המחברים מבקשים להודות Ikiel Ptito לקבלת תמיכה טכנית המשיכה שלו.

חלק 1: טרום עיבוד של רקמות <ol style=";text-align:right;direction:rtl"><li style=";text-align:right;direction:rtl"> רקמות צריך להיות perfused היטב עם paraformaldehyde, glutaraldehyde, או פורמלין. זו יכולה להיות מושגת באמצעות זלוף transcardial הסטנדרטית המשמשת בדרך כלל כדי לקצור איברים אחרים. במחקר הנוכחי הנושא היה מורדמת עם קטמין הידרוכלוריד (10 מ&quot;ג / ק&quot;ג, IM), מורדמים עם ממנת יתר של pentobarbital נתרן (25 מ&quot;ג / ק&quot;ג, ד) ו perfused transcardially עם 0.1 מ PBS עד exsanguinated לחלוטין. זה ואחריו פתרון paraformaldehyde 4% PBS דקות 5 (~ 1 ליטר). </li><li style=";text-align:right;direction:rtl"> המוח צריך להיות חסום stereotaxically, להסיר הגולגולת, שקל, ונפח נחוש 2. רקמה אמור להיות cryoprotected בפתרונות סוכרוז מדורגים עם ריכוז סופי של סוכרוז 30% חיץ פוספט. גושי רקמה אמור להיות קפוא isopentane ב -65 ° C ו המאוחסן ב -80 ° C עד שאתה מוכן לחתוך. </li></ol> חלק 2: דגימה שיטתית <ol style=";text-align:right;direction:rtl"><li style=";text-align:right;direction:rtl"> דגימה שיטתית מסתמך על תוכנית ראויה לפני חתך. האזור כולו של עניין הן את גודל מקורי, הזנב ואת הגב, הגחון צריך להיות מוגדר היטב. אטלס stereotaxic של המין בשימוש עוזר להגדיר את האזור של הריבית כדי לקבוע את אורך האזור של עניין. לאחר אורך האזור של עניין נקבע יש צורך להקים את מרווח הדגימה. לדוגמה, מאטלס stereotaxic לך לקבוע כי היקף כולו rostro-הזנב של האזור של עניין הוא כ 3 מ&quot;מ אורך, כאשר חתך במישור העטרה. לאחר מכן אתה מחליט להגדיר את cryostat סעיף ב 50μm, בקצב הזה יהיה לך 60 סעיפים המכסים את היקף מקורי, הזנב של האזור של עניין. לצורך המחקר טיפוסי stereological משוחדת 6-10 חלקים שנדגמו באופן שיטתי נדרשים לקבל תוצאות אמינות. בדוגמה זו 10 סעיפים ברווחים שווים לאורך כל תוצאה rostro-הזנב היקף במרווח דגימה של 1 / 6 חלקים. </li></ol> חלק 3: יצירה של הבנק המוח <ol style=";text-align:right;direction:rtl"><li style=";text-align:right;direction:rtl"> לאחר תדר הדגימה סעיף נקבע, הצעד הבא הוא לקבוע איזה כתמים יבוצע מיד. לדוגמה, אם אתה מכתים עם סגול cresyl תוכלו לתפוס 1 / 6 חלקים בשקופית. זו תהיה סדרה 1. אם אתה יודע אולי יש נוגדנים פוטנציאל כמה שאתה רוצה לרוץ, אבל גם רוצה לראות את הנתונים מספק כתם הראשי שלך או פשוט אין לך את הזמן כדי לבצע את אימונוהיסטוכימיה, במקום סדרה הנותרים 5 סעיפים כדי שהם היו פרוסים לשמר אנטיגן בארות המכיל (1 פוליוויניל% pyrrolidone, אתילן גליקול 50% o.1M PBS, Ph 7.4). הטבלה שלהלן מספקת תוכנית דגימה המורכב שקופית 1 ו 1 צלחת (צלחת 48-היטב רגיל) סעיפים כחלק הבנק מוח (טבלה 1). <img src="/files/ftp_upload/1260/table1.jpg" alt="טבלה 1" /> טבלה 1. הדגימה סעיף תדר כאן הוא 1 / 6. בסעיף הראשון מושם על שקופית עבור מכתים סגול cresyl ולמחרת 5 סעיפים ממוקמות לתוך אנטיגן לשמר. מספר סעיף 7 הוא נתפס אז בשקופית וחתכים 9-12 ממוקמים אנטיגן לשמר. מחזור זה חוזר על עצמו עד לאזור של עניין או בלוק של רקמות מחולק ממצה. </li><li style=";text-align:right;direction:rtl"> השלב הבא הוא סעיף ממצה את גוש של רקמה. מניחים כמות קטנה של הטבעה בינוני על צ&#39;אק ולתת לו להקפיא. מניחים את צ&#39;אק לתוך הראש microtome ו לגלח מספיק של המדיום קפוא יש משטח שטוח. הסר את צ&#39;אק מהראש microtome מקום שטוח בתוך cryostat. יוצקים הטבעה בינוני על צ&#39;אק ועל בתוקף במקום לחסום את המוח עם הצד החתוך stereotaxically ממוצבת ביובש על צ&#39;אק. לאט לחלוטין להטביע את המוח הרכבה בינונית. מניחים את צ&#39;אק עם המוח לתוך הראש microtome וסעיף באמצעות פרמטרים שנקבעו מראש עבור תדר הדגימה סעיף. </li><li style=";text-align:right;direction:rtl"> לאחר בלוק של רקמות כבר מחולק ממצה וחתכים ממוקמות לתוך בארות, מכסים את הצלחת עם מכסה, עוטפים את המכסה עם parafilm, ומכניסים למקפיא -20 מעלות צלזיוס. התחבר המספר הכולל של החלקים שצולמו עבור כל סדרה, רווח בסעיף הדגימה, ומספר הסדרה עבור כל חיה. יומן זה יהיה חיוני כדי לעקוב אחר הסרת הבאות של חלקים מהבנק המוח אימונוהיסטוכימיה בעתיד. </li></ol> חלק 4: תוצאות נציג: דגימה שיטתית באופן זה כבר מקובל במעבדה שלנו במשך 3 השנים האחרונות. היו לנו הרבה הצלחה בביצוע אימונוהיסטוכימיה על החומר אוחסן ב אנטיגן לשמר שלוש שנים אחרי זה היה פרוס ללא הידרדרות של האות (איור 1). יתר על כן, כחלק הבנק המוח שלנו יש לנו מחובר קרוב ל 20,000 חלקים שיטתית של הלא אנושיהמוח הפרימטים (איור 2) במסגרת תוכניות לטווח ארוך המחקר שלנו. <img src="/files/ftp_upload/1260/1260figure1.jpg" alt="" /> באיור 1. זהו קטע מן המוט העורפית של פרימטים לא אנושיים immunostained עבור NeuN מראה הן שכבה VI ו ביניים נוירונים החומר הלבן. סעיף זה בפרט אוחסן אנטיגן לשמור על -20 מעלות צלזיוס למשך תקופה של 2 שנים. בר סולם = 100μm. <img src="/files/ftp_upload/1260/1260fig2.jpg" alt="figure2" /> איור 2. Vervet הבנק המוח שלנו מורכב כיום קרוב ל 20,000 חלקים שיטתית מ 30 קופים.

<ol>
	<li>West, M., Slomianka, L., Gundersen, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Unbiased+stereological+estimation+of+the+total+number+of+neurons+in+the+subdivisions+of+the+rat+hippocampus+using+the+optical+fractionator.+Aat+Rec.">Unbiased stereological estimation of the total number of neurons in the subdivisions of the rat hippocampus using the optical fractionator. Aat Rec.</a> 231, 482-497 (1991).</li><li>Burke, M. W., Zangenehpour, S., Boire, D., Ptito, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Dissecting+the+Non-human+Primate+Brain+in+Stereotaxic+Space.">Dissecting the Non-human Primate Brain in Stereotaxic Space.</a> J Vis Exp. , (2009).</li></ol>

The authors have nothing to disclose.

דגימה שיטתית היא שיטה זולה שנועדה למקסם את חומר מחקרי. אסטרטגיה זו הדגימה נועדה לציית לכללי stereology משוחדת הדורש דגימה שיטתי ברחבי האזור. זה קריטי, כי את סדר הסעיפים עבור כל סידרת נשמר. במעבדה שלנו השתמשנו בהצלחה בשיטה זו עבור הבנקאות הן אוגר והלא אנושי חלקים במוח הפרימ...

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<td>Ethylene glycol</td>
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<td>Thermal Scientific Embedding Medium</td>
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brain banking: making the most of your research specimens

Stereology משוחדת היא שיטה מדויקת ויעילה לאמוד את מספר נוירון הכולל (או סוג תא אחר) באזור מסוים של עניין<sup&gt; 1</sup&gt;. כדי להשיג מטרה זו בסעיפים 60-10 שיטתי צריך להיות נחקר מכסה את המבנה כולו. בדרך כלל זה כרוך עיבוד 1 / 5 חלקים אשר משאיר כמות משמעותית של חומר לא מעובד. על מנת למקסם את החומר, אנו מציעים שיטה זולה לשימור רקמות קבוע במסגרת תוכנית ארוכת טווח מחקר אחסון. כאשר הרקמה פרוס ומעובד הרצוי, קטעי כתם או נוגדן חלופי צריך להיות ממוקם באופן שיטתי אנטיגן לשמור על -20 ° C עבור עיבוד עתידי. שימוש של 24 צלחות היטב, קטעי ניתן להציב על מנת בעתיד אחזור. באמצעות שיטה זו, רקמות יכול להיות מאוחסן ומעובד אימונוהיסטוכימיה במשך שנים.

Watch this Scientific Journal Video about Brain Banking: Making the Most of your Research Specimens at JoVE.com

Brain Banking: Making the Most of your Research Specimens

בנקאות המוח דגימה שיטתית של חומר ביולוגי מספק את הבסיס stereology משוחדת למקסימום את פוטנציאל נתונים המתקבלים הדגימה כל אחד.

בנקאות המוח: הפיכת רוב דוגמאות המחקר שלך

Unbiased stereology is a method for accurately and efficiently estimating the total neuron number (or other cell type) in a given area of interest1.  To ...

brain-banking-making-the-most-of-your-research-specimens

Research

JoVE Journal

Biology

9.9K Views.  University of Montreal. בנקאות המוח דגימה שיטתית של חומר ביולוגי מספק את הבסיס stereology משוחדת למקסימום את פוטנציאל נתונים המתקבלים הדגימה כל אחד.

Video: Brain Banking: Making the Most of your Research Specimens

Fabrication of a Microfluidic Device for the Compartmentalization of Neuron Soma and Axons

In this video, we demonstrate the technique of soft lithography with polydimethyl siloxane (PDMS) which we use to fabricate a microfluidic device for culturing neurons. Previously, a silicon wafer was patterned with the design for the neuron microfluidic device using SU-8 and photolithography to create a master mold, or what we simply refer to as a "master". Next, we pour the silicon polymer PDMS on top of the master which is then cured by heating the PDMS to 80&deg;C for 1 hour. The PDMS forms a negative mold of the device. The PDMS is then carefully cut and lifted away from the master. Holes are punched where the reservoirs will be and the excess PDMS trimmed away from the device. Nitrogen is used to blow away any excess debris from the device. At this point the devices are now ready for use and can either bonded to corning No. 1 cover glass with a plasma sterilizer/cleaner or can be reversibly bound to the cover glass by simply placing the device on top of the cover glass. The reversible bonding of the device to glass is covered in a separate video and requires first that the device be sterilized either with 70% ethanol or by autoclaving. Plasma treating sterilizes the devices so no further treatment is necessary. It is, however, important, when plasma-treating the devices, to add liquid to the devices within 10 minutes of the plasma treatment while the surfaces are still hydrophilic. Waiting longer than 10 minutes to add liquid to the device makes it difficult for the liquid to enter the device. The neuron devices are typically plasma-bound to cover glass and 0.5 mg/ml poly-L-lysine (PLL) in pH 8.5 borate buffer is immediately added to the device. After a minimum of 3 hours incubating with PLL, the devices are washed with dH2O water a minimum of 3 times with at least 15 minutes between each wash. Next, the water is removed and fresh media is added to the device. At this point the device is ready for use. It is important to remember at this point to never remove all the media from the device. Always leave media in the main channel.

In this video we demonstrate the technique of soft lithography with polydimethyl siloxane (PDMS) which we use to farbricate a microfluidic device for culturing neurons.

המצאה של המכשיר microfluidic עבור מידור של Neuron סומה האקסונים

In this video, we demonstrate the technique of soft lithography with polydimethyl siloxane (PDMS) which we use to fabricate a microfluidic device for ...

Micro-dissection of Rat Brain for RNA or Protein Extraction from Specific Brain Region

Micro-dissection of rat brain into various regions is extremely important for the study of different neurodegenerative diseases. This video demonstrates micro-dissection of four major brain regions include olfactory bulb, frontal cortex, striatum and hippocampus in fresh rat brain tissue. Useful tips for quick removal of respective regions to avoid RNA and protein degradation of the tissue are given.

Micro-dissection of rat brain into various regions is extremely important for the study of different neurodegenerative diseases. This video demonstrates micro-dissection of four major brain regions include olfactory bulb, frontal cortex, striatum and hippocampus in fresh rat brain tissue. Useful tips for quick removal of respective regions to avoid RNA and protein degradation of the tissue are given.

Micro-דיסקציה של מוח החולדה עבור הפקת RNA או חלבון מן אזור מסוים במוח

Micro-dissection of rat brain into various regions is extremely important for the study of different neurodegenerative diseases. This video demonstrates ...

Single Cell Electroporation in vivo within the Intact Developing Brain

Single-cell electroporation (SCE) is a specialized technique allowing the delivery of DNA or other macromolecules into individual cells within intact tissue, including in vivo preparations. The distinct advantage of this technique is that experimental manipulations may be performed on individual cells while leaving the surrounding tissue unaltered, thereby distinguishing cell-autonomous effects from those resulting from global treatments. When combined with advanced in vivo imaging techniques, SCE of fluorescent markers permits direct visualization of cellular morphology, cell growth, and intracellular events over timescales ranging from seconds to days. While this technique is used in a variety of in vivo and ex vivo preparations, we have optimized this technique for use in Xenopus laevis tadpoles. In this video article, we detail the procedure for SCE of a fluorescent dye or plasmid DNA into neurons within the intact brain of the albino Xenopus tadpole. We also discuss methods to optimize yield, and show examples of live two-photon fluorescence imaging of neurons fluorescently labeled by SCE.

Single-cell electroporation (SCE) is a specialized technique allowing delivery of DNA or other macromolecules into individual cells within intact tissue, including in vivo preparations. Here we detail the procedure for SCE of a fluorescent dye or plasmid DNA into neurons within the intact brain of the Xenopus laevis tadpole.

תא יחיד electroporation in vivo בתוך המוח המתפתח אינטקט

Single-cell electroporation (SCE) is a specialized technique allowing the delivery of DNA or other macromolecules into individual cells within intact ...

Live Imaging of the Zebrafish Embryonic Brain by Confocal Microscopy

In this video, we demonstrate the method our lab has developed to analyze the cell shape changes and rearrangements required to bend and fold the developing zebrafish brain (Gutzman et al, 2008). Such analysis affords a new understanding of the underlying cell biology required for development of the 3D structure of the vertebrate brain, and significantly increases our ability to study neural tube morphogenesis. The embryonic zebrafish brain is shaped beginning at 18 hours post fertilization (hpf) as the ventricles within the neuroepithelium inflate. By 24 hpf, the initial steps of neural tube morphogenesis are complete. Using the method described here, embryos at the one cell stage are injected with mRNA encoding membrane-targeted green fluorescent protein (memGFP). After injection and incubation, the embryo, now between 18 and 24 hpf, is mounted, inverted, in agarose and imaged by confocal microscopy. Notably, the zebrafish embryo is transparent making it an ideal system for fluorescent imaging. While our analyses have focused on the midbrain-hindbrain boundary and the hindbrain, this method could be extended for analysis of any region in the zebrafish to a depth of 
80-100 &mu;m.

In this video, we demonstrate a method by which to analyze the developing vertebrate brain in live zebrafish embryos at single cell resolution by confocal microscopy. This includes the method by which we inject the single-cell zebrafish embryo and subsequently mount and image the developing brain.

חיים הדמיה של המוח על ידי עוברי דג הזברה מיקרוסקופיה confocal

In this video, we demonstrate the method our lab has developed to analyze the cell shape changes and rearrangements required to bend and fold the ...

Using the Horseshoe Crab, Limulus Polyphemus, in Vision Research

The American horseshoe crab, Limulus Polyphemus is one of the oldest creatures on earth, and the animal continues to play an indispensable role in biomedical research. Not only does their blood contain special cells that scientists use to detect bacteriotoxins in our medicines, but their eyes also contain a neural network that has provided much insight about physiological processes operating in our visual system, such as light adaptation and lateral inhibition. The horseshoe crab remains an attractive model for vision research because the animal is large and hardy for an invertebrate, its retinal neurons are big and easily accessible, its visual system is compact and extensively studied, and its visual behavior is well defined. Moreover, the structure and function of the eyes are modulated on a daily basis by a circadian clock in the animal s brain. In short, the visual system of horseshoe crabs is simple enough to be understood yet complex enough to be interesting.
In this video we present three electrophysiological paradigms for investigating the neural basis of vision that can be performed in vivo with Limulus. They are electroretinogram recording, optic nerve recording, and intraretinal recording. Electroretinogram (ERG) recordings measure with a surface electrode the summed electrical response of all cells in the eye to a flash of light. They can be used to monitor the overall sensitivity of the eye for prolong periods of time. Optic nerve recordings measure the spiking activity of single nerve fibers with an extracellular microsuction electrode. They can be used to study visual messages conveyed from the eye to the brain as well as circadian-clock messages fed back from the brain to the eye. Intraretinal recordings measure with an intracellular microelectrode the voltage fluctuations induced by light in individual cells of the eye. They can be used to elucidate cellular mechanisms of retinal processing.

Using the Horseshoe Crab, Limulus Polyphemus, in Vision Research

In this video we perform electroretinogram recording, optic nerve recording, and intraretinal recording with the American horseshoe crab, Limulus Polyphemus. These electrophysiological paradigms can be used for investigating the neural basis of vision in a research or teaching lab.

שימוש סרטן הפרסה, Limulus פוליפמוס, מחקר חזון

The American horseshoe crab, Limulus Polyphemus is one of the oldest creatures on earth, and the animal continues to play an indispensable role in ...

Making Gynogenetic Diploid Zebrafish by Early Pressure

Heterozygosity in diploid eukaryotes often makes genetic studies cumbersome. Methods that produce viable homozygous diploid offspring directly from heterozygous females allow F1 mutagenized females to be screened directly for deleterious mutations in an accelerated forward genetic screen. Streisinger et al.1,2 described methods for making gynogenetic (homozygous) diploid zebrafish by activating zebrafish eggs with ultraviolet light-inactivated sperm and preventing either the second meiotic or the first zygotic cell division using physical treatments (heat or pressure) that deploymerize microtubules. The "early pressure" (EP) method blocks the meiosis II, which occurs shortly after fertilization. The EP method produces a high percentage of viable embryos that can develop to fertile adults of either sex. The method generates embryos that are homozygous at all loci except those that were separated from their centromere by recombination during meiosis I. Homozygous mutations are detected in EP clutches at between 50% for centromeric loci and less than 1% for telomeric loci. This method is reproduced verbatim from the Zebrafish Book3.

This is a method for generating gynogenetic diploid zebrafish embryos (embryos whose only genetic contribution comes from the mother) by blocking the second meiotic division immediately after fertilization with ultraviolet light-inactivated sperm. EP embryos are not fully homozygous due to recombination during the first meiotic division, however they are homozygous at all loci that have not been separated from their centromere by recombination.

ביצוע דיפלואידי דג הזברה Gynogenetic על ידי לחץ מוקדמת

Heterozygosity in diploid eukaryotes often makes genetic studies cumbersome. Methods that produce viable homozygous diploid offspring directly from ...

Long-term Culture of Human Breast Cancer Specimens and Their Analysis Using Optical Projection Tomography

Breast cancer is a leading cause of mortality in the Western world. It is well established that the spread of breast cancer, first locally and later distally, is a major factor in patient prognosis. Experimental systems of breast cancer rely on cell lines usually derived from primary tumours or pleural effusions. Two major obstacles hinder this research: (i) some known sub-types of breast cancers (notably poor prognosis luminal B tumours) are not represented within current line collections; (ii) the influence of the tumour microenvironment is not usually taken into account.
We demonstrate a technique to culture primary breast cancer specimens of all sub-types. This is achieved by using three-dimensional (3D) culture system in which small pieces of tumour are embedded in soft rat collagen I cushions. Within 2-3 weeks, the tumour cells spread into the collagen and form various structures similar to those observed in human tumours1. Viable adipocytes, epithelial cells and fibroblasts within the original core were evident on histology. Malignant epithelial cells with squamoid morphology were demonstrated invading into the surrounding collagen. Nuclear pleomorphism was evident within these cells, along with mitotic figures and apoptotic bodies.
We have employed Optical Projection Tomography (OPT), a 3D imaging technology, in order to quantify the extent of tumour spread in culture. We have used OPT to measure the bulk volume of the tumour culture, a parameter routinely measured during the neo-adjuvant treatment of breast cancer patients to assess response to drug therapy. 
Here, we present an opportunity to culture human breast tumours without sub-type bias and quantify the spread of those ex vivo. This method could be used in the future to quantify drug sensitivity in original tumour. This may provide a more predictive model than currently used cell lines.

We have developed a collagen-based in vitro assay which promotes proliferation and invasion from samples of all breast cancer subtypes. Optical Projection Tomography, a three dimensional microscopy technique was utilised to visualise and quantify tumour expansion. This assay may be used to quantify drug response of individual tumour samples.

לטווח ארוך של תרבות האדם סרטן השד דוגמאות וניתוח שלהם באמצעות טומוגרפיה הקרנה אופטית

Breast cancer is a leading cause of mortality in the Western world. It is well established that the spread of breast cancer, first locally and later ...

Mapping the After-effects of Theta Burst Stimulation on the Human Auditory Cortex with Functional Imaging

Auditory cortex pertains to the processing of sound, which is at the basis of speech or music-related processing1. However, despite considerable recent progress, the functional properties and lateralization of the human auditory cortex are far from being fully understood. Transcranial Magnetic Stimulation (TMS) is a non-invasive technique that can transiently or lastingly modulate cortical excitability via the application of localized magnetic field pulses, and represents a unique method of exploring plasticity and connectivity. It has only recently begun to be applied to understand auditory cortical function 2. 
An important issue in using TMS is that the physiological consequences of the stimulation are difficult to establish. Although many TMS studies make the implicit assumption that the area targeted by the coil is the area affected, this need not be the case, particularly for complex cognitive functions which depend on interactions across many brain regions 3. One solution to this problem is to combine TMS with functional Magnetic resonance imaging (fMRI). The idea here is that fMRI will provide an index of changes in brain activity associated with TMS. Thus, fMRI would give an independent means of assessing which areas are affected by TMS and how they are modulated 4. In addition, fMRI allows the assessment of functional connectivity, which represents a measure of the temporal coupling between distant regions. It can thus be useful not only to measure the net activity modulation induced by TMS in given locations, but also the degree to which the network properties are affected by TMS, via any observed changes in functional connectivity.
Different approaches exist to combine TMS and functional imaging according to the temporal order of the methods. Functional MRI can be applied before, during, after, or both before and after TMS. Recently, some studies interleaved TMS and fMRI in order to provide online mapping of the functional changes induced by TMS 5-7. However, this online combination has many technical problems, including the static artifacts resulting from the presence of the TMS coil in the scanner room, or the effects of TMS pulses on the process of MR image formation. But more importantly, the loud acoustic noise induced by TMS (increased compared with standard use because of the resonance of the scanner bore) and the increased TMS coil vibrations (caused by the strong mechanical forces due to the static magnetic field of the MR scanner) constitute a crucial problem when studying auditory processing. 
This is one reason why fMRI was carried out before and after TMS in the present study. Similar approaches have been used to target the motor cortex 8,9, premotor cortex 10, primary somatosensory cortex 11,12 and language-related areas 13, but so far no combined TMS-fMRI study has investigated the auditory cortex. The purpose of this article is to provide details concerning the protocol and considerations necessary to successfully combine these two neuroscientific tools to investigate auditory processing. 
Previously we showed that repetitive TMS (rTMS) at high and low frequencies (resp. 10 Hz and 1 Hz) applied over the auditory cortex modulated response time (RT) in a melody discrimination task 2. We also showed that RT modulation was correlated with functional connectivity in the auditory network assessed using fMRI: the higher the functional connectivity between left and right auditory cortices during task performance, the higher the facilitatory effect (i.e. decreased RT) observed with rTMS. However those findings were mainly correlational, as fMRI was performed before rTMS. Here, fMRI was carried out before and immediately after TMS to provide direct measures of the functional organization of the auditory cortex, and more specifically of the plastic reorganization of the auditory neural network occurring after the neural intervention provided by TMS. 
Combined fMRI and TMS applied over the auditory cortex should enable a better understanding of brain mechanisms of auditory processing, providing physiological information about functional effects of TMS. This knowledge could be useful for many cognitive neuroscience applications, as well as for optimizing therapeutic applications of TMS, particularly in auditory-related disorders.

Auditory processing is the basis of speech and music-related processing. Transcranial Magnetic Stimulation (TMS) has been used successfully to study cognitive, sensory and motor systems but has rarely been applied to audition. Here we investigated TMS combined with functional Magnetic Resonance Imaging to understand the functional organization of auditory cortex.

מיפוי לאחר הלוואי של תטא Burst גירוי בקורטקס השמיעתי האנושי בהדמיה תפקודית

Auditory cortex pertains to the processing of sound, which is at the basis of speech or music-related processing1. However, despite considerable recent ...

Obtaining Specimens with Slowed, Accelerated and Reversed Aging in the Honey Bee Model

Societies of highly social animals feature vast lifespan differences between closely related individuals. Among social insects, the honey bee is the best established model to study how plasticity in lifespan and aging is explained by social factors.
The worker caste of honey bees includes nurse bees, which tend the brood, and forager bees, which collect nectar and pollen. Previous work has shown that brain functions and flight performance senesce more rapidly in foragers than in nurses. However, brain functions can recover, when foragers revert back to nursing tasks. Such patterns of accelerated and reversed functional senescence are linked to changed metabolic resource levels, to alterations in protein abundance and to immune function. Vitellogenin, a yolk protein with adapted functions in hormonal control and cellular defense, may serve as a major regulatory element in a network that controls the different aging dynamics in workers.
Here we describe how the emergence of nurses and foragers can be monitored, and manipulated, including the reversal from typically short-lived foragers into longer-lived nurses. Our representative results show how individuals with similar chronological age differentiate into foragers and nurse bees under experimental conditions. We exemplify how behavioral reversal from foragers back to nurses can be validated. Last, we show how different cellular senescence can be assessed by measuring the accumulation of lipofuscin, a universal biomarker of senescence.
For studying mechanisms that may link social influences and aging plasticity, this protocol provides a standardized tool set to acquire relevant sample material, and to improve data comparability among future studies.

In honey bee workers, aging depends on social behaviors rather than on chronological age. Here we show how worker-types with very different aging patterns can be obtained and analyzed for cellular senescence.

קבלת דגימות עם מואט, מואצת והפוך הזדקנות של דבורת דבש הדגם

Societies of highly social animals feature vast lifespan differences between closely related individuals. Among social insects, the honey bee is the best ...

Sampling Blood from the Lateral Tail Vein of the Rat

Blood samples are commonly obtained in many experimental contexts to measure targets of interest, including hormones, immune factors, growth factors, proteins, and glucose, yet the composition of the blood is dynamically regulated and easily perturbed. One factor that can change the blood composition is the stress response triggered by the sampling procedure, which can contribute to variability in the measures of interest. Here we describe a procedure for blood sampling from the lateral tail vein in the rat. This procedure offers significant advantages over other more commonly used techniques. It permits rapid sampling with minimal pain or invasiveness, without anesthesia or analgesia. Additionally, it can be used to obtain large volume samples (upwards of 1 ml in some rats), and it may be used repeatedly across experimental days. By minimizing the stress response and pain resulting from blood sampling, measures can more accurately reflect the true basal state of the animal, with minimal influence from the sampling procedure itself.

Blood samples are useful for assessing biomarkers of physiological states or disease in vivo. Here we describe the methodology to sample blood from the lateral tail vein in the rat. This method provides rapid samples with minimal pain and invasiveness.

דגימת דם מוריד הזנב לרוחב של העכברוש

Blood samples are commonly obtained in many experimental contexts to measure targets of interest, including hormones, immune factors, growth factors, ...