وأيد هذا العمل من قبل &quot;الوكالة الوطنية بحوث دي لا&quot; منحة ANR - 09 - JCJC - 0117 - 01 و &quot;دي Neuropôle بحوث Francilien - NERF&quot; منحة لشيري رومان. نشكر فرانسواز يفبفر لتطوير البرمجيات في كيو تي سي + + / ، وبينوت لوران وجانفييه باتيست للمساعدة في تطوير برنامج الإعداد التصوير الضوئي

<ol>
	<li>Grinvald, A., Lieke, E., Frostig, R. D., Gilbert, C. D., Wiesel, T. N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Functional+architecture+of+cortex+revealed+by+optical+imaging+of+intrinsic+signals.">Functional architecture of cortex revealed by optical imaging of intrinsic signals.</a> Nature. 324, 361-364 (1986).</li><li>Rubin, B. D., Katz, 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=Optical+imaging+of+odorant+representations+in+the+mammalian+olfactory+bulb.">Optical imaging of odorant representations in the mammalian olfactory bulb.</a> Neuron. 23, 499-511 (1999).</li><li>Chance, B., Cohen, P., J&amp;246, b. s. i. s., &amp;amp, F., Schoener, B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Intracellular+oxidation-reduction+states+in+vivo.">Intracellular oxidation-reduction states in vivo.</a> Science. 137, 499-508 (1962).</li><li>Shibuki, K., Hishida, R., Murakami, H., Kudoh, M., Kawaguchi, T., Watanabe, M., Watanabe, S., Kouuchi, T., Tanaka, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Dynamic+imaging+of+somatosensory+cortical+activity+in+the+rat+visualized+by+flavoprotein+autofluorescence.">Dynamic imaging of somatosensory cortical activity in the rat visualized by flavoprotein autofluorescence.</a> J. Physiol. 549 (Pt. 3, 919-9127 (2003).</li><li>Uchida, N., Takahashi, Y. K., Tanifuji, M., Mori, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Odor+maps+in+the+mammalian+olfactory+bulb:+domain+organization+and+odorant+structural+features.">Odor maps in the mammalian olfactory bulb: domain organization and odorant structural features.</a> Nat. Neurosci. 3, 1035-1043 (2000).</li><li>Gurden, H., Uchida, N., Mainen, Z. F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Sensory-evoked+intrinsic+optical+signals+in+the+olfactory+bulb+are+coupled+to+glutamate+release+and+uptake.">Sensory-evoked intrinsic optical signals in the olfactory bulb are coupled to glutamate release and uptake.</a> Neuron. 52, 335-345 (2006).</li><li>Soucy, E. R., Albeanu, D. F., Fantana, A. L., Murthy, V. N., Meister, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Precision+and+diversity+in+an+odor+map+on+the+olfactory+bulb.">Precision and diversity in an odor map on the olfactory bulb.</a> Nat. Neurosci. 12, 210-220 (2009).</li><li>Frostig, R. D., Lieke, E. E., Ts'o, D. Y., Grinvald, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cortical+functional+architecture+and+local+coupling+between+neuronal+activity+and+the+microcirculation+revealed+by+in+vivo+high-resolution+optical+imaging+of+intrinsic+signals.">Cortical functional architecture and local coupling between neuronal activity and the microcirculation revealed by in vivo high-resolution optical imaging of intrinsic signals.</a> Proc. Natl. Acad. Sci. U.S.A. 87, 6082-6086 (1990).</li><li>Meister, M., Bonhoeffer, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Tuning+and+topography+in+an+odor+map+on+the+rat+olfactory+bulb.">Tuning and topography in an odor map on the rat olfactory bulb.</a> J. Neurosci. 21, 1351-1360 (2001).</li><li>Dunn, A. K., Devor, A., Bolay, H., Andermann, M. L., Moskowitz, M. A., Dale, A. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Simultaneous+imaging+of+total+cerebral+hemoglobin+concentration,+oxygenation,+and+blood+flow+during+functional+activation.">Simultaneous imaging of total cerebral hemoglobin concentration, oxygenation, and blood flow during functional activation.</a> Opt. Lett. 28, 28-30 (2003).</li><li>L'Heureux, B., Gurden, H., Pain, F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Autofluorescence+imaging+of+NADH+and+flavoproteins+in+the+rat+brain:+insights+from+Monte+Carlo+simulations.">Autofluorescence imaging of NADH and flavoproteins in the rat brain: insights from Monte Carlo simulations.</a> Optics. Express. 17, 9477-9490 (2009).</li><li>Renaud, R., Gurden, H., Chery, R., Bendhamane, M., Martin, C., Pain, F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Multispectral+imaging+of+the+olfactory+bulb+activation:+influence+of+realistic+differential+pathlength+correction+factors+on+the+derivation+of+oxygenation+and+total+hemoglobin+concentration+maps+(Proceedings+Paper).">Multispectral imaging of the olfactory bulb activation: influence of realistic differential pathlength correction factors on the derivation of oxygenation and total hemoglobin concentration maps (Proceedings Paper).</a> , (2011).</li></ol>

ليس لدينا شيء في الكشف عنها.

في هذه المقالة نقدم IOS وفاس لتقنيات التصوير في تسجيلات فيفو الأنشطة ذات رائحة أثار في OB الماوس. لتحقيق هذا الهدف بسيط نسبيا وبأسعار معقولة على نطاق ومجال التصوير الضوئي الإعداد أمر ضروري. الحصول على البيانات والتصوير ويتطلب التدريب لتنفيذ الإجراءات الجراحية الدقيقة ، وتجنب أي ضرر يلحق الجافية أو أنسجة المخ. على وجه الخصوص ، سوف تمتص فوتونات نزيف الرئيسية المسجلة في التصوير وحتى نهاية التجربة. واحد يستفيد من IOS والتصوير FAS هو تجنب حقن استشفاف الفلورسنت التي يمكن أن تؤدي سمية الخلوية أو آثار جانبية غير مرغوب فيها. أنها تجعل من الممكن لمعالجة القضايا المتعلقة خرائط الشمية الترميز المكاني وبالتالي من المحفزات الحسية. خلافا ل2 - DeoxyGlucose التصوير ، لأنها توفر إمكانية الروائح الصور عدة في حيوان واحد. ومع ذلك ، منذ يقتصر الاختراق الفوتون في الأنسجة ، ويتم تقييد IOS وفاس على الجزء الظهري من OBولا يمكن تسجيلها من مناطق البطني. التصوير الضوئي الذاتية إشارة القرار المكانية عروض ممتازة للتصوير في الجسم الحي. التحسينات التقنية الحسابات القلق كمية من المكونات الأوعية الدموية في إشارات الانعكاس 8،9 فضلا عن ديناميات الأوكسجين في الدم وحجم أثناء التنشيط الحسي 10. يتم تطويرها حاليا التصوير Multiwavelength النهج التصوير IOS في مختبرنا لقياس تركيز الهيموجلوبين بشكل كامل مجموع والأوكسجين في التوليد أثناء التنشيط الحسي. وهذه القياسات الطيفية الضوئية إضافة إلى التصوير FAS إعطاء فرصة للرد على العلاقة بين الديناميات التي لم تحل بين الخلايا والأوعية الدموية أثناء التنشيط الحسي 11،12.

<table border="1" style=";text-align:right;direction:rtl"><tbody><tr><td> اسم الوصي </td><td> شركة </td><td> فهرس العدد </td></tr><tr><td> Imalgene </td><td> Merial </td><td></td></tr><tr><td> Rompun </td><td> باير </td><td></td></tr><tr><td> Agarose ، النوع الثالث - A </td><td> سيغما الدريخ </td><td> A9793 - 50G </td></tr><tr><td> Hexanal 98 ٪ </td><td> الدريتش </td><td> 115606 - 100ML </td></tr></tbody></table>

imaging odor-evoked activities in the mouse olfactory bulb using optical reflectance and autofluorescence signals

في المخ وينشط الحواس توزيع السكان بين الخلايا العصبية التي تشارك وحدات وظيفية للترميز من التحفيز. تقنيات التصوير الضوئي وظيفية هي مفيدة لتصور تفعيل هذه الوحدات في القشور الحسية مع القرار المكانية العالية. في هذا السياق ، الإشارات الضوئية الذاتية التي تنشأ عن الآليات الجزيئية مرتبطة neuroenergetics هي على النقيض من مصادر قيمة لتسجيل الخرائط المساحية من المحفزات الحسية على حقول واسعة في الدماغ القوارض. هنا ، نقدم اثنين من التقنيات القائمة على التغيرات في الخصائص البصرية الذاتية للأنسجة المخ أثناء التنشيط. ويتم إنتاج أول الإشارات الضوئية الجوهرية (IOS) عن طريق تغيير المحلية في انعكاس الضوء الأحمر للأسباب التالية : (ط) من خلال استيعاب التغيرات في مستوى الأوكسجين في الدم وحجم الدم (II) الفوتون نثر. بدأ استخدام في الجسم الحي IOS لتسجيل الخرائط المساحية في منتصف عام 1980 مع observatiعلى الخرائط البصرية للبرميل الطولي في الفئران وأعمدة التوجه في القشرة البصرية القط 1. وقد تجلى لاحقا IOS التصوير من سطح البصلة الشمية القوارض الرئيسي (OB) استجابة لodorants من قبل مجموعة لاري كاتز 2. النهج الثاني يعتمد على الإشارات بروتين فلافيني تألق ذاتي (FAS) وذلك بسبب التغييرات في حالة الأكسدة هذه سيطة الأيضية الميتوكوندريا. أكثر تحديدا ، وتستند هذه التقنية على مضان أخضر بسبب حالة من flavoproteins عندما تتأكسد هو متحمس النسيج مع ضوء أزرق. على الرغم من أن هذه الإشارات ربما كانت بين جزيئات الفلورسنت سجلت أول لدراسة نشاط الدماغ عن طريق الدراسات رائدا من احتمالات بريتون وزملاؤه 3 ، فإنه لم يكن حتى وقت قريب التي تم استخدامها لرسم خرائط لتنشيط الدماغ في الجسم الحي. وكان أول تطبيق FAS التصوير إلى القشرة الحسية الجسدية في القوارض استجابة لتحريض من قبل مجموعة hindpaw Katsuei Shibuki 4. نظام حاسة الشم هي من الأهمية المركزية لبقاء الغالبية العظمى من أنواع الكائنات الحية لأنه يتيح الكشف عن كفاءة وتحديد المواد الكيميائية في البيئة (الغذاء والحيوانات المفترسة). الحريق المكشوف هو التتابع الأولى من معالجة المعلومات الشمية في الدماغ. يتلقى التوقعات وارد من الخلايا العصبية الحسية الشمية الأولية التي تكشف عن جزيئات الرائحة المتطايرة. كل الخلايا العصبية الحسية عن نوع واحد فقط من مستقبلات الرائحة والخلايا العصبية التي تحمل نفس النوع من العمليات ترسل مستقبلات أعصابهم لمتناهية الصغر واضحة المعالم نفسه من 3 100μm ~ مؤلفة من neuropil منفصلة ، والكبيبة الشمية (الشكل 1). في العقد الماضي ، عزز IOS التصوير استكشاف الفنية للOB 5 و 6 و 7 والتي أصبحت واحدة من أكثر هياكل الحسية التي شملتها الدراسة. لم يتم تعيين النشاط OB مع التصوير FAS يؤديها حتى الان. هنا ، ثه تبين الخطوات المتتالية لبروتوكول فعال لIOS والتصوير FAS لرسم خريطة للرائحة أثار الأنشطة في OB الماوس.

Watch this Scientific Journal Video about Imaging Odor-Evoked Activities in the Mouse Olfactory Bulb using Optical Reflectance and Autofluorescence Signals at JoVE.com

Imaging Odor-Evoked Activities in the Mouse Olfactory Bulb using Optical Reflectance and Autofluorescence Signals

هذا المقال يعرض بروتوكولات الإشارات الضوئية الذاتية وتألق ذاتي flavoproteins إشارات التصوير لأنشطة رائحة أثارت الخريطة على سطح البصلة الشمية في الفئران.

التصوير رائحة مستدعى الأنشطة في لمبة الماوس الشم باستخدام الانعكاس الضوئية وإشارات تألق ذاتي

In the brain, sensory stimulation activates distributed populations of neurons among functional modules which participate to the coding of the stimulus. ...

imaging-odor-evoked-activities-mouse-olfactory-bulb-using-optical

Research

JoVE Journal

Neuroscience

15.9K Views.  UMR8165 Université Paris Sud 11, Paris Diderot 7 – CNRS. هذا المقال يعرض بروتوكولات الإشارات الضوئية الذاتية وتألق ذاتي flavoproteins إشارات التصوير لأنشطة رائحة أثارت الخريطة على سطح البصلة الشمية في الفئران.

Video: Imaging Odor-Evoked Activities in the Mouse Olfactory Bulb using Optical Reflectance and Autofluorescence Signals

Analyzing Responses of Mouse Olfactory Sensory Neurons Using the Air-phase Electroolfactogram Recording

Animals depend on olfaction for many critical behaviors, such as finding food sources, avoiding predators, and identifying conspecifics for mating and other social interactions. The electroolfactogram (EOG) recording is an informative, easy to conduct, and reliable method to assay olfactory function at the level of the olfactory epithelium. Since the 1956 description of the EOG by Ottoson in frogs1, the EOG recording has been applied in many vertebrates including salamanders, rabbits, rats, mice, and humans (reviewed by Scott and Scott-Johnson, 2002, ref. 2). The recent advances in genetic modification in mice have rekindled interest in recording the EOG for physiological characterization of olfactory function in knock-out and knock-in mice. EOG recordings have been successfully applied to demonstrate the central role of olfactory signal transduction components3-8, and more recently to characterize the contribution of certain regulatory mechanisms to OSN responses9-12.
Odorant detection occurs at the surface of the olfactory epithelium on the cilia of OSNs, where a signal transduction cascade leads to opening of ion channels, generating a current that flows into the cilia and depolarizes the membrane13. The EOG is the negative potential recorded extracellularly at the surface of the olfactory epithelium upon odorant stimulation, resulting from a summation of the potential changes caused by individual responsive OSNs in the recording field2. Comparison of the amplitude and kinetics of the EOG thus provide valuable information about how genetic modification and other experimental manipulations influence the molecular signaling underlying the OSN response to odor. 
Here we describe an air-phase EOG recording on a preparation of mouse olfactory turbinates. Briefly, after sacrificing the mouse, the olfactory turbinates are exposed by bisecting the head along the midline and removing the septum. The turbinate preparation is then placed in the recording setup, and a recording electrode is placed at the surface of the olfactory epithelium on one of the medial turbinates. A reference electrode is electrically connected to the tissue through a buffer solution. A continuous stream of humidified air is blown over the surface of the epithelium to keep it moist. The vapor of odorant solutions is puffed into the stream of humidified air to stimulate the epithelium. Responses are recorded and digitized for further analysis.

The electroolfactogram (EOG) recording is an informative, easy-to-conduct, and reliable way of assessing olfactory function at the level of the olfactory epithelium. This protocol describes a recording setup, mouse tissue preparation, data collection, and basic data analysis.

تحليل الردود من الخلايا العصبية الحسية الشمية ماوس باستخدام تسجيل Electroolfactogram الهواء المرحلة

Animals depend on olfaction for many critical behaviors, such as finding food sources, avoiding predators, and identifying conspecifics for mating and ...

Multiple-mouse Neuroanatomical Magnetic Resonance Imaging

The field of mouse phenotyping with magnetic resonance imaging (MRI) is rapidly growing, motivated by the need for improved tools for characterizing and evaluating mouse models of human disease. MRI is an excellent modality for investigating genetically altered animals. It is capable of whole brain coverage, can be used in vivo, and provides multiple contrast mechanisms for investigating different aspects of neuranatomy and physiology. The advent of high-field scanners along with the ability to scan multiple mice simultaneously allows for rapid phenotyping of novel mutations.
Effective mouse MRI studies require attention to many aspects of experiment design. In this article, we will describe general methods to acquire quality images for mouse phenotyping using a system that images mice concurrently in shielded transmit/receive radio frequency (RF) coils in a common magnet (Bock et al., 2003). We focus particularly on anatomical phenotyping, an important and accessible application that has shown a high potential for impact in many mouse models at our imaging centre. Before we can provide the detailed steps to acquire such images, there are important practical considerations for both in vivo brain imaging (Dazai et al., 2004) and ex vivo brain imaging (Spring et al., 2007) that should be noted. These are discussed below.

Magnetic resonance imaging (MRI) has become an increasingly popular tool for examining the phenotype of genetically altered mice. This article illustrates the methods necessary to achieve high-throughput phenotyping of genetically altered mice using multiple-mouse MRI.

متعددة الماوس تشريحي عصبي التصوير بالرنين المغناطيسي

The field of mouse phenotyping with magnetic resonance imaging (MRI) is rapidly growing, motivated by the need for improved tools for characterizing and ...

Targeting Olfactory Bulb Neurons Using Combined In Vivo Electroporation and Gal4-Based Enhancer Trap Zebrafish Lines

In vivo electroporation is a powerful method for delivering DNA expression plasmids, RNAi reagents, and morpholino anti-sense oligonucleotides to specific regions of developing embryos, including those of C. elegans, chick, Xenopus, zebrafish, and mouse 1. In zebrafish, in vivo electroporation has been shown to have excellent spatial and temporal resolution for the delivery of these reagents 2-7. The temporal resolution of this method is important because it allows for incorporation of these reagents at specific stages in development. Furthermore, because expression from electroporated vectors occurs within 6 hours 7, this method is more timely than transgenic approaches. While the spatial resolution can be extremely precise when targeting a single cell 2, 6, it is often preferable to incorporate reagents into a specific cell population within a tissue or structure. When targeting multiple cells, in vivo electroporation is efficient for delivery to a specific region of the embryo; however, particularly within the developing nervous system, it is difficult to target specific cell types solely through spatially discrete electroporation. Alternatively, enhancer trap transgenic lines offer excellent cell type-specific expression of transgenes 8. Here we describe an approach that combines transgenic Gal4-based enhancer trap lines 8 with spatially discrete in vivo electroporation 7, 9 to specifically target developing neurons of the zebrafish olfactory bulb. The Et(zic4:Gal4TA4,UAS:mCherry)hzm5 (formerly GA80_9) enhancer trap line previously described 8, displays targeted transgenic expression of mCherry mediated by a zebrafish optimized Gal4 (KalTA4) transcriptional activator in multiple regions of the developing brain including hindbrain, cerebellum, forebrain, and the olfactory bulb. To target GFP expression specifically to the olfactory bulb, a plasmid with the coding sequence of GFP under control of multiple Gal4 binding sites (UAS) was electroporated into the anterior end of the forebrain at 24-28 hours post-fertilization (hpf). Although this method incorporates plasmid DNA into multiple regions of the forebrain, GFP expression is only induced in cells transgenically expressing the KalTA4 transcription factor. Thus, by using the GA080_9 transgenic line, this approach led to GFP expression exclusively in the developing olfactory bulb. GFP expressing cells targeted through this approach showed typical axonal projections, as previously described for mitral cells of the olfactory bulb 10. This method could also be used for targeted delivery of other reagents including short-hairpin RNA interference expression plasmids, which would provide a method for spatially and temporally discrete loss-of-function analysis.

Targeting Olfactory Bulb Neurons Using Combined In Vivo Electroporation and Gal4-Based Enhancer Trap Zebrafish Lines

The temporal and spatial resolution of genetic manipulations determines the spectrum of biological phenomena that they can perturb. Here we use temporally and spatially discrete in vivo electroporation, combined with transgenic lines of zebrafish, to induce expression of a GFP transgene specifically in neurons of the developing olfactory bulb.

استهداف الخلايا العصبية عن طريق الجمع بين البصلة الشمية في فيفو Electroporation وGal4 محسن المستندة إلى خطوط مصيدة اسماك الزرد

In vivo electroporation is a powerful method for delivering DNA expression plasmids, RNAi reagents, and morpholino anti-sense oligonucleotides to specific ...

Calcium Imaging of Odor-evoked Responses in the Drosophila Antennal Lobe

The antennal lobe is the primary olfactory center in the insect brain and represents the anatomical and functional equivalent of the vertebrate olfactory bulb1-5. Olfactory information in the external world is transmitted to the antennal lobe by olfactory sensory neurons (OSNs), which segregate to distinct regions of neuropil called glomeruli according to the specific olfactory receptor they express. Here, OSN axons synapse with both local interneurons (LNs), whose processes can innervate many different glomeruli, and projection neurons (PNs), which convey olfactory information to higher olfactory brain regions.
Optical imaging of the activity of OSNs, LNs and PNs in the antennal lobe - traditionally using synthetic calcium indicators (e.g. calcium green, FURA-2) or voltage-sensitive dyes (e.g. RH414) - has long been an important technique to understand how olfactory stimuli are represented as spatial and temporal patterns of glomerular activity in many species of insects6-10. Development of genetically-encoded neural activity reporters, such as the fluorescent calcium indicators G-CaMP11,12 and Cameleon13, the bioluminescent calcium indicator GFP-aequorin14,15, or a reporter of synaptic transmission, synapto-pHluorin16 has made the olfactory system of the fruitfly, Drosophila melanogaster, particularly accessible to neurophysiological imaging, complementing its comprehensively-described molecular, electrophysiological and neuroanatomical properties2,4,17. These reporters can be selectively expressed via binary transcriptional control systems (e.g. GAL4/UAS18, LexA/LexAop19,20, Q system21) in defined populations of neurons within the olfactory circuitry to dissect with high spatial and temporal resolution how odor-evoked neural activity is represented, modulated and transformed22-24.
Here we describe the preparation and analysis methods to measure odor-evoked responses in the Drosophila antennal lobe using G-CaMP25-27. The animal preparation is minimally invasive and can be adapted to imaging using wide-field fluorescence, confocal and two-photon microscopes.

Calcium Imaging of Odor-evoked Responses in the Drosophila Antennal Lobe

We describe an established technique to measure and analyze odor-evoked calcium responses in the antennal lobe of living Drosophila melanogaster.

التصوير الكالسيوم من الردود، رائحة أثار في ذبابة الفاكهة لوب Antennal

The antennal lobe is the primary olfactory center in the insect brain and represents the anatomical and functional equivalent of the vertebrate olfactory ...

Ex Vivo Preparations of the Intact Vomeronasal Organ and Accessory Olfactory Bulb

The mouse accessory olfactory system (AOS) is a specialized sensory pathway for detecting nonvolatile social odors, pheromones, and kairomones. The first neural circuit in the AOS pathway, called the accessory olfactory bulb (AOB), plays an important role in establishing sex-typical behaviors such as territorial aggression and mating. This small (&#60;1 mm3) circuit possesses the capacity to distinguish unique behavioral states, such as sex, strain, and stress from chemosensory cues in the secretions and excretions of conspecifics. While the compact organization of this system presents unique opportunities for recording from large portions of the circuit simultaneously, investigation of sensory processing in the AOB remains challenging, largely due to its experimentally disadvantageous location in the brain. Here, we demonstrate a multi-stage dissection that removes the intact AOB inside a single hemisphere of the anterior mouse skull, leaving connections to both the peripheral vomeronasal sensory neurons (VSNs) and local neuronal circuitry intact. The procedure exposes the AOB surface to direct visual inspection, facilitating electrophysiological and optical recordings from AOB circuit elements in the absence of anesthetics. Upon inserting a thin cannula into the vomeronasal organ (VNO), which houses the VSNs, one can directly expose the periphery to social odors and pheromones while recording downstream activity in the AOB. This procedure enables controlled inquiries into AOS information processing, which can shed light on mechanisms linking pheromone exposure to changes in behavior.

Ex Vivo Preparations of the Intact Vomeronasal Organ and Accessory Olfactory Bulb

The mouse accessory olfactory bulb (AOB) has been difficult to study in the context of sensory coding. Here, we demonstrate a dissection that produces an ex vivo preparation in which AOB neurons remain functionally connected to their peripheral inputs, facilitating research into information processing of mouse pheromones and kairomones.

فيفو السابقين الاستعدادات للالميكعي الجهاز سليمة والإكسسوار الشم لمبة

The mouse accessory olfactory system (AOS) is a specialized sensory pathway for detecting nonvolatile social odors, pheromones, and kairomones. The first ...

Recording Temperature-induced Neuronal Activity through Monitoring Calcium Changes in the Olfactory Bulb of Xenopus laevis

The olfactory system, specialized in the detection, integration and processing of chemical molecules is likely the most thoroughly studied sensory system. However, there is piling evidence that olfaction is not solely limited to chemical sensitivity, but also includes temperature sensitivity. Premetamorphic Xenopus laevis are translucent animals, with protruding nasal cavities deprived of the cribriform plate separating the nose and the olfactory bulb. These characteristics make them well suited for studying olfaction, and particularly thermosensitivity. The present article describes the complete procedure for measuring temperature responses in the olfactory bulb of X. laevis larvae. Firstly, the electroporation of olfactory receptor neurons (ORNs) is performed with spectrally distinct dyes loaded into the nasal cavities in order to stain their axon terminals in the bulbar neuropil. The differential staining between left and right receptor neurons serves to identify the &#947;-glomerulus as the only structure innervated by contralateral presynaptic afferents. Secondly, the electroporation is combined with focal bolus loading in the olfactory bulb in order to stain mitral cells and their dendrites. The 3D brain volume is then scanned under line-illumination microscopy for the acquisition of fast calcium imaging data while small temperature drops are induced at the olfactory epithelium. Lastly, the post-acquisition analysis allows the morphological reconstruction of the thermosensitive network comprising the &#947;-glomerulus and its innervating mitral cells, based on specific temperature-induced Ca2+ traces. Using chemical odorants as stimuli in addition to temperature jumps enables the comparison between thermosensitive and chemosensitive networks in the olfactory bulb.

Recording Temperature-induced Neuronal Activity through Monitoring Calcium Changes in the Olfactory Bulb of Xenopus laevis

Here we describe a protocol for measuring and analyzing temperature responses in the olfactory bulb of Xenopus laevis. Olfactory receptor neurons and mitral cells are differentially stained, after which calcium changes are recorded, reflecting a sensitivity of some neural networks in the bulb to temperature drops induced at the nose.

تسجيل درجة الحرارة التي يسببها العصبية آخر من خلال مراقبة التغيرات الكالسيوم في البصلة الشمية من<em&gt; القيطم المورق</em

The olfactory system, specialized in the detection, integration and processing of chemical molecules is likely the most thoroughly studied sensory system. ...

Simultaneous Evaluation of Cerebral Hemodynamics and Light Scattering Properties of the In Vivo Rat Brain Using Multispectral Diffuse Reflectance Imaging

The simultaneous evaluation of cerebral hemodynamics and the light scattering properties of in vivo rat brain tissue is demonstrated using a conventional multispectral diffuse reflectance imaging system. This system is constructed from a broadband white light source, a motorized filter wheel with a set of narrowband interference filters, a light guide, a collecting lens, a video zoom lens, and a monochromatic charged-coupled device (CCD) camera. An ellipsoidal cranial window is made in the skull bone of a rat under isoflurane anesthesia to capture in vivo multispectral diffuse reflectance images of the cortical surface. Regulation of the fraction of inspired oxygen using a gas mixture device enables the induction of different respiratory states such as normoxia, hyperoxia, and anoxia. A Monte Carlo simulation-based multiple regression analysis for the measured multispectral diffuse reflectance images at nine wavelengths (500, 520, 540, 560, 570, 580, 600, 730, and 760 nm) is then performed to visualize the two-dimensional maps of hemodynamics and the light scattering properties of the in vivo rat brain.

Simultaneous Evaluation of Cerebral Hemodynamics and Light Scattering Properties of the In Vivo Rat Brain Using Multispectral Diffuse Reflectance Imaging

The simultaneous evaluation of cerebral hemodynamics and the light scattering properties of in vivo rat brain tissue is demonstrated using a conventional multispectral diffuse reflectance imaging system.

تقييم في وقت واحد من الشلل ديناميكا الدم وتشتت الضوء خصائص<em&gt; في فيفو</em&gt; الجرذ الدماغ عن طريق المتعددة الأطياف منتشر الانعكاس التصوير

The simultaneous evaluation of cerebral hemodynamics and the light scattering properties of in vivo rat brain tissue is demonstrated using a conventional ...

Optical Coherence Tomography: Imaging Mouse Retinal Ganglion Cells In Vivo

Structural changes in the retina are common manifestations of ophthalmic diseases. Optical coherence tomography (OCT) enables their identification in vivo&#8212;rapidly, repetitively, and at a high resolution. This protocol describes OCT imaging in the mouse retina as a powerful tool to study optic neuropathies (OPN). The OCT system is an interferometry-based, non-invasive alternative to common post mortem histological assays. It provides a fast and accurate assessment of retinal thickness, allowing the possibility to track changes, such as retinal thinning or thickening. We present the imaging process and analysis with the example of the Opa1delTTAG mouse line. Three types of scans are proposed, with two quantification methods: standard and homemade calipers. The latter is best for use on the peripapillary retina during radial scans; being more precise, is preferable for analyzing thinner structures. All approaches described here are designed for retinal ganglion cells (RGC) but are easily adaptable to other cell populations. In conclusion, OCT is efficient in mouse model phenotyping and has the potential to be used for the reliable evaluation of therapeutic interventions.

Optical Coherence Tomography: Imaging Mouse Retinal Ganglion Cells In Vivo

This manuscript describes a protocol for the in vivo imaging of the mouse retina with high-resolution spectral domain optical coherence tomography (SD-OCT). It focuses on retinal ganglion cells (RGC) in the peripapillary region, with several scanning and quantifying approaches described.

التصوير المقطعي التماسك البصري: التصوير الماوس العقدة الشبكية خلايا في فيفو

Structural changes in the retina are common manifestations of ophthalmic diseases. Optical coherence tomography (OCT) enables their identification in ...

Quadruple Immunostaining of the Olfactory Bulb for Visualization of Olfactory Sensory Axon Molecular Identity Codes

The mouse olfactory system is often used to study mechanisms of neural circuit formation&#12288;because of its simple anatomical structure. An Olfactory Sensory Neuron (OSN) is a bipolar cell with a single dendrite and a single unbranched axon. An OSN expresses only one Olfactory Receptor (OR) gene, OSNs expressing a given type of OR converge their axons to a few sets of invariant glomeruli in the Olfactory Bulb (OB). A remarkable feature of OSN projection is that the expressed ORs play instructive roles in axonal projection. ORs regulate the expression of multiple axon-sorting molecules and generate the combinatorial molecular code of axon-sorting molecules at the OSN axon termini. Thus, to understand the molecular mechanisms of OR-specific axon guidance mechanisms, it is vital to characterize their expression profiles at the OSN axon termini within the same glomerulus. The aim of this article was to introduce methods for collecting as many glomeruli as possible on a single OB section and for performing immunostaining using multiple antibodies. This would allow the comparison and analysis of the expression patterns of axon-sorting molecules without staining variation between OB sections.

Olfactory sensory neurons express a wide variety of axon-sorting molecules to establish proper neural circuitry. This protocol describes an immunohistochemical staining method to visualize combinatorial expressions of axon-sorting molecules at the axon termini of olfactory sensory neurons.

رباعية المناعية من لمبة الشمية لتصور رموز حسية أكسون الجزيئية الحسية

The mouse olfactory system is often used to study mechanisms of neural circuit formation&#12288;because of its simple anatomical structure. An Olfactory ...

Stereological Estimation of Dopaminergic Neuron Number in the Mouse Substantia Nigra Using the Optical Fractionator and Standard Microscopy Equipment

In pre-clinical Parkinson&#39;s disease research, analysis of the nigrostriatal tract, including quantification of dopaminergic neuron loss within the substantia nigra, is essential. To estimate the total dopaminergic neuron number, unbiased stereology using the optical fractionator method is currently considered the gold standard. Because the theory behind the optical fractionator method is complex and because stereology is difficult to achieve without specialized equipment, several commercially available complete stereology systems that include the necessary software do exist, purely for cell counting reasons. Since purchasing a specialized stereology setup is not always feasible, for many reasons, this report describes a method for the stereological estimation of dopaminergic neuronal cell counts using standard microscopy equipment, including a light microscope, a motorized object table (x, y, z plane) with imaging software, and a computer for analysis. A step-by-step explanation is given on how to perform stereological quantification using the optical fractionator method, and pre-programmed files for the calculation of estimated cell counts are provided. To assess the accuracy of this method, a comparison to data obtained from a commercially available stereology apparatus was performed. Comparable cell numbers were found using this protocol and the stereology device, thus demonstrating the precision of this protocol for unbiased stereology.

This work presents a step-by-step protocol for the unbiased stereological estimation of dopaminergic neuronal cell numbers in the mouse substantia nigra using standard microscopy equipment (i.e., a light microscope, a motorized object table (x, y, z plane), and public domain software for digital image analysis.

ستيريولوجيكال تقدير عدد الخلايا العصبية تتميز في أسود الدماغ الماوس باستخدام فراكتيوناتور الضوئية ومعدات مجهرية القياسية

In pre-clinical Parkinson&#39;s disease research, analysis of the nigrostriatal tract, including quantification of dopaminergic neuron loss within the ...