כאן. האיור 4A מראה דוגמה של interneuron אחת DiI מלא שphotoconverted נקי. זה יפה ממחיש את כמות הפרטים אלה הצעת הכנות. כאשר מתחת אופטיקה DIC ההקשר המרחבי של התא המסומן בתוך הרקמה הסובבת הלא מתויגת-הופך לגלוי, למשל</em...

Watch this Scientific Journal Video about Labeling of Single Cells in the Central Nervous System of Drosophila melanogaster at JoVE.com

Labeling of Single Cells in the Central Nervous System of Drosophila melanogaster

אנו מציגים טכניקה לסימון תאי עצב בודדים במערכת העצבים המרכזית (CNS) של<em&gt; דרוזופילה</em&gt; עוברים, המאפשר הניתוח של מורפולוגיה עצבית על ידי אור או שודר או מיקרוסקופיה confocal.

תיוג של תאים בודדים במערכת העצבים המרכזית של<em&gt; דרוזופילה melanogaster</em

In this  article we describe how to individually label neurons in the embryonic CNS of Drosophila melanogaster by juxtacellular injection of the ...

labeling-single-cells-central-nervous-system-drosophila

Universidad de Cartagena UDEC

Research

JoVE Journal

Neuroscience

Labeling of Single Cells in the Central Nervous System of Drosophila melanogaster

12.7K Views.  University of Mainz. אנו מציגים טכניקה לסימון תאי עצב בודדים במערכת העצבים המרכזית (CNS) של דרוזופילה עוברים, המאפשר הניתוח של מורפולוגיה עצבית על ידי אור או שודר או מיקרוסקופיה confocal.

Video: Labeling of Single Cells in the Central Nervous System of Drosophila melanogaster

Dissection of Larval CNS in Drosophila Melanogaster

The central nervous system (CNS) of Drosophila larvae is complex and poorly understood.  One way to investigate the CNS is to use immunohistochemistry to examine the expression of various novel and marker proteins.  Staining of whole larvae is impractical because the tough cuticle prevents antibodies from penetrating inside the body cavity.  In order to stain these tissues it is necessary to dissect the animal prior to fixing and staining.  In this article we demonstrate how to dissect Drosophila larvae without damaging the CNS.  Begin by tearing the larva in half with a pair of fine forceps, and then turn the cuticle "inside-out" to expose the CNS.  If the dissection is performed carefully the CNS will remain attached to the cuticle.  We usually keep the CNS attached to the cuticle throughout the fixation and staining steps, and only completely remove the CNS from the cuticle just prior to mounting the samples on glass slides.  We also show some representative images of a larval CNS stained with Eve, a transcription factor expressed in a subset of neurons in the CNS.  The article concludes with a discussion of some of the practical uses of this technique and the potential difficulties that may arise.

In this article we demonstrate how to dissect the central nervous system from third instar Drosophila larvae.

Dissection של מערכת העצבים המרכזית הזחל ב תסיסנית melanogaster

The central nervous system (CNS) of Drosophila larvae is complex and poorly understood.  One way to investigate the CNS is to use immunohistochemistry to ...

Biology

Purification of Low-abundant Cells in the Drosophila Visual System

Recent improvements in the sensitivity of next generation sequencing have facilitated the application of transcriptomic and genomic analyses to small numbers of cells. Utilizing this technology to study development in the Drosophila visual system, which boasts a wealth of cell type-specific genetic tools, provides a powerful approach for addressing the molecular basis of development with precise cellular resolution. For such an approach to be feasible, it is crucial to have the capacity to reliably and efficiently purify cells present at low abundance within the brain. Here, we present a method that allows efficient purification of single cell clones in genetic mosaic experiments. With this protocol, we consistently achieve a high cellular yield after purification using fluorescence activated cell sorting (FACS) (~25% of all labeled cells), and successfully performed transcriptomics analyses on single cell clones generated through mosaic analysis with a repressible cell marker (MARCM). This protocol is ideal for applying transcriptomic and genomic analyses to specific cell types in the visual system, across different stages of development and in the context of different genetic manipulations.

Here, we present a cell dissociation protocol for efficiently isolating cells present at low abundance within the Drosophila visual system through fluorescence activated cell sorting (FACS).

טיהור של התאים נמוך-שפע ב מערכת הראייה דרוזופילה

Recent improvements in the sensitivity of next generation sequencing have facilitated the application of transcriptomic and genomic analyses to small ...

Genetics

Cell Lineage Analyses and Gene Function Studies Using Twin-spot MARCM

Mosaic analysis with a repressible cell marker (MARCM) is a positive mosaic labeling system that has been widely applied in Drosophila neurobiological studies to depict intricate morphologies and to manipulate the function of genes in subsets of neurons within otherwise unmarked and unperturbed organisms. Genetic mosaics generated in the MARCM system are mediated through site-specific recombination between homologous chromosomes within dividing precursor cells to produce both marked (MARCM clones) and unmarked daughter cells during mitosis. An extension of the MARCM method, called twin-spot MARCM (tsMARCM), labels both of the twin cells derived from a common progenitor with two distinct colors. This technique was developed to enable the retrieval of useful information from both hemi-lineages. By comprehensively analyzing different pairs of tsMARCM clones, the tsMARCM system permits high-resolution neural lineage mapping to reveal the exact birth-order of the labeled neurons produced from common progenitor cells. Furthermore, the tsMARCM system also extends gene function studies by permitting the phenotypic analysis of identical neurons of different animals. Here, we describe how to apply the tsMARCM system to facilitate studies of neural development in Drosophila.

Here, we present a protocol for a mosaic labeling technique that permits the visualization of neurons derived from a common progenitor cell in two distinct colors. This facilitates neural lineage analysis with the capability of birth-dating individual neurons and studying gene function in the same neurons of different individuals.

ניתוח שושלת תא ולימודי תפקוד גן באמצעות Twin-spot MARCM

Mosaic analysis with a repressible cell marker (MARCM) is a positive mosaic labeling system that has been widely applied in Drosophila neurobiological ...

Developmental Biology

Application of MultiColor FlpOut Technique to Study High Resolution Single Cell Morphologies and Cell Interactions of Glia in Drosophila

Cells display different morphologies and complex anatomical relationships. How do cells interact with their neighbors? Do the interactions differ between cell types or even within a given type? What kinds of spatial rules do they follow? The answers to such fundamental questions in vivo have been hampered so far by a lack of tools for high resolution single cell labeling. Here, a detailed protocol to target single cells with a MultiColor FlpOut (MCFO) technique is provided. This method relies on three differently tagged reporters (HA, FLAG and V5) under UAS control that are kept silent by a transcriptional terminator flanked by two FRT sites (FRT-stop-FRT). A heat shock pulse induces the expression of a heat shock-induced Flp recombinase, which randomly removes the FRT-stop-FRT cassettes in individual cells: expression occurs only in cells that also express a GAL4 driver. This leads to an array of differently colored cells of a given cell type that allows the visualization of individual cell morphologies at high resolution. As an example, the MCFO technique can be combined with specific glial GAL4 drivers to visualize the morphologies of the different glial subtypes in the adult Drosophila brain.

Application of MultiColor FlpOut Technique to Study High Resolution Single Cell Morphologies and Cell Interactions of Glia in Drosophila

Cells display different morphologies and establish a variety of interactions with their neighbors. This protocol describes how to reveal the morphology of single cells and to investigate cell-cell interaction by using the well-established Gal4/UAS expression system.

יישום טכניקת FlpOut ססגוניות ללמוד ברזולוציה גבוהה תא בודד מורפולוגיות ו תא אינטראקציות של עכשיו, דונלד ב דרוזופילה

Cells display different morphologies and complex anatomical relationships. How do cells interact with their neighbors? Do the interactions differ between ...

Behavior

Immunohistological Labeling of Microtubules in Sensory Neuron Dendrites, Tracheae, and Muscles in the Drosophila Larva Body Wall

To understand how differences in complex cell shapes are achieved, it is important to accurately follow microtubule organization. The Drosophila larval body wall contains several cell types that are models to study cell and tissue morphogenesis. For example tracheae are used to examine tube morphogenesis1, and the dendritic arborization (DA) sensory neurons of the Drosophila larva have become a primary system for the elucidation of general and neuron-class-specific mechanisms of dendritic differentiation2-5 and degeneration6.
The shape of dendrite branches can vary significantly between neuron classes, and even among different branches of a single neuron7,8. Genetic studies in DA neurons suggest that differential cytoskeletal organization can underlie morphological differences in dendritic branch shape4,9-11. We provide a robust immunological labeling method to assay in vivo microtubule organization in DA sensory neuron dendrite arbor (Figures 1, 2, Movie 1). This protocol illustrates the dissection and immunostaining of first instar larva, a stage when active sensory neuron dendrite outgrowth and branching organization is occurring 12,13.
In addition to staining sensory neurons, this method achieves robust labeling of microtubule organization in muscles (Movies 2, 3), trachea (Figure 3, Movie 3), and other body wall tissues. It is valuable for investigators wishing to analyze microtubule organization in situ in the body wall when investigating mechanisms that control tissue and cell shape.

Immunohistological Labeling of Microtubules in Sensory Neuron Dendrites, Tracheae, and Muscles in the Drosophila Larva Body Wall

To understand how complex cell shapes, such as neuronal dendrites, are achieved during development, it is important to be able to accurately assay microtubule organization. Here we describe a robust immunohistological labeling method to examine microtubule organization of dendritic arborization neuron sensory dendrites, trachea, muscle, and other Drosophila larva body wall tissues.

תיוג Immunohistological של microtubules ב דנדריטים Neuron חושי, Tracheae, והשרירים של תסיסנית גוף Larva הכותל

To understand how differences in complex cell shapes are achieved, it is important to accurately follow microtubule organization. The Drosophila larval ...

Dissection and Immunofluorescent Staining of Mushroom Body and Photoreceptor Neurons in Adult Drosophila melanogaster Brains

Nervous system development involves a sequential series of events that are coordinated by several signaling pathways and regulatory networks. Many of the proteins involved in these pathways are evolutionarily conserved between mammals and other eukaryotes, such as the fruit fly Drosophila melanogaster, suggesting that similar organizing principles exist during the development of these organisms. Importantly, Drosophila has been used extensively to identify cellular and molecular mechanisms regulating processes that are required in mammals including neurogenesis, differentiation, axonal guidance, and synaptogenesis. Flies have also been used successfully to model a variety of human neurodevelopmental diseases. Here we describe a protocol for the step-by-step microdissection, fixation, and immunofluorescent localization of proteins within the adult Drosophila brain. This protocol focuses on two example neuronal populations, mushroom body neurons and retinal photoreceptors, and includes optional steps to trace individual mushroom body neurons using Mosaic Analysis with a Repressible Cell Marker (MARCM) technique. Example data from both wild-type and mutant brains are shown along with a brief description of a scoring criteria for axonal guidance defects. While this protocol highlights two well-established antibodies for investigating the morphology of mushroom body and photoreceptor neurons, other Drosophila brain regions and the localization of proteins within other brain regions can also be investigated using this protocol.

Dissection and Immunofluorescent Staining of Mushroom Body and Photoreceptor Neurons in Adult Drosophila melanogaster Brains

This protocol describes the dissection and immunostaining of adult Drosophila melanogaster brain tissues. Specifically, this protocol highlights the use of Drosophila mushroom body and photoreceptor neurons as example neuronal subsets that can be accurately used to uncover general principles underlying many aspects of neuronal development.

ניתוח Immunofluorescent מכתים של פטריות הגוף והנפש ומה קולט אור נוירונים במוח למבוגרים דרוזופילה melanogaster

Nervous system development involves a sequential series of events that are coordinated by several signaling pathways and regulatory networks. Many of the ...

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.

הדמיה חיה של<em&gt; דרוזופילה</em&gt; הזחל Neuroblasts

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

Electrophysiological Recording of The Central Nervous System Activity of Third-Instar Drosophila Melanogaster

The majority of the currently available insecticides target the nervous system and genetic mutations of invertebrate neural proteins oftentimes yield deleterious consequences, yet the current methods for recording nervous system activity of an individual animal is costly and laborious. This suction electrode preparation of the third-instar larval central nervous system of Drosophila melanogaster, is a tractable system for testing the physiological effects of neuroactive agents, determining the physiological role of various neural pathways to CNS activity, as well as the influence of genetic mutations to neural function. This ex vivo preparation requires only moderate dissecting skill and electrophysiological expertise to generate reproducible recordings of insect neuronal activity. A wide variety of chemical modulators, including peptides, can then be applied directly to the nervous system in solution with the physiological saline to measure the influence on the CNS activity. Further, genetic technologies, such as the GAL4/UAS system, can be applied independently or in tandem with pharmacological agents to determine the role of specific ion channels, transporters, or receptors to arthropod CNS function. In this context, the assays described herein are of significant interest to insecticide toxicologists, insect physiologists, and developmental biologists for which D. melanogaster is an established model organism. The goal of this protocol is to describe an electrophysiological method to enable the measurement of electrogenesis of the central nervous system in the model insect, Drosophila melanogaster, which is useful for testing a diversity of scientific hypotheses.

Electrophysiological Recording of The Central Nervous System Activity of Third-Instar  Drosophila Melanogaster

This protocol describes a method to record the descending electrical activity of the Drosophila melanogaster central nervous system to enable the cost-efficient and convenient testing of pharmacological agents, genetic mutations of neural proteins, and/or the role of unexplored physiological pathways.

הקלטה אלקטרופיזיולוגיות של פעילות מערכת העצבים המרכזית השלישית-לחלל דרוזופילה Melanogaster

The majority of the currently available insecticides target the nervous system and genetic mutations of invertebrate neural proteins oftentimes yield ...

Stimulating and Analyzing Adult Neurogenesis in the Drosophila Central Brain

The molecular and cellular mechanisms underlying neurogenesis in response to disease or injury are not well understood. However, understanding these mechanisms is crucial for developing neural regenerative therapies. Drosophila melanogaster is a leading model for studies of neural development but historically has not been exploited to investigate adult brain regeneration. This is primarily because the adult brain exhibits very low mitotic activity. Nonetheless, penetrating traumatic brain injury (PTBI) to the adult Drosophila central brain triggers the generation of new neurons and new glia. The powerful genetic tools available in Drosophila combined with the simple but rigorous injury protocol described here now make adult Drosophila brain a robust model for neural regeneration research. Provided here are detailed instructions for (1) penetrating injuries to the adult central brain and (2) dissection, immunohistochemistry, and imaging post-injury. These protocols yield highly reproducible results and will facilitate additional studies to dissect mechanisms underlying neural regeneration.

Stimulating and Analyzing Adult Neurogenesis in the Drosophila Central Brain

This article provides detailed protocols for inflicting Penetrating Traumatic Brain Injury (PTBI) to adult Drosophila and examining the resulting neurogenesis.

גירוי וניתוח נוירוגנזה למבוגרים במוח המרכזי של דרוזופילה

The molecular and cellular mechanisms underlying neurogenesis in response to disease or injury are not well understood. However, understanding these ...

Retrograde Labeling of Drosophila Motor Neurons Using Fluorescent Lipophilic Dyes

<a href='https://appjove.unicartagenaproxy.elogim.com/v/60716/retrograde-tracing-drosophila-embryonic-motor-neurons-using'>Source: Inal, M. A. et. al., Retrograde Tracing of Drosophila Embryonic Motor Neurons Using Lipophilic Fluorescent Dyes. J. Vis. Exp. (2020)</a>This video demonstrates the method of retrograde labeling of Drosophila motor neurons using lipophilic dyes. The dye enters the axon, travels retrogradely, and labels the neurons green, allowing detailed visualization of their morphology and projections under a confocal microscope.

Source: Inal, M. A. et. al., Retrograde Tracing of Drosophila Embryonic Motor Neurons Using Lipophilic Fluorescent Dyes. J. Vis. Exp. (2020)This video ...

תיוג של תאים בודדים במערכת העצבים המרכזית של

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