이 작품은 안토니오와 한나 Damasio에 매더스는 재단과 국립 보건원 (부여 번호 5P50NS019632 - 27)에 의해 부여에 의해 지원되었다.

<ol>
	<li>Haynes, J. -. D., Rees, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Decoding+mental+states+from+brain+activity+in+humans.">Decoding mental states from brain activity in humans.</a> Nat. Rev. Neurosci. 7, 523-534 (2006).</li><li>Norman, K. A., Polyn, S. M., Detre, G. J., Haxby, J. V. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Beyond+mind-reading:+multi-voxel+pattern+analysis+of+fMRI+data.">Beyond mind-reading: multi-voxel pattern analysis of fMRI data.</a> Trends. Cogn. Sci. 10, 424-430 (2006).</li><li>Kriegeskorte, N., Bandettini, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Analyzing+for+information,+not+activation,+to+exploit+high-resolution+fMRI.">Analyzing for information, not activation, to exploit high-resolution fMRI.</a> NeuroImage. 38, 649-662 (2007).</li><li>Mur, M., Bandettini, P. A., Kriegeskorte, N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Revealing+representational+content+with+pattern-information+fMRI+-+an+introductory+guide.">Revealing representational content with pattern-information fMRI - an introductory guide.</a> Soc. Cogn. Affect. Neurosci. 4, 101-109 (2009).</li><li>Kamitani, Y., Tong, F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Decoding+the+visual+and+subjective+contents+of+the+human+brain.">Decoding the visual and subjective contents of the human brain.</a> Nat. Neurosci. 8, 679-685 (2005).</li><li>Formisano, E., De Martino, F., Bonte, M., Goebel, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term="Who"+is+saying+"what"?+Brain-based+decoding+of+human+voice+and+speech?.">"Who" is saying "what"? Brain-based decoding of human voice and speech?.</a> Science. 322, 970-973 (2008).</li><li>Meyer, K., Kaplan, J. T., Essex, R., Webber, C., Damasio, H., Damasio, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Predicting+visual+stimuli+on+the+basis+of+activity+in+auditory+cortices.">Predicting visual stimuli on the basis of activity in auditory cortices.</a> Nat. Neurosci. 13, 667-668 (2010).</li><li>Meyer, K., Kaplan, J. T., Essex, R., Damasio, H., Damasio, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Seeing+touch+is+correlated+with+content-specific+activity+in+primary+somatosensory+cortex.">Seeing touch is correlated with content-specific activity in primary somatosensory cortex.</a> Cereb. Cortex. 21, 2113-2121 (2011).</li><li>Damasio, A. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Time-locked+multiregional+retroactivation:+a+systems-level+proposal+for+the+neural+substrates+of+recall+and+recognition.">Time-locked multiregional retroactivation: a systems-level proposal for the neural substrates of recall and recognition.</a> Cognition. 33, 25-62 (1989).</li><li>Meyer, K., Damasio, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Convergence+and+divergence+in+a+neural+architecture+for+recognition+and+memory.">Convergence and divergence in a neural architecture for recognition and memory.</a> Trends. Neurosci. 32, 376-382 (2009).</li><li>Friston, K. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Modalities,+modes,+and+models+in+functional+neuroimaging.">Modalities, modes, and models in functional neuroimaging.</a> Science. 326, 399-403 (2009).</li><li>Hanke, M., Halchenko, Y. O., Sederberg, P. B., Hanson, S. J., Haxby, J. V., Pollmann, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=PyMVPA:+a+Python+toolbox+for+multivariate+pattern+analysis+of+fMRI+data.">PyMVPA: a Python toolbox for multivariate pattern analysis of fMRI data.</a> Neuroinformatics. 7, 37-53 (2009).</li><li>Harrison, S. A., Tong, F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Decoding+reveals+the+contents+of+visual+working+memory+in+early+visual+areas.">Decoding reveals the contents of visual working memory in early visual areas.</a> Nature. 458, 632-635 (2009).</li><li>Serences, J. T., Ester, E. F., Vogel, E. K., Awh, E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Stimulus-specific+delay+activity+in+human+primary+visual+cortex.">Stimulus-specific delay activity in human primary visual cortex.</a> Psych Sci. 20, 207-214 (2009).</li></ol>

관심 없음 충돌 선언하지 않습니다.

우리의 이전 연구의 결과는 크로스 모달 MVPA가 &quot;마음의 귀&quot;와 &quot;마음의 손길&quot;의 경험이 정신적 이미지의 신경 상호 연구를하는 유용한 도구입니다 보여줍니다. 특히, 그 결과는 이미지의 내용이 융합 - 발산 영역의 Damasio의 프레임 워크에 대한 직접적인 경험 지원을 제공, 각각 초기 청각과 somatosensory cortices의 신경 활동과 상관 것을 보여줍니다. 우리가 설명하는 기본적인 패러다임은 여러 가지 방법으로 확장될 수 있습니다. 대부분 분명, 유사한 연구는 감각 modalities의 다른 조합을 사용하여 수행하실 수 있습니다. 이러한 측면에서, 그것은 이전 실험 특정 크로스 모달 연관 관계를 확립하는 것이 성공의 기회를 증가 수도 있습니다. 예를 들어, 후각 cortices에서 크로스 모달 표현을 공부하기 위해, 하나는 시력 및 식품 항목의 숫자의 냄새 동시에 그들을 노출하여 주요 과목 수 있습니다. Shortly 후, fMRI 스캐너 안에 냄새의 후각 기억은 시각적인 단서에 의해 트리거 될 것이며, MVPA은 전적으로 후각 cortices의 활동에 따라 올바른 식품 항목에 시련을 지정하기 위해 사용될 수 있습니다. 관심의 또 다른 질문은 초기 감각 cortices의 활동의 상호 modally 유발 패턴은 사운드 또는 터치가 실제로 경험이 때 유도되는 패턴과 유사성을 부담하는지 여부이며 즉, 유리 꽃병이 산산조각과 비슷한 패턴을 호출 보는 않습니다 초기 청각 cortices의 신경 활동은 실제로 같은 이벤트를 듣고? 이 질문은, 다시, MVPA을 통해 해결하실 수 있습니다 주제가 제대로 과목 해당 비디오 클립을 시청하면서 기록된 데이터를 차별 할 수 특정한 소리를 듣고있는 동안 기록된 데이터에 대한 훈련이되어 버린 분류입니까? 우리의 청각 연구에서 우리는 그러한 분류를 시도했지만, 결과는 (그림 참조 3. 심판 인치 7 한계되었습니다). 그 연구에서는, 그러나, 참가자의 청각 협회는 어떠한 방법으로 제어되지 않은, 즉, 우리는 비디오와 관련된 각 과목의 분류를 양성하는 데 사용되는 오디오 트랙에 얼마나 유사한 청각 영상 모르겠어요. 다시 말하지만, 그것은 구체적인 크로스 모달 연관이로 visuo - 후각 협회 위에서 설명한, 과목 준비하는 후에 같은 질문을 해결하기 위해 흥미로운 수 있습니다. 이것은 안정적으로 비디오 재판을하는 동안 대상의 정신적 경험을 제어하기 때문에 분류의 예측 성능을 향상시킬 수 있도록합니다. 결론 위해, 우리는 자극이 시간뿐만 아니라 예측할 수 수 보여주는 고전적인 MVPA 패러다임의 확장을 도입뿐만 아니라, 감각 modalities에 걸쳐있다. 따라서, 우리는 MVPA의 사용은 외부 감각 자극에 의​​해 직접적으로 유도 지각 표현의 상호를 조사에 국한되지 것을 보여줍니다. 오히려 MVPA도 있습니다내부 트리거되는 정신적 이미지의 신경 기판을 평가 : Damasio의 융합 - 발산 영역 프레임 워크에 따라, 우리의 연구 결과는 기억에 따라 복원 아르 정신적 이미지의 의식 경험 초기 감각 콘텐츠 특정 신경 진술과 상관 것을 제안 cortices.

cross-modal multivariate pattern analysis

복수 변수 패턴 분석 (MVPA)은 기능적 자기 공명 영상 (fMRI) 데이터 1-4 분석 점점 많이 쓰이는 방법입니다. 일반적으로 메서드는 두뇌의 특정 지역에서 신경 활동 주제의 지각 경험을 식별하는 데 사용됩니다. 예를 들어, 그것은 피사체가 초기 청각 cortices 6 활동 연설의 내용, analogously, 초기 시각 cortices 5 활동이나 인식 시각 통같이의 방향을 예측하는 채용되었습니다. 여기서 우리는 지각 자극은 이내지만, 감각 시스템에서 예측되지 않은 어떤에 따르면, 전통적인 MVPA 패러다임의 확장을 제시. 특히, 우리가 설명하는 방법은 그들이 제시하는 통해서가 아닌 다른 modalities에서 메모리 연결을 보여주고 자극이 그 다른 modalities의 감각 cortices 콘텐츠 관련 활동 패턴을 유도하는지 여부의 문제를 해결합니다.초기의 특정 신경 활동 패턴과 상관있는 &quot;마음의 귀에&quot;이 이미지의 경험, 예를 들어, 바닥에 산산조각 유리 꽃병의 음소거 비디오 클립을보고하면 자동으로 대부분의 관측에 관련된 소리의 청각 이미지가 트리거 청각 cortices? 피사체가 짖는 강아지의 비디오 클립을보고, 대신이라면 또한, 관찰 될 수있는 패턴에서 뚜렷한 활동이 패턴인가? 이전의 두 연구 7,8에서, 우리는 예측할 수 있었던 사운드와 터치 뜻 각각 초기 청각과 somatosensory cortices의 신경 활동을 기반으로 비디오 클립. 우리의 결과는 어떤에 따르면, Damasio 9,10 제안한 neuroarchitectural 프레임 워크 라인에 추억을 기반으로하는 정신 이미지의 경험 - 등 해당 동영상을보고시 &quot;마음의 귀에&quot;에 꽃병을 깜짝 놀라게 할 소리를 듣기로 클립 - 콘텐츠의 다시 공사에서 지원특정 초기 감각 cortices에서 신경 활동 패턴.

Watch this Scientific Journal Video about Cross-Modal Multivariate Pattern Analysis at JoVE.com

Cross-Modal Multivariate Pattern Analysis

고전 복수 변수 패턴 분석 피사체가 해당 cortices (시각 피질의 활동의 예 시각적 자극)의 신경 활동의 인식 감각 자극을 예측합니다. 여기, 우리는 상호 modally 패턴 분석을 적용하고 표시 그 소리와 터치 뜻 시각적 자극이 것은 각각 청각과 somatosensory cortices의 활동에서 예측할 수 있습니다.

크로스 모달 변수 패턴 분석

Multivariate pattern analysis (MVPA) is an increasingly popular method of analyzing functional magnetic resonance imaging (fMRI) data1-4. Typically, the ...

cross-modal-multivariate-pattern-analysis

Research

JoVE Journal

Neuroscience

19.8K 조회수.  University of Southern California. 고전 복수 변수 패턴 분석 피사체가 해당 cortices (시각 피질의 활동의 예 시각적 자극)의 신경 활동의 인식 감각 자극을 예측합니다. 여기, 우리는 상호 modally 패턴 분석을 적용하고 표시 그 소리와 터치 뜻 시각적 자극이 것은 각각 청각과 somatosensory cortices의 활동에서 예측할 수 있습니다.

비디오: Cross-Modal Multivariate Pattern Analysis

Basics of Multivariate Analysis in Neuroimaging Data

Multivariate analysis techniques for neuroimaging data have recently received increasing attention as they have many attractive features that cannot be easily realized by the more commonly used univariate, voxel-wise, techniques1,5,6,7,8,9. Multivariate approaches evaluate correlation/covariance of activation across brain regions, rather than proceeding on a voxel-by-voxel basis. Thus, their results can be more easily interpreted as a signature of neural networks. Univariate approaches, on the other hand, cannot directly address interregional correlation in the brain. Multivariate approaches can also result in greater statistical power when compared with univariate techniques, which are forced to employ very stringent corrections for voxel-wise multiple comparisons. Further, multivariate techniques also lend themselves much better to prospective application of results from the analysis of one dataset to entirely new datasets. Multivariate techniques are thus well placed to provide information about mean differences and correlations with behavior, similarly to univariate approaches, with potentially greater statistical power and better reproducibility checks. In contrast to these advantages is the high barrier of entry to the use of multivariate approaches, preventing more widespread application in the community. To the neuroscientist becoming familiar with multivariate analysis techniques, an initial survey of the field might present a bewildering variety of approaches that, although algorithmically similar, are presented with different emphases, typically by people with mathematics backgrounds. We believe that multivariate analysis techniques have sufficient potential to warrant better dissemination. Researchers should be able to employ them in an informed and accessible manner. The current article is an attempt at a didactic introduction of multivariate techniques for the novice. A conceptual introduction is followed with a very simple application to a diagnostic data set from the Alzheimer s Disease Neuroimaging Initiative (ADNI), clearly demonstrating the superior performance of the multivariate approach.

The current article describes the basics of multivariate analysis and contrasts it to the more commonly used voxel-wise univariate analysis. Both types of analysis are applied to a clinical-neuroscience data set. Supplementary split-half simulations show better replication of the multivariate results in independent data sets.

데이터 Neuroimaging에서 복수 변수 분석의 기초

Multivariate analysis techniques for neuroimaging data have recently received increasing attention as they have many attractive features that cannot be ...

연구

신경과학

Brain Imaging Investigation of the Impairing Effect of Emotion on Cognition

Emotions can impact cognition by exerting both enhancing (e.g., better memory for emotional events) and impairing (e.g., increased emotional distractibility) effects (reviewed in 1). Complementing our recent protocol 2 describing a method that allows investigation of the neural correlates of the memory-enhancing effect of emotion (see also 1, 3-5), here we present a protocol that allows investigation of the neural correlates of the detrimental impact of emotion on cognition. The main feature of this method is that it allows identification of reciprocal modulations between activity in a ventral neural system, involved in 'hot' emotion processing (HotEmo system), and a dorsal system, involved in higher-level 'cold' cognitive/executive processing (ColdEx system), which are linked to cognitive performance and to individual variations in behavior (reviewed in 1). Since its initial introduction 6, this design has proven particularly versatile and influential in the elucidation of various aspects concerning the neural correlates of the detrimental impact of emotional distraction on cognition, with a focus on working memory (WM), and of coping with such distraction 7,11, in both healthy 8-11 and clinical participants 12-14.

We present a protocol that allows investigation of the neural mechanisms mediating the detrimental impact of emotion on cognition, using functional magnetic resonance imaging. This protocol can be used with both healthy and clinical participants.

인식에 대한 감정의 혼란 효과의 두뇌 이미징 조사

Emotions can impact cognition by exerting both enhancing (e.g., better memory for emotional events) and impairing (e.g., increased emotional ...

Transcriptome Analysis of Single Cells

Many gene expression analysis techniques rely on material isolated from heterogeneous populations of cells from tissue homogenates or cells in culture.1,2,3 In the case of the brain, regions such as the hippocampus contain a complex arrangement of different cell types, each with distinct mRNA profiles. The ability to harvest single cells allows for a more in depth investigation into the molecular differences between and within cell populations. We describe a simple and rapid method for harvesting cells for further processing. Pipettes often used in electrophysiology are utilized to isolate (using aspiration) a cell of interest and conveniently deposit it into an Eppendorf tube for further processing with any number of molecular biology techniques. Our protocol can be modified for the harvest of dendrites from cell culture or even individual cells from acute slices.
We also describe the aRNA amplification method as a major downstream application of single cell isolations. This method was developed previously by our lab as an alternative to other gene expression analysis techniques such as reverse-transcription or real-time polymerase chain reaction (PCR).4,5,6,7,8 This technique provides for linear amplification of the polyadenylated RNA beginning with only femtograms of material and resulting in microgram amounts of antisense RNA. The linearly amplified material provides a more accurate estimation than PCR exponential amplification of the relative abundance of components of the transcriptome of the isolated cell. The basic procedure consists of two rounds of amplification. Briefly, a T7 RNA polymerase promoter site is incorporated into double stranded cDNA created from the mRNA transcripts. An overnight in vitro transcription (IVT) reaction is then performed in which T7 RNA polymerase produces many antisense transcripts from the double stranded cDNA. The second round repeats this process but with some technical differences since the starting material is antisense RNA. It is standard to repeat the second round, resulting in three rounds of amplification. Often, the third round in vitro transcription reaction is performed using biotinylated nucleoside triphosphates so that the antisense RNA produced can be hybridized and detected on a microarray.7,8

In this article we describe a simple method for the harvesting of single cells from rat primary neuronal cultures and subsequent transcriptome analysis using aRNA amplification. This approach is generalizable to any cell type.

단일 셀 Transcriptome 분석

Many gene expression analysis techniques rely on material isolated from heterogeneous populations of cells from tissue homogenates or cells in ...

T-maze Forced Alternation and Left-right Discrimination Tasks for Assessing Working and Reference Memory in Mice

Forced alternation and left-right discrimination tasks using the T-maze have been widely used to assess working and reference memory, respectively, in rodents. In our laboratory, we evaluated the two types of memory in more than 30 strains of genetically engineered mice using the automated version of this apparatus. Here, we present the modified T-maze apparatus operated by a computer with a video-tracking system and our protocols in a movie format. The T-maze apparatus consists of runways partitioned off by sliding doors that can automatically open downward, each with a start box, a T-shaped alley, two boxes with automatic pellet dispensers at one side of the box, and two L-shaped alleys. Each L-shaped alley is connected to the start box so that mice can return to the start box, which excludes the effects of experimenter handling on mouse behavior. This apparatus also has an advantage that in vivo microdialysis, in vivo electrophysiology, and optogenetics techniques can be performed during T-maze performance because the doors are designed to go down into the floor. In this movie article, we describe T-maze tasks using the automated apparatus and the T-maze performance of &alpha;-CaMKII+/- mice, which are reported to show working memory deficits in the eight-arm radial maze task. Our data indicated that &alpha;-CaMKII+/- mice showed a working memory deficit, but no impairment of reference memory, and are consistent with previous findings using the eight-arm radial maze task, which supports the validity of our protocol. In addition, our data indicate that mutants tended to exhibit reversal learning deficits, suggesting that &alpha;-CaMKII deficiency causes reduced behavioral flexibility. Thus, the T-maze test using the modified automatic apparatus is useful for assessing working and reference memory and behavioral flexibility in mice.

This article presents the protocol of T-maze tests using a modified automated apparatus for assessing the learning and memory functions in mice.

T-미로 강제 교대 및 근무와 마우스의 레퍼런스 메모리를 평가하기위한 왼쪽 오른쪽 차별 작업

Forced alternation and left-right discrimination tasks using the T-maze have been widely used to assess working and reference memory, respectively, in ...

Creating Objects and Object Categories for Studying Perception and Perceptual Learning

In order to quantitatively study object perception, be it perception by biological systems or by machines, one needs to create objects and object categories with precisely definable, preferably naturalistic, properties1. Furthermore, for studies on perceptual learning, it is useful to create novel objects and object categories (or object classes) with such properties2.
Many innovative and useful methods currently exist for creating novel objects and object categories3-6 (also see refs. 7,8). However, generally speaking, the existing methods have three broad types of shortcomings.
First, shape variations are generally imposed by the experimenter5,9,10, and may therefore be different from the variability in natural categories, and optimized for a particular recognition algorithm. It would be desirable to have the variations arise independently of the externally imposed constraints.
Second, the existing methods have difficulty capturing the shape complexity of natural objects11-13. If the goal is to study natural object perception, it is desirable for objects and object categories to be naturalistic, so as to avoid possible confounds and special cases.
Third, it is generally hard to quantitatively measure the available information in the stimuli created by conventional methods. It would be desirable to create objects and object categories where the available information can be precisely measured and, where necessary, systematically manipulated (or 'tuned'). This allows one to formulate the underlying object recognition tasks in quantitative terms.
Here we describe a set of algorithms, or methods, that meet all three of the above criteria. Virtual morphogenesis (VM) creates novel, naturalistic virtual 3-D objects called 'digital embryos' by simulating the biological process of embryogenesis14. Virtual phylogenesis (VP) creates novel, naturalistic object categories by simulating the evolutionary process of natural selection9,12,13. Objects and object categories created by these simulations can be further manipulated by various morphing methods to generate systematic variations of shape characteristics15,16. The VP and morphing methods can also be applied, in principle, to novel virtual objects other than digital embryos, or to virtual versions of real-world objects9,13. Virtual objects created in this fashion can be rendered as visual images using a conventional graphical toolkit, with desired manipulations of surface texture, illumination, size, viewpoint and background. The virtual objects can also be 'printed' as haptic objects using a conventional 3-D prototyper.
We also describe some implementations of these computational algorithms to help illustrate the potential utility of the algorithms. It is important to distinguish the algorithms from their implementations. The implementations are demonstrations offered solely as a 'proof of principle' of the underlying algorithms. It is important to note that, in general, an implementation of a computational algorithm often has limitations that the algorithm itself does not have.
Together, these methods represent a set of powerful and flexible tools for studying object recognition and perceptual learning by biological and computational systems alike. With appropriate extensions, these methods may also prove useful in the study of morphogenesis and phylogenesis.

We describe a novel methodology for creating naturalistic 3-D objects and object categories with precisely defined feature variations. We use simulations of the biological processes of morphogenesis and phylogenesis to create novel, naturalistic virtual 3-D objects and object categories that can then be rendered as visual images or haptic objects.

지각과 지각 학습 유학을위한 객체와 객체 카테고리 만들기

In order to quantitatively study object perception, be it perception by biological systems or by machines, one needs to create objects and object ...

Visualization and Genetic Manipulation of Dendrites and Spines in the Mouse Cerebral Cortex and Hippocampus using In utero Electroporation

In utero electroporation (IUE) has become a powerful technique to study the development of different regions of the embryonic nervous system 1-5. To date this tool has been widely used to study the regulation of cellular proliferation, differentiation and neuronal migration especially in the developing cerebral cortex 6-8. Here we detail our protocol to electroporate in utero the cerebral cortex and the hippocampus and provide evidence that this approach can be used to study dendrites and spines in these two cerebral regions.
Visualization and manipulation of neurons in primary cultures have contributed to a better understanding of the processes involved in dendrite, spine and synapse development. However neurons growing in vitro are not exposed to all the physiological cues that can affect dendrite and/or spine formation and maintenance during normal development. Our knowledge of dendrite and spine structures in vivo in wild-type or mutant mice comes mostly from observations using the Golgi-Cox method 9. However, Golgi staining is considered to be unpredictable. Indeed, groups of nerve cells and fiber tracts are labeled randomly, with particular areas often appearing completely stained while adjacent areas are devoid of staining. Recent studies have shown that IUE of fluorescent constructs represents an attractive alternative method to study dendrites, spines as well as synapses in mutant / wild-type mice 10-11 (Figure 1A). Moreover in comparison to the generation of mouse knockouts, IUE represents a rapid approach to perform gain and loss of function studies in specific population of cells during a specific time window. In addition, IUE has been successfully used with inducible gene expression or inducible RNAi approaches to refine the temporal control over the expression of a gene or shRNA 12. These advantages of IUE have thus opened new dimensions to study the effect of gene expression/suppression on dendrites and spines not only in specific cerebral structures (Figure 1B) but also at a specific time point of development (Figure 1C).
Finally, IUE provides a useful tool to identify functional interactions between genes involved in dendrite, spine and/or synapse development. Indeed, in contrast to other gene transfer methods such as virus, it is straightforward to combine multiple RNAi or transgenes in the same population of cells. 
In summary, IUE is a powerful method that has already contributed to the characterization of molecular mechanisms underlying brain function and disease and it should also be useful in the study of dendrites and spines.

Visualization and Genetic Manipulation of Dendrites and Spines in the Mouse Cerebral Cortex and Hippocampus using In utero Electroporation

This article describes in detail a protocol to electroporate in utero the cerebral cortex and the hippocampus at E14.5 in mice. We also show that this is a valuable method to study dendrites and spines in these two cerebral regions.

사용하는 마우스 대뇌 피질과 해마의 시각화 및 Dendrites와 쪽이의 유전자 조작<em&gt; utero에서</em&gt; Electroporation

In utero electroporation (IUE) has become a powerful technique to study the development of different regions of the embryonic nervous system 1-5. To date ...

The Swimmeret System of Crayfish: A Practical Guide for the Dissection of the Nerve Cord and Extracellular Recordings of the Motor Pattern

Here we demonstrate the dissection of the crayfish abdominal nerve cord. The preparation comprises the last two thoracic ganglia (T4, T5) and the chain of abdominal ganglia (A1 to A6). This chain of ganglia includes the part of the central nervous system (CNS) that drives coordinated locomotion of the pleopods (swimmerets): the swimmeret system. It is known for over five decades that in crayfish each swimmeret is driven by its own independent pattern generating kernel that generates rhythmic alternating activity 1-3. The motor neurons innervating the musculature of each swimmeret comprise two anatomically and functionally distinct populations 4. One is responsible for the retraction (power stroke, PS) of the swimmeret. The other drives the protraction (return stroke, RS) of the swimmeret. Motor neurons of the swimmeret system are able to produce spontaneously a fictive motor pattern, which is identical to the pattern recorded in vivo 1.
The aim of this report is to introduce an interesting and convenient model system for studying rhythm generating networks and coordination of independent microcircuits for students&#8217; practical laboratory courses. The protocol provided includes step-by-step instructions for the dissection of the crayfish&#8217;s abdominal nerve cord, pinning of the isolated chain of ganglia, desheathing the ganglia and recording the swimmerets fictive motor pattern extracellularly from the isolated nervous system.
Additionally, we can monitor the activity of swimmeret neurons recorded intracellularly from dendrites. Here we also describe briefly these techniques and provide some examples. Furthermore, the morphology of swimmeret neurons can be assessed using various staining techniques. Here we provide examples of intracellular (by iontophoresis) dye filled neurons and backfills of pools of swimmeret motor neurons. In our lab we use this preparation to study basic functions of fictive locomotion, the effect of sensory feedback on the activity of the CNS, and coordination between microcircuits on a cellular level.

Here we describe the dissection of the crayfish abdominal nerve cord. We also demonstrate an electrophysiological technique to record fictive locomotion from swimmeret motor neurons.

왕새우의 Swimmeret 시스템 : 신경 코드 및 모터 패턴의 세포 외 녹음의 해부를위한 실무 가이드

Here we demonstrate the dissection of the crayfish abdominal nerve cord. The preparation comprises the last two thoracic ganglia (T4, T5) and the chain of ...

Peering at Brain Polysomes with Atomic Force Microscopy

The translational machinery, i.e., the polysome or polyribosome, is one of the biggest and most complex cytoplasmic machineries in cells. Polysomes, formed by ribosomes, mRNAs, several proteins and non-coding RNAs, represent integrated platforms where translational controls take place. However, while the ribosome has been widely studied, the organization of polysomes is still lacking comprehensive understanding. Thus much effort is required in order to elucidate polysome organization and any novel mechanism of translational control that may be embedded. Atomic force microscopy (AFM) is a type of scanning probe microscopy that allows the acquisition of 3D images at nanoscale resolution. Compared to electron microscopy (EM) techniques, one of the main advantages of AFM is that it can acquire thousands of images both in air and in solution, enabling the sample to be maintained under near physiological conditions without any need for staining and fixing procedures. Here, a detailed protocol for the accurate purification of polysomes from mouse brain and their deposition on mica substrates is described. This protocol enables polysome imaging in air and liquid with AFM and their reconstruction as three-dimensional objects. Complementary to cryo-electron microscopy (cryo-EM), the proposed method can be conveniently used for systematically analyzing polysomes and studying their organization.

While ribosome structure has been extensively characterized, the organization of polysomes is still understudied. To overcome this lack of knowledge, we present here a detailed preparation protocol for accurate imaging of mammalian polysomes by atomic force microscopy (AFM) in air and liquid.

원자 힘 현미경과 뇌 Polysom​​es에서 피어링

The translational machinery, i.e., the polysome or polyribosome, is one of the biggest and most complex cytoplasmic machineries in cells. Polysomes, ...

Analyzing Synaptic Modulation of Drosophila melanogaster Photoreceptors after Exposure to Prolonged Light

The nervous system has the remarkable ability to adapt and respond to various stimuli. This neural adjustment is largely achieved through plasticity at the synaptic level. The Active Zone (AZ) is the region at the presynaptic membrane that mediates neurotransmitter release and is composed of a dense collection of scaffold proteins. AZs of Drosophila melanogaster (Drosophila) photoreceptors undergo molecular remodeling after prolonged exposure to natural ambient light. Thus the level of neuronal activity can rearrange the molecular composition of the AZ and contribute to the regulation of the functional output.
Starting from the light exposure set-up preparation to the immunohistochemistry, this protocol details how to quantify the number, the spatial distribution, and the delocalization level of synaptic molecules at AZs in Drosophila photoreceptors. Using image analysis software, clusters of the GFP-fused AZ component Bruchpilot were identified for each R8 photoreceptor (R8) axon terminal. Detected Bruchpilot spots were automatically assigned to individual R8 axons. To calculate the distribution of spot frequency along the axon, we implemented a customized software plugin. Each axon&#39;s start-point and end-point were manually defined and the position of each Bruchpilot spot was projected onto the connecting line between start and end-point. Besides the number of Bruchpilot clusters, we also quantified the delocalization level of Bruchpilot-GFP within the clusters. These measurements reflect in detail the spatially resolved synaptic dynamics in a single neuron under different environmental conditions to stimuli.

Analyzing Synaptic Modulation of Drosophila melanogaster Photoreceptors after Exposure to Prolonged Light

Here we show how to quantify the number and spatial distribution of synaptic active zones in Drosophila melanogaster&#160;photoreceptors, highlighted with a genetically encoded molecular marker, and their modulation after prolonged exposure to light.

의 시냅틱 변조 분석<em&gt; 노랑 초파리</em&gt; 장시간 빛에 노출 후 광수

The nervous system has the remarkable ability to adapt and respond to various stimuli. This neural adjustment is largely achieved through plasticity at ...

Visualization of the Axonal Projection Pattern of Embryonic Motor Neurons in Drosophila

The establishment of functional neuromuscular circuits relies on precise connections between developing motor axons and target muscles. Motor neurons extend growth cones to navigate along specific pathways by responding to a large number of axon guidance cues that emanate from the surrounding extracellular environment. Growth cone target recognition also plays a critical role in neuromuscular specificity. This work presents a standard immunohistochemistry protocol to visualize motor neuron projections of late stage-16 Drosophila melanogaster embryos. This protocol includes a few key steps, including a genotyping procedure, to sort the desired mutant embryos; an immunostaining procedure, to tag embryos with fasciclin II (FasII) antibody; and a dissection procedure, to generate filleted preparations from fixed embryos. Motor axon projections and muscle patterns in the periphery are much better visualized in flat preparations of filleted embryos than in whole-mount embryos. Therefore, the filleted preparation of fixed embryos stained with FasII antibody provides a powerful tool to characterize the genes required for motor axon pathfinding and target recognition, and it can also be applied to both loss-of-function and gain-of-function genetic screens.

Visualization of the Axonal Projection Pattern of Embryonic Motor Neurons in Drosophila

This work details a standard immunohistochemistry method to visualize motor neuron projections of late stage-16 Drosophila melanogaster embryos. The filleted preparation of fixed embryos stained with FasII antibody provides a powerful tool to characterize the genes required for motor axon pathfinding and target recognition during neural development.

배아 운동 뉴런의 Axonal 투영 패턴의 시각화<em&gt; Drosophila</em

The establishment of functional neuromuscular circuits relies on precise connections between developing motor axons and target muscles. Motor neurons ...