This work was supported by grants from the Natural Sciences and Engineering Research Council (NSERC) of Canada and The Canada Foundation for Innovation. Thanks to Dr Paul Mains (University of Calgary, Calgary, Canada) for let-502(sb118ts) strain. Some of the strains were provided by the Caenorhabditis Genetics Center, which is funded by NIH Office of Research Infrastructure Programs (P40 OD010440).

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The authors declare that they have no competing financial interests.

이 프로토콜은 초기 특성화하는 새로운 메트릭 및 배아 신장의 후반 단계에 대해 설명합니다. 섹션 1에서 중요한 단계는 패드 박테리아의 잠재적 인 존재이다. 배아는 밀봉 패드와 이미지 수집시 커버 슬립 사이에 동봉되어 있습니다. 슬라이드를 씰링하는 두 시간 이상 지속 취득 중에 동물의 탈수를 방지하기 위해 필요하다. 우리의 지식, 슬라이드와 커버 슬립 사이의 아가?...

짧은 라이프 사이클 : 거의 50 년 동안 선충 예쁜 꼬마 선충 개발, 신경 생물학, 진화, 호스트 병원체 상호 작용 등의 중요한 문제를 연구하는 강력한 모델로 자리 매김 한 개발의 연구에서이 모델의 강도에 달려있다 삼일의; 이러한 동물은 유전자 조작 될 수있는 용이성; 대부분 여분의 자궁이다 살아있는 동물과 개발에 세포 변위 및 형태의 관찰을 가능하게 투명성. 선충의 발달 단계는 성인이 다음에 배아 네 애벌레 단계 (L4에 L1)를 포함한다. 배아 발생 동안, 표피 형태 형성 세포는 그들의 접합부를 재구성하는 방법을 그룹으로 마이그레이션 방법 상피의 더 나은 이해를 가능하게하고 개별적인 형태뿐만 아니라 기능적 상피 세포 내에서의 상대 위치를 수정하는 능력에 상당한 관심을 끌었다.표피의 형태 형성은 네 단계로 구분됩니다 지느러미 표피 세포의 구조 조정에 구성된 지느러미 인터에서, 하피라고 함; 따라서 상피 세포 단층의 배아를 감싸는 복부 정중선으로 복부 피하 세포의 이동에 구성된 복부 인클로저; 초기와 후기 신장 벌레 모양의 유충에 콩 모양의 배아를 변환. 다음과 같은 형태 형성, 배아 해치 및 L1 유충은 즉시 환경에서 사용 가능한 박테리아를 사용하여 먹이를 시작합니다. 배아 신장 따라서 배아 발달의 후기이다. 그것은 그것의 길이 방향 축을 따라 배아의 확장과 가로 직경의 감소로 구성되어 있습니다. 이것은 피하 세포의 형상 급격한 변형을 포함한다. 신장은 초기와 후기로 나누어진다. 몸 벽 근육이 w의 1.75 배 단계에서 계약 시작할 때 초기 단계는 쉼표 단계에서 시작 및 종료ILD 형 (중량) 배아 - 비 신장 배아에 비해 길이의 1.75 배입니다 배아에 대응. 그 단계에서 발생하는 형태 형성 공정은 주로 두 배의 안테 - 후방 축에 그들의 연장 운전 피하 세포의 꼭대기에 위치한 극 액틴 필라멘트 다발 FBS ()의 수축에 의해 구동된다. 이물 제거의 수축 세 키나제 LET-502 / ROCK, MRCK-1, PAK-1 (5)에 의해 미오신 라이트 체인의 인산화에 의해 컨트롤입니다. 몸 벽 근육의 기능이되어 계약을 시작할 때 신장의 후기가 시작됩니다. 그것은 지느러미와 복부 피하 세포에 몸 벽 근육에서 mechanotransduction 신호를 포함하고 동물 3 부화 할 때 끝납니다. 및 L1 유충 (애벌레 체포 표현형으로 그 체포 그들의 개발, 신장 결함은 일반적으로 배아로 죽어가는 동물의 비율 (EMB 배아 치사)을 특징으로 레벨a) 및 중량보다 훨씬 짧은 것을. 발달 체포의 단계의 확인은 죽은 태아의 현미경 관찰을 필요로하고 유충 3-6을 체포했다. 그것은 최근에 같은 Cdc42 / 라 세미 레귤레이터 및 이펙터 PIX-1과 박-1, 여러 유전자가, 초기와 후기 신장 3,7 둘 중에 형태 형성 과정을 제어하는 것으로 나타났다. 우리는 또한 최근에 형태 형성 과정 초기 신장 3 ~ 7시 배아의 안테 - 후방 축을 따라 차이가 있음을 보여 주었다. 이러한 연구 결과는 특히 초기 신장 동안 자신의 안테 - 후방 축을 따라 배아의 형태의 특성을 가능하게 일찍 또는 늦게 신장 단계 및 기타 측정을 대상으로 새로운 지표의 개발 동기를 부여. 이러한 새로운 방법은 폭뿐만 아니라 처음부터 초기 신장 끝에 배아의 길이를 측정하는 구성 그광고 및 꼬리. 7 두 프로토콜은 또한 L1 단계 7에서 동기화 새로 부화 유충의 길이를 측정하기 위해 개발되었다. 유충, 성인 및 배양액에 존재하는 세균이 처리에 의해 용해되는 동안 배아 달걀 껍질 알칼리 처리 염소산으로부터 보호. 이러한 처리 후, 잘 공급 성인 (8)의 대부분을 함유하는 비 - 동기화 집단으로부터 배아를 정화하는데 사용된다. 식품 제한은 새로 부화 유충을 동기화하는 데 사용됩니다. 이들 유충의 길이를 측정하는 것은 다음 신장 결함을 검출하기 위해 사용된다. 에서 부화 유충이 아닌 완전히 신장 배아가 공급 될 때 &quot;정상 길이&quot;로 복구 할 수 있지만, 음식이없는 구속 할 때 자신의 소형화를 유지하기 때문에이 측정은 배양 플레이트에 구속 유충의 측정보다 선호된다. 여기서는 르의 측정을 가능 상세한 프로토콜을 제시경과 DIC 현미경과 화상 해석 (1 프로토콜)을 사용하여 배아뿐만 아니라 헤드의 폭과 신장의 꼬리 ngth. 또한 화상 해석 (프로토콜 2) 플로우 세포 계측법 (3 프로토콜)을 사용하여 동기화 된 유충의 길이를 측정하기위한 구체적인 프로토콜을 제공한다.

<table border="0" cellpadding="0" cellspacing="0" width="696">
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			<td height="21" style="height:21px;width:216px;">Agar&#160;</td>
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			<td style="width:195px;">Bio Basic Inc.</td>
			<td style="width:119px;">DB0483</td>
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</table>

novel metrics to characterize embryonic elongation of the nematode caenorhabditis elegans

Dissecting the signaling pathways that control the alteration of morphogenic processes during embryonic development requires robust and sensitive metrics. Embryonic elongation of the nematode Caenorhabditis elegans is a late developmental stage consisting of the elongation of the embryo along its longitudinal axis. This developmental stage is controlled by intercellular communication between hypodermal cells and underlying body-wall muscles. These signaling mechanisms control the morphology of hypodermal cells by remodeling the cytoskeleton and the cell-cell junctions. Measurement of embryonic lethality and developmental arrest at larval stages as well as alteration of cytoskeleton and cell-cell adhesion structures in hypodermal and muscle cells are classical phenotypes that have been used for more than 25 years to dissect these signaling pathways. Recent studies required the development of novel metrics specifically targeting either early or late elongation and characterizing morphogenic defects along the antero-posterior axis of the embryo. Here, we provide detailed protocols enabling the accurate measurement of the length and the width of the elongating embryos as well as the length of synchronized larvae. These methods constitute useful tools to identify genes controlling elongation, to assess whether these genes control both early and late phases of this stage and are required evenly along the antero-posterior axis of the embryo.

헤드폰, Tail- 및 헤드 / 테일 폭 비율은 강력한 지표입니다. 여기에 설명 된 프로토콜이 성공적으로 조절기와의 Rho GTP 아제 PIX-1의 이펙터 기능, 박-1 특성화 렛 502 초 신장 7 동안 사용되어왔다. ...

Watch this Scientific Journal Video about Novel Metrics to Characterize Embryonic Elongation of the Nematode Caenorhabditis elegans at JoVE.com

Novel Metrics to Characterize Embryonic Elongation of the Nematode Caenorhabditis elegans

소설 메트릭은 선충의 배아 신장의 특성을<em&gt; 예쁜 꼬마 선충</em

Here we detail protocols specifically designed to monitor morphogenic defects that occur during early and late phases of embryonic elongation of the nematode Caenorhabditis elegans. Ultimately, these protocols are designed to identify genes that regulate these phases and to characterize their differential requirements along the antero-posterior axis of the embryo.

Dissecting the signaling pathways that control the alteration of morphogenic processes during embryonic development requires robust and sensitive metrics. ...

novel-metrics-to-characterize-embryonic-elongation-nematode

Research

JoVE Journal

Neuroscience

Novel Metrics to Characterize Embryonic Elongation of the Nematode Caenorhabditis elegans

6.0K 조회수.  Université du Québec à Montréal. Here we detail protocols specifically designed to monitor morphogenic defects that occur during early and late phases of embryonic elongation of the nematode Caenorhabditis elegans. Ultimately, these protocols are designed to identify genes that regulate these phases and to characterize their differential requirements along the antero-posterior axis of the embryo. 

비디오: Novel Metrics to Characterize Embryonic Elongation of the Nematode Caenorhabditis elegans

Multi-unit Recording Methods to Characterize Neural Activity in the Locust (Schistocerca Americana) Olfactory Circuits

Detection and interpretation of olfactory cues are critical for the survival of many organisms. Remarkably, species across phyla have strikingly similar olfactory systems suggesting that the biological approach to chemical sensing has been optimized over evolutionary time1. In the insect olfactory system, odorants are transduced by olfactory receptor neurons (ORN) in the antenna, which convert chemical stimuli into trains of action potentials. Sensory input from the ORNs is then relayed to the antennal lobe (AL; a structure analogous to the vertebrate olfactory bulb). In the AL, neural representations for odors take the form of spatiotemporal firing patterns distributed across ensembles of principal neurons (PNs; also referred to as projection neurons)2,3. The AL output is subsequently processed by Kenyon cells (KCs) in the downstream mushroom body (MB), a structure associated with olfactory memory and learning4,5. Here, we present electrophysiological recording techniques to monitor odor-evoked neural responses in these olfactory circuits.
First, we present a single sensillum recording method to study odor-evoked responses at the level of populations of ORNs6,7. We discuss the use of saline filled sharpened glass pipettes as electrodes to extracellularly monitor ORN responses. Next, we present a method to extracellularly monitor PN responses using a commercial 16-channel electrode3. A similar approach using a custom-made 8-channel twisted wire tetrode is demonstrated for Kenyon cell recordings8. We provide details of our experimental setup and present representative recording traces for each of these techniques.

Multi-unit Recording Methods to Characterize Neural Activity in the Locust Schistocerca Americana Olfactory Circuits

We demonstrate variations of the extracellular multi-unit recording technique to characterize odor-evoked responses in the first three stages of the invertebrate olfactory pathway. These techniques can easily be adapted to examine ensemble activity in other neural systems as well.

메뚜기에 신경 활동을 특성화하기 위해 다중 단위 기록 방법 (<em&gt; Schistocerca 아메리카나</em&gt;) 강한 회로

Detection and interpretation of olfactory cues are critical for the survival  of many organisms. Remarkably, species across phyla have strikingly similar ...

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A Caenorhabditis elegans Model System for Amylopathy Study

Amylopathy is a term that describes abnormal synthesis and accumulation of amyloid beta (A&beta;) in tissues with time. A&beta; is a hallmark of Alzheimer's disease (AD) and is found in Lewy body dementia, inclusion body myositis and cerebral amyloid angiopathy 1-4. Amylopathies progressively develop with time. For this reason simple organisms with short lifespans may help to elucidate molecular aspects of these conditions. Here, we describe experimental protocols to study A&beta;-mediated neurodegeneration using the worm Caenorhabditis elegans. Thus, we construct transgenic worms by injecting DNA encoding human A&beta;42 into the syncytial gonads of adult hermaphrodites. Transformant lines are stabilized by a mutagenesis-induced integration. Nematodes are age synchronized by collecting and seeding their eggs. The function of neurons expressing A&beta;42 is tested in opportune behavioral assays (chemotaxis assays). Primary neuronal cultures obtained from embryos are used to complement behavioral data and to test the neuroprotective effects of anti-apoptotic compounds.

A Caenorhabditis elegans Model System for Amylopathy Study

We describe methods to study aspects of amylopathies in the worm C. elegans. We show how to construct worms expressing human A&beta;42 in neurons and how to test their function in behavioral assays. We further show how to obtain primary neuronal cultures that can be used for pharmacological testing.

<em&gt; Caenorhabditis의 엘레</emAmylopathy 연구에 대한&gt; 모델 시스템

Amylopathy is a term that describes abnormal synthesis and accumulation of amyloid beta (A&beta;) in tissues with time. A&beta; is a hallmark of ...

Methods for Studying the Mechanisms of Action of Antipsychotic Drugs in Caenorhabditis elegans

Caenorhabditis elegans is a simple genetic organism amenable to large-scale forward and reverse genetic screens and chemical genetic screens. The C. elegans genome includes potential antipsychotic drug (APD) targets conserved in humans, including genes encoding proteins required for neurotransmitter synthesis and for synaptic structure and function. APD exposure produces developmental delay and/or lethality in nematodes in a concentration-dependent manner. These phenotypes are caused, in part, by APD-induced inhibition of pharyngeal pumping1,2. Thus, the developmental phenotype has a neuromuscular basis, making it useful for pharmacogenetic studies of neuroleptics. Here we demonstrate detailed procedures for testing APD effects on nematode development and pharyngeal pumping. For the developmental assay, synchronized embryos are placed on nematode growth medium (NGM) plates containing APDs, and the stages of developing animals are then scored daily. For the pharyngeal pumping rate assay, staged young adult animals are tested on NGM plates containing APDs. The number of pharyngeal pumps per unit time is recorded, and the pumping rate is calculated. These assays can be used for studying many other types of small molecules or even large molecules.

Methods for Studying the Mechanisms of Action of Antipsychotic Drugs in Caenorhabditis elegans

Approaches for testing the effects of antipsychotic drugs (APDs) in Caenorhabditis elegans are demonstrated. Assays are described for testing drug effects on development and viability and on pharyngeal pumping rate. These methods are also applicable for pharmacogenetic experiments with drug classes other than APDs.

에서 항 정신병 약물의 작용 메커니즘을 공부하는 방법<em&gt; 예쁜 꼬마 선충</em

Caenorhabditis elegans is a simple genetic organism amenable to  large-scale forward and reverse genetic screens and chemical genetic screens. The C.  ...

An Assay for Measuring the Effects of Ethanol on the Locomotion Speed of Caenorhabditis elegans

Alcohol use disorders are a significant public health concern, for which there are few effective treatment strategies. One difficulty that has delayed the development of more effective treatments is the relative lack of understanding of the molecular underpinnings of the effects of ethanol on behavior. The nematode, Caenorhabditis elegans (C.&#160;elegans), provides a useful model in which to generate and test hypotheses about the molecular effects of ethanol. Here, we describe an assay that has been developed and used to examine the roles of particular genes and environmental factors in behavioral responses to ethanol, in which locomotion is the behavioral output. Ethanol dose-dependently causes an acute depression of crawling on an agar surface. The effects are dynamic; animals exposed to a high concentration demonstrate an initial strong depression of crawling, referred to here as initial sensitivity, and then partially recover locomotion speed despite the continued presence of the drug. This ethanol-induced behavioral plasticity is referred to here as the development of acute functional tolerance. This assay has been used to demonstrate that these two phenotypes are distinct and genetically separable. The straightforward locomotion assay described here is suitable for examining the effects of both genetic and environmental manipulations on these acute behavioral responses to ethanol in C.&#160;elegans.

An Assay for Measuring the Effects of Ethanol on the Locomotion Speed of Caenorhabditis elegans

C. elegans is a useful model for studying the effects of ethanol on behavior. We present a behavioral assay that quantifies the effects of ethanol on the locomotion speed of crawling worms; both initial sensitivity and the development of acute functional tolerance to ethanol can be measured with this assay.

의 로코 속도에 에탄올의 효과를 측정하는 분석<em&gt; 예쁜 꼬마 선충</em

Alcohol use disorders are a significant public health concern, for which there are few effective treatment strategies. One difficulty that has delayed the ...

Agarose Microchambers for Long-term Calcium Imaging of Caenorhabditis elegans

Behavior is controlled by the nervous system. Calcium imaging is a straightforward method in the transparent nematode Caenorhabditis elegans to measure the activity of neurons during various behaviors. To correlate neural activity with behavior, the animal should not be immobilized but should be able to move. Many behavioral changes occur during long time scales and require recording over many hours of behavior. This also makes it necessary to culture the worms in the presence of food. How can worms be cultured and their neural activity imaged over long time scales? Agarose Microchamber Imaging (AMI) was previously developed to culture and observe small larvae and has now been adapted to study all life stages from early L1 until the adult stage of C. elegans. AMI can be performed on various life stages of C. elegans. Long-term calcium imaging is achieved without immobilizing the animals by using short externally triggered exposures combined with an electron multiplying charge-coupled device (EMCCD) camera recording. Zooming out or scanning can scale up this method to image up to 40 worms in parallel. Thus, a method is described to image behavior and neural activity over long time scales in all life stages of C. elegans.

Agarose Microchambers for Long-term Calcium Imaging of Caenorhabditis elegans

Imaging behavior and neural activity over long time scales without immobilization of the animal is a prerequisite to understand behavior. Agarose microfluidic chambers imaging (AMI) can be used to image neural activity and behavior for all life stages of Caenorhabditis elegans.

의 장기 칼슘 이미징을위한 아가로 오스 Microchambers<em&gt; 예쁜 꼬마 선충</em

Behavior is controlled by the nervous system. Calcium imaging is a straightforward method in the transparent nematode Caenorhabditis elegans to measure ...

Dynamic Quantitative Sensory Testing to Characterize Central Pain Processing

Central facilitation and modulation of incoming nociceptive signals play an important role in the perception of pain. Disruption in central pain processing is present in many chronic pain conditions and can influence responses to specific therapies. Thus, the ability to precisely describe the state of central pain processing has profound clinical significance in both prognosis and prediction. Because it is not practical to record neuronal firings directly in the human spinal cord, surrogate behavior tests become an important tool to assess the state of central pain processing. Dynamic QST is one such test, and can probe both the ascending facilitation and descending modulation of incoming nociceptive signals via TS and CPM, respectively. Due to the large between-individual variability in the sensitivity to noxious signals, standardized TS and CPM tests may not yield any meaningful data in up to 50% of the population due to floor or ceiling effects. We present methodologies to individualize TS and CPM so we can capture these measures in a broader range of individuals than previously possible. We have used these methods successfully in several studies at the lab, and data from one ongoing study will be presented to demonstrate feasibility and potential applications of the methods.

By assessing pain in response to repetitive or different types of standardized stimuli, dynamic quantitative sensory testing (QST) can reveal changes in the central processing of pain. We present methods to optimize and individualize two dynamic QST measures: temporal summation (TS) and conditioned pain modulation (CPM).

동적 정량적 감각 테스트는 중앙 통증 처리의 특성을

Central facilitation and modulation of incoming nociceptive signals play an important role in the perception of pain. Disruption in central pain ...

A Caenorhabditis elegans Nutritional-status Based Copper Aversion Assay

To ensure survival, organisms must be capable of avoiding unfavorable habitats while ensuring a consistent food source. Caenorhabditis elegans alter their locomotory patterns upon detection of diverse environmental stimuli and can modulate their suite of behavioral responses in response to starvation conditions. Nematodes typically exhibit a decreased aversive response when removed from a food source for over 30 min. Observation of behavioral changes in response to a changing nutritional status can provide insight into the mechanisms that regulate the transition from a well-fed to starved state.
We have developed an assay that measures a nematode&#39;s ability to cross an aversive barrier (i.e. copper) then reach a food source over a prolonged period of time. This protocol builds upon previous work by integrating multiple variables in a manner that allows for continued data collection as the organisms shift towards an increasingly starved condition. Moreover, this assay permits an increased sample size so that larger populations of nematodes can be simultaneously evaluated.
Organisms defective for the ability to detect or respond to copper immediately cross the chemical barrier, while wild type nematodes are initially repelled. As wild type worms are increasingly starved, they begin to cross the barrier and reach the food source. We designed this assay to evaluate a mutant that is incapable of responding to diverse environmental cues, including food sensation or detection of aversive chemicals. When evaluated via this protocol, the defective organisms immediately crossed the barrier, but were also incapable of detecting a food source. Hence, these mutants repeatedly cross the chemical barrier despite temporarily reaching a food source. This assay can straightforwardly test populations of worms to evaluate potential pathway defects related to aversion and starvation.

A Caenorhabditis elegans Nutritional-status Based Copper Aversion Assay

Here, we present a Caenorhabditis elegans-specific assay designed to evaluate changes in copper aversion behavior and the ability to locate a common food source, as the organism progresses from a well-fed to starved nutritional state.

에이<em&gt; Caenorhabditis elegans</em&gt; 영양 상태 기반 구리 혐오 검정

To ensure survival, organisms must be capable of avoiding unfavorable habitats while ensuring a consistent food source. Caenorhabditis elegans alter their ...

Ratiometric Calcium Imaging of Individual Neurons in Behaving Caenorhabditis Elegans

It has become increasingly clear that neural circuit activity in behaving animals differs substantially from that seen in anesthetized or immobilized animals. Highly sensitive, genetically encoded fluorescent reporters of Ca2+ have revolutionized the recording of cell and synaptic activity using non-invasive optical approaches in behaving animals. When combined with genetic and optogenetic techniques, the molecular mechanisms that modulate cell and circuit activity during different behavior states can be identified.
Here we describe methods for ratiometric Ca2+ imaging of single neurons in freely behaving Caenorhabditis elegans worms. We demonstrate a simple mounting technique that gently overlays worms growing on a standard Nematode Growth Media (NGM) agar block with a glass coverslip, permitting animals to be recorded at high-resolution during unrestricted movement and behavior. With this technique, we use the sensitive Ca2+ reporter GCaMP5 to record changes in intracellular Ca2+ in the serotonergic Hermaphrodite Specific Neurons (HSNs) as they drive egg-laying behavior. By co-expressing mCherry, a Ca2+-insensitive fluorescent protein, we can track the position of the HSN within ~ 1 &#181;m and correct for fluctuations in fluorescence caused by changes in focus or movement. Simultaneous, infrared brightfield imaging allows for behavior recording and animal tracking using a motorized stage. By integrating these microscopic techniques and data streams, we can record Ca2+ activity in the C. elegans egg-laying circuit as it progresses between inactive and active behavior states over tens of minutes.

Ratiometric Calcium Imaging of Individual Neurons in Behaving Caenorhabditis Elegans

This protocol describes the use of genetically encoded Ca2+ reporters to record changes in neural activity in behaving Caenorhabditis elegans worms. &#8195;

꼬마 선 충 을 행동에 개별 뉴런의 비율 칼슘 이미징

It has become increasingly clear that neural circuit activity in behaving animals differs substantially from that seen in anesthetized or immobilized ...

Combining Optogenetics with Artificial microRNAs to Characterize the Effects of Gene Knockdown on Presynaptic Function within Intact Neuronal Circuits

The purpose of this protocol is to characterize the effect of gene knockdown on presynaptic function within intact neuronal circuits. We describe a workflow on how to combine artificial microRNA (miR)-mediated RNA interference with optogenetics to achieve selective stimulation of manipulated presynaptic boutons in acute brain slices. The experimental approach involves the use of a single viral construct and a single neuron-specific promoter to drive the expression of both an optogenetic probe and artificial miR(s) against presynaptic gene(s). When stereotactically injected in the brain region of interest, the expressed construct makes it possible to stimulate with light exclusively the neurons with reduced expression of the gene(s) under investigation. This strategy does not require the development and maintenance of genetically modified mouse lines and can in principle be applied to other organisms and to any neuronal gene of choice. We have recently applied it to investigate how the knockdown of alternative splice isoforms of presynaptic P/Q-type voltage-gated calcium channels (VGCCs) regulates short-term synaptic plasticity at CA3 to CA1 excitatory synapses in acute hippocampal slices. A similar approach could also be used to manipulate and probe the neuronal circuitry in vivo.

This protocol provides a workflow on how to combine artificial microRNA-mediated RNA interference with optogenetics to stimulate specifically presynaptic boutons with reduced expression of selective gene(s) within intact neuronal circuits.

연 접 기능 그대로 신경 회로 내에 유전자 최저 효과 특성 인공 microRNAs Optogenetics 결합

The purpose of this protocol is to characterize the effect of gene knockdown on presynaptic function within intact neuronal circuits. We describe a ...

Imaging Dendritic Spines in Caenorhabditis elegans

Dendritic spines are specialized sites of synaptic innervation modulated by activity and serve as substrates for learning and memory. Recently, dendritic spines have been described for DD GABAergic neurons as the input sites from presynaptic cholinergic neurons in the motor circuit of Caenorhabditis elegans. This synaptic circuit can now serve as a powerful new in vivo model of spine morphogenesis and function that exploits the facile genetics and ready accessibility of C. elegans to live-cell imaging. 
This protocol describes experimental strategies for assessing DD spine structure and function. In this approach, a super-resolution imaging strategy is used to visualize the intricate shapes of actin-rich dendritic spines. To evaluate the DD spine function, the light-activated opsin, Chrimson, stimulates the presynaptic cholinergic neurons, and the calcium indicator, GCaMP, reports the evoked calcium transients in postsynaptic DD spines. Together, these methods comprise powerful approaches for identifying genetic determinants of dendritic spines in C. elegans that could also direct spine morphogenesis and function in the brain.

Imaging Dendritic Spines in Caenorhabditis elegans

Dendritic spines are important cellular features of the nervous system. Here live imaging methods are described for assessing the structure and function of dendritic spines in C. elegans.&#160;These approaches support the development of mutant screens for genes that define dendritic spine shape or function.

Caenorhabditis elegans의 수지상 가시 이미징

Dendritic spines are specialized sites of synaptic innervation modulated by activity and serve as substrates for learning and memory. Recently, dendritic ...