우리는 원고의 비판적 읽기를 위해 열 플레이트 분석, 플라이 주식에 대한 블루밍턴 Drosophila 증권 센터, Galko 연구소 회원 제안에 대한 애벌레 열 프로브 분석, 숀 스위니을 개발하기 위해 열 프로브 설계, 다니엘 Babcock위한 기독교 랜드리 감사드립니다. 이 작품은 MJG 및 연구 경력 (AVG)에 소수 액세스를위한 NIH MARC U-STAR 교육 그랜트 (휴스턴 - 다운 타운 학자 아카데미 대학 T34GM079088)에 NIH R01 NS069828에 의해 지원되었다.

<ol>
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J., Ma, Q. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Nociceptors--noxious+stimulus+detectors.">Nociceptors--noxious stimulus detectors.</a> Neuron. 55, 353-364 (2007).</li><li>Hwang, R. Y. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Nociceptive+neurons+protect+Drosophila+larvae+from+parasitoid+wasps.">Nociceptive neurons protect Drosophila larvae from parasitoid wasps.</a> Curr. Biol. 17, 2105-2116 (2007).</li><li>Xiang, Y. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Light-avoidance-mediating+photoreceptors+tile+the+Drosophila+larval+body+wall.">Light-avoidance-mediating photoreceptors tile the Drosophila larval body wall.</a> Nature. 468, 921-926 (2010).</li><li>Grueber, W. 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우리는 공개 할게 없다.

여기에 설명된 assays는 질적 및 양적 유해 열 자극에 대한 반응에 대해 다른 genotypes의 유충을 평가하는 데 사용할 수 있습니다. 열 프로브 분석의 주요 기능은 자극은 단지 하나의 현장에서 주어진다는 것입니다. 이것은 아마도 아마도 유일한 작은 탐침으로 연락을 세그먼트에있는 클래스 IV 뉴런-것들의 집합과 바로 인접 세그먼트 11에서 이들의 발포로 안내합니다. 때문에 현지 자극, 열 프로브 분석은 지역과 같은 뜨거운 난로에 문의 손으로로서 특정 몸에 현지되는 유해 자극을 감지의 일반적인 감각 경험을 모방한 것이었 지요. ) I) 사용자가 유충에 프로브를 적용있는 압력, II 프로브의 정확한 위치 열 프로브 분석의 단점은 그것이 가능성이 세 가지 요소에 의한 것이 일부 사용자에 대한 사용자의 다양성을 가지고있다는 것입니다 기본 nociceptive 뉴런, 그리고, 3) 정확한 중앙에 상대적으로 유충에르 어떤 탐사선이 접점 유충의 표면에서. 우리는 이전에 주어진 온도를 1로 자신의 탈퇴 대기 시간을 기준으로 비 답장, 느린 조치 및 신속한 조치로 분류 애벌레의 양적 전략을보고했다. 여기에서 우리는 더 높은 온도에 애벌레 대응에 대한 리포트입니다. 흥미롭게도, 우리는 애벌레 열 nociceptive 답변에 천정이있다는 것을 발견하고이 천장 52 54 사이에 놓여 있다고 ° C. 이것은 일시적인 수용체 잠재적인 (TRP) 52보다 높은 온도에서 게이팅 수있는 채널 ° C를 소유하지 않는 애벌레 나타낼 수 또는 그것이 사용되는 뉴런이나 근육이 시작하거나 심지어 그들 aversive 탈퇴에서 작동하기 전에 운동 반응이 손상되어 수행하는 것이 좋습니다 수 있습니다. 우리는 또한 금단의 진폭의 다른 분석 반응을 이용하여 응답의 &quot;견고&quot;의 지표로서 롤의 숫자 중 하나를보고합니다. Naively 한이러한 매개 변수가 자극을 증가 온도 또​​는 시간으로 증가할 것이라고 기대합니다. 놀랍게도, 우리는이 사건 아닌 것을 알게됩니다. 애벌레는 유해 범위 (42 ℃)의 낮은 끝에 온도에서 긴 시간 동안 자극하는 것은 더 많은 롤을 보여 주며 더 많은 시간이 애벌레는 높은 온도 (48-52 ° C)에서 탐지보다 압연 보냈다. 이것은 유해 온도 창 안에 그것이 주로 응답의 진폭을 결정하는 노출의 기간입니다 것이 좋습니다. 애벌레는 매우 유해 온도에 노출되어 있기 때문에 (48-52 ° C) 매우 빠른 속도로 평균 반응, 그들은 시간의 긴 기간 동안 덜 유해 온도에 노출 애벌레 많은 롤스로 전시하지 않습니다.에게 여기에 보고된 응답 진폭의 분석은 다른 genotypes이나 환경 조작가를 비교할 수있는 더불어 또 다른 양적 차원을 추가합니다. 열 플레이트 분석의 주된 기능은 유해에 대한 글로벌 노출과 관련된 것입니다더위. 따라서 그것은 뜨거운 난로를 만져보다 가열 가마솥에 앉아 동물에 더 가깝다입니다. 유충이 글로벌 노출에 대한 행동 반응은 현지 자극에 따라 관찰하는 것보다 더 복잡 실험실에서, 야생에서 세계적으로 유해 자극을 경험할 수 있습니다했을 때 그것은 명확하지 않지만. 또 다른 3 언급 열 플레이트 분석의 강점은, 유충을 만지는 것은 프로토콜의 구성 요소가 아니므로 그것이 작은 사용자로 사용자의 다양성을 가지고있다는 것입니다. 유일한 상당한 편차는 각 행동 개시하고 이것이 반복 시청 / 친숙로 최소화할 수있을 때 정의하는 것으로 보인다. assays 사이에 흥미로운 차이는 aversive 동작이 시작되는 온도입니다. 이것은 열 프로브보다 열 플레이트 분석에서 훨씬 낮은 수 있습니다. 열 프로브 (머리와 꼬리 인상)으로 연락 애벌레가 전시 예비 동작 ° C 열 작은지면으로 ~ 27에서 관찰 헤드 록 음악의 한 형식으로 상관 관계를 수전자 분석. 그것은이 반응은 &#39;고통&#39;보다 더 &#39;불편&#39;을 반영하는 것이 가능합니다. 우리는 심지어 높은에서 압류를 흔든다의 상관 관계, 그리고 마비를 감찰하지 않은 (최대 48 ° C) 열 프로브 분석 기온 그리고 신체의 하나 이상의 지역에서 소성 감각 신경 세포의 중요한 대량임을있을 수 있습니다 이러한 행동을 가지고 필요했습니다. 그 세계 자극 로컬로 행동을 실행하는 데 충분하지가의 연결 응답 합산 포함됩니다 나타내는 열 프로브로 관찰 nociceptive 임계값의 하단 끝 아래 - 흥미롭게도, 발작과 마비 행동은 온도 (37 ° C ~ 34)에서 관찰된다 열 프로브의 응용. 이러한 온도 실제로 애벌레에 대한 유해으로 인식하고있는가 대부분의 경우하지 마비 동작을 시작하고 이후 플라이 음식을 복구하는 데 사용할 수 있습니다 애벌레는 (그림 5C) 살아남을 것을 관찰에 의해 지원됩니다. 또한 인수를 지원하는 그 열기 PL먹은 분석은 nociceptive 답변을 읽고있는 것은 잘 알려진 nociceptive 감각 뉴런에서 시냅스 전송을 차단하는 것이 관찰된 행동 (그림 6) 대부분의 대기 시간을 증가한다는 사실입니다. 높은 온도를 일으키는 행동을위한 대기 시간에는 증가가 없다는 것을 관찰 MD-Gal4을 표현하지 않는 감각 뉴런이 동작이 필요할 수 있습니다하는 것이 좋습니다. 글로벌 분석의 급속 가열 물 한 방울의 지역 분석 및 침수에 작은 금속 탐침의 핫 팁 - 요컨대, 두 assays는 정의된 온도의 유해 열 자극에 개별 애벌레를 노출하는 방식입니다. 유전 배경 및 / 변화 또는 환경 조건 (예 : 플러스 또는 마이너스 조직 손상에) 변화에 노출의 Drosophila의 유충의 행동 반응은, 공부하고 이러한 assays를 사용하여 계량하실 수 있습니다. 궁극적으로 이러한 assays 결과에서 우리가 더 NOC을 제어하는​​ 유전자 네트워크를 이해하는 데 도움이됩니다Drosophila 및 기타 관련 종의 iception.

<table border="1"><tbody><tr><td> 시약의 이름 </td><td> 회사 </td><td> 카탈로그 번호 </td><td> 댓글 </td></tr><tr><td> 온도 프로브 </td><td> 프로 - 데브 공학 </td><td> 수요에 맞춤 건설한 </td><td> 연락처 정보는 요청시 제공될 수 있습니다 </td></tr><tr><td> 드라이 바스 인큐베이터 </td><td> 피셔 과학 </td><td> 11-718 </td><td> 한 고체 가열 블록 및 16mm 우물 1 개 가열 블록 </td></tr><tr><td> Leica DFC290 12v/400mA 컬러 카메라 </td><td> Leica 마이크로 시스템즈 </td><td> 12730080 </td><td> 이와 동등한 카메라는 할 것입니다. </td></tr><tr><td> Leica MZ6 현미경 </td><td> Leica 마이크로 시스템즈 </td><td> MZ6 줌 몸에 부품 번호 (광학 캐리어)는 10445614입니다 </td><td></td></tr><tr><td> SCHOTT 에이스 Modulamp 단위 </td><td> SCHOTT 북미, 주식 회사 </td><td> A20500 </td><td></td></tr><tr><td> SCHOTT 듀얼 거위 목처럼 생긴 것 23 인치 광섬유 라이트 가이드 </td><td> SCHOTT 북미, 주식 회사 </td><td> SCHOTT A08575 </td><td></td></tr><tr><td> 온도 제어 장치 </td><td> TSCI 사. </td><td> 사용자 정의가 내장 </td><td> 자세한 내용은 요청시 제공될 수 있습니다 </td></tr><tr><td> 자이스 혈구 Stemi 2,000 현미경 </td><td> 자이스 혈구 </td><td> NT55-605 </td><td> 이와 동등한 현미경할 것입니다. </td></tr><tr><td> 집게 </td><td> FST </td><td> FS-1670 </td><td></td></tr><tr><td> 1mm 메쉬 </td><td> Genesee 과학 </td><td> 57-101 </td><td></td></tr><tr><td> 화필 </td><td> 딕 Blick 미술 재료 </td><td> 06762-1002 </td><td></td></tr><tr><td> 자외선 crosslinker </td><td> 피셔 과학 </td><td> 1,199,289 </td><td></td></tr><tr><td> Coplin 단지 </td><td> 피셔 과학 </td><td> 08-816 </td><td></td></tr><tr><td> 10ml의 비이커 </td><td> 피셔 과학 </td><td> 02-540C </td><td></td></tr><tr><td> 디에틸 에테르 </td><td> 피셔 과학 </td><td> E138-500 </td><td></td></tr><tr><td> 35 X 10 밀리미터 폴리스티렌 페트리 접시 </td><td> 매 </td><td> 351,008 </td><td> 우리는 테스트하지 않았습니다 대안 요리. </td></tr><tr><td> 유리 현미경 슬라이드 </td><td> 코닝 </td><td> 26,003 </td><td></td></tr><tr><td> 열전대 </td><td> 오메가 엔지니어링 주식 회사 </td><td> HH802U </td><td></td></tr><tr><td> 비닐 조각 </td><td> 오피스 디포 </td><td> 480,009 </td><td></td></tr><tr><td> Microcentrifuge 관 </td><td> Denville 과학주식 회사 </td><td> C-2170 </td><td></td></tr></tbody></table>

local and global methods of assessing thermal nociception in drosophila larvae

이 문서에서는 Drosophila의 애벌레에서 열 nociception 공부를 assays를 보여줍니다. 두 번째는 대부분의 도매 (전역) 활성화 또는 이러한 모든 뉴런 3을 포함하면서 한 분석은 열 nociceptors 1,2의 (로컬) spatially - 제한 자극을 포함한다. 함께 이러한 기법은 Drosophila nociceptive 감각 뉴런의 행동 기능의 시각화 및 부량을 허용합니다. Drosophila의 유충은 온도 nociception, evolutionarily 종의 1,2 걸쳐 보존되고 잠재적으로 유해한 온도 감각 반응을 연구하기 위해 설립 모델 시스템입니다. 이러한 연구 Drosophila의 장점은 respons, 모든 metazoans에서와 같이, 그 신경계의 상대적인 단순 성과 Drosophila에서 근본적인 생물학 4-6의 분자 기초를 해부하는 데 사용할 수있는 유전자 기술의 고도화이다유해 열 자극에 전자는 일반적으로 제시 자극 7 &quot;nocifensive&quot;aversive 탈퇴 관련이 있습니다. 이러한 자극은 무료 신경 엔딩이나 nociceptors 통해 감지하고 organismal 응답의 진폭은 유해 자극에게 8을받은 nociceptors의 숫자에 달려있다. Drosophila에서는 photoreceptors 10들은 최근에 발견된 역할 이외에 유해 열 및 기계적 자극에 9 감지 클래스 IV의 돌기를 arborization 감각 뉴런이다. 잘 발달 수준에서 연구되어 이들 뉴런은 장벽 표피 시트 위에 arborize과 11,12 거의 모든 상피 세포와 연락처를 확인하십시오. 그들은 두 번째 순서 뉴런에 연결할 수있는 중추 신경계 11 복부 신경 코드로 각 클래스 IV 뉴런 프로젝트의 단일 축삭 그 뇌로 프로젝트. 기준 조건 하에서 nociceptive 감각 신경상대적으로 높은 임계값에 도달할 때까지 NS는 해고되지 않습니다. 여기에 설명된 assays는 조사가 기준선 행동 반응이나, 아마도, 다음과 같은 조직의 손상을 ensues sensitization.를 정할 수 있습니다 각 분석은 유해 열 자극에 뚜렷한하지만 관련 locomotory 행동 반응을 유발하고 Drosophila의 애벌레에 열 nociception의 다양한 측면을 시각화하고 수치 연구자를 허용합니다. assays는 원하는 genotypes의 유충이나 nociception에 영향을 미칠 수도 서로 다른 환경 조건 하에서 사육 유충에 적용할 수 있습니다. 열 nociception는 수종에 걸쳐 보존되어 있기 때문에, Drosophila의 유전자 해부에서 gleaned 결과 가능성이 척추 동물 등 다른 생물에서 열 nociception 우리의 이해를 통지합니다.

Watch this Scientific Journal Video about Local and Global Methods of Assessing Thermal Nociception in Drosophila Larvae at JoVE.com

Local and Global Methods of Assessing Thermal Nociception in Drosophila Larvae | Neuroscience | Video

Local and Global Methods of Assessing Thermal Nociception in Drosophila Larvae

이 문서에서 우리는 assays가 열 nociception 공부를 입증<em&gt; Drosophila</em&gt; 애벌레. 한 분석은 열 nociceptors의 (로컬) spatially - 제한 자극을 포함<sup&gt; 1,2</sup&gt; 반면 두 번째는 대부분의 도매 (전역) 활성화 또는 이러한 모든 뉴런을 포함<sup&gt; 3</sup&gt;. 함께 이러한 기술의 행동 기능의 시각화 및 부량을 허용<em&gt; Drosophila</em&gt; nociceptive 감각 뉴런.

에서 열 Nociception를 평가 지역 및 글로벌 방법<em&gt; Drosophila</em&gt; 애벌레

In this article, we demonstrate assays to study thermal nociception in Drosophila larvae. One assay involves spatially-restricted (local) stimulation of ...

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Local and Global Methods of Assessing Thermal Nociception in Drosophila Larvae

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15.7K 조회수.  The University of Texas MD Anderson Cancer Center. 이 문서에서 우리는 assays가 열 nociception 공부를 입증 Drosophila 애벌레. 한 분석은 열 nociceptors의 (로컬) spatially - 제한 자극을 포함 1,2 반면 두 번째는 대부분의 도매 (전역) 활성화 또는 이러한 모든 뉴런을 포함 3. 함께 이러한 기술의 행동 기능의 시각화 및 부량을 허용 Drosophila nociceptive 감각 뉴런.

비디오: Local and Global Methods of Assessing Thermal Nociception in Drosophila Larvae

Electrophysiological Measurements and Analysis of Nociception in Human Infants

Pain is an unpleasant sensory and emotional experience. Since infants cannot verbally report their experiences, current methods of pain assessment are based on behavioural and physiological body reactions, such as crying, body movements or changes in facial expression. While these measures demonstrate that infants mount a response following noxious stimulation, they are limited: they are based on activation of subcortical somatic and autonomic motor pathways that may not be reliably linked to central sensory processing in the brain. Knowledge of how the central nervous system responds to noxious events could provide an insight to how nociceptive information and pain is processed in newborns.
The heel lancing procedure used to extract blood from hospitalised infants offers a unique opportunity to study pain in infancy. In this video we describe how electroencephalography (EEG) and electromyography (EMG) time-locked to this procedure can be used to investigate nociceptive activity in the brain and spinal cord.
This integrative approach to the measurement of infant pain has the potential to pave the way for an effective and sensitive clinical measurement tool.

The assessment and treatment of pain in infants is difficult because infants cannot verbally report their experience. In this video we describe quantitative electrophysiological methods and analysis techniques that can be used to measure the response to noxious events from the infant nervous system.

Electrophysiological 측정과 인간 유아의 Nociception 분석

Pain is an unpleasant sensory and emotional experience. Since infants cannot verbally report their experiences, current methods of pain assessment are ...

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신경과학

Methods to Assay Drosophila Behavior

Drosophila melanogaster, the fruit fly, has been used to study molecular mechanisms of a wide range of human diseases such as cancer, cardiovascular disease and various neurological diseases1. We have optimized simple and robust behavioral assays for determining larval locomotion, adult climbing ability (RING assay), and courtship behaviors of Drosophila. These behavioral assays are widely applicable for studying the role of genetic and environmental factors on fly behavior. Larval crawling ability can be reliably used for determining early stage changes in the crawling abilities of Drosophila larvae and also for examining effect of drugs or human disease genes (in transgenic flies) on their locomotion. The larval crawling assay becomes more applicable if expression or abolition of a gene causes lethality in pupal or adult stages, as these flies do not survive to adulthood where they otherwise could be assessed. This basic assay can also be used in conjunction with bright light or stress to examine additional behavioral responses in Drosophila larvae. Courtship behavior has been widely used to investigate genetic basis of sexual behavior, and can also be used to examine activity and coordination, as well as learning and memory. Drosophila courtship behavior involves the exchange of various sensory stimuli including visual, auditory, and chemosensory signals between males and females that lead to a complex series of well characterized motor behaviors culminating in successful copulation. Traditional adult climbing assays (negative geotaxis) are tedious, labor intensive, and time consuming, with significant variation between different trials2-4. The rapid iterative negative geotaxis (RING) assay5 has many advantages over more widely employed protocols, providing a reproducible, sensitive, and high throughput approach to quantify adult locomotor and negative geotaxis behaviors. In the RING assay, several genotypes or drug treatments can be tested simultaneously using large number of animals, with the high-throughput approach making it more amenable for screening experiments.

Methods to Assay Drosophila Behavior

Drosophila melanogaster is a genetically and behaviorally tractable model system that has been used to understand the molecular and cellular basis of many important biological processes for over a century 1. Drosophila has been well exploited to gain insights into the genetic basis of fly behavior.

분석의 방법 Drosophila 행동

Drosophila melanogaster, the fruit fly, has been used to study molecular mechanisms of a wide range of human diseases such as cancer, cardiovascular ...

Visualization of Proprioceptors in Drosophila Larvae and Pupae

Proprioception is the ability to sense the motion, or position, of body parts by responding to stimuli arising within the body. In fruitflies and other insects proprioception is provided by specialized sensory organs termed chordotonal organs (ChOs) 2. Like many other organs in Drosophila, ChOs develop twice during the life cycle of the fly. First, the larval ChOs develop during embryogenesis. Then, the adult ChOs start to develop in the larval imaginal discs and continue to differentiate during metamorphosis.
The development of larval ChOs during embryogenesis has been studied extensively 10,11,13,15,16. The centerpiece of each ChO is a sensory unit composed of a neuron and a scolopale cell. The sensory unit is stretched between two types of accessory cells that attach to the cuticle via specialized epidermal attachment cells 1,9,14. When a fly larva moves, the relative displacement of the epidermal attachment cells leads to stretching of the sensory unit and consequent opening of specific transient receptor potential vanilloid (TRPV) channels at the outer segment of the dendrite 8,12. The elicited signal is then transferred to the locomotor central pattern generator circuit in the central nervous system.
Multiple ChOs have been described in the adult fly 7. These are located near the joints of the adult fly appendages (legs, wings and halters) and in the thorax and abdomen. In addition, several hundreds of ChOs collectively form the Johnston's organ in the adult antenna that transduce acoustic to mechanical energy 3,5,17,4.
In contrast to the extensive knowledge about the development of ChOs in embryonic stages, very little is known about the morphology of these organs during larval stages. Moreover, with the exception of femoral ChOs 18 and Johnston's organ, our knowledge about the development and structure of ChOs in the adult fly is very fragmentary.
Here we describe a method for staining and visualizing ChOs in third instar larvae and pupae. This method can be applied together with genetic tools to better characterize the morphology and understand the development of the various ChOs in the fly.

Visualization of Proprioceptors in Drosophila Larvae and Pupae

A method to immunostain and visualize chordotonal organs in larvae and pupae of Drosophila melanogaster is described.

의 Proprioceptors의 시각화<em&gt; Drosophila</em&gt; 애벌레와 Pupae

Proprioception is the ability to sense the motion, or position, of body parts by responding to stimuli arising within the body. In fruitflies and other ...

Appetitive Associative Olfactory Learning in Drosophila Larvae

In the following we describe the methodological details of appetitive associative olfactory learning in Drosophila larvae. The setup, in combination with genetic interference, provides a handle to analyze the neuronal and molecular fundamentals of specifically associative learning in a simple larval brain.
Organisms can use past experience to adjust present behavior. Such acquisition of behavioral potential can be defined as learning, and the physical bases of these potentials as memory traces1-4. Neuroscientists try to understand how these processes are organized in terms of molecular and neuronal changes in the brain by using a variety of methods in model organisms ranging from insects to vertebrates5,6. For such endeavors it is helpful to use model systems that are simple and experimentally accessible. The Drosophila larva has turned out to satisfy these demands based on the availability of robust behavioral assays, the existence of a variety of transgenic techniques and the elementary organization of the nervous system comprising only about 10,000 neurons (albeit with some concessions: cognitive limitations, few behavioral options, and richness of experience questionable)7-10.
Drosophila larvae can form associations between odors and appetitive gustatory reinforcement like sugar11-14. In a standard assay, established in the lab of B. Gerber, animals receive a two-odor reciprocal training: A first group of larvae is exposed to an odor A together with a gustatory reinforcer (sugar reward) and is subsequently exposed to an odor B without reinforcement 9. Meanwhile a second group of larvae receives reciprocal training while experiencing odor A without reinforcement and subsequently being exposed to odor B with reinforcement (sugar reward). In the following both groups are tested for their preference between the two odors. Relatively higher preferences for the rewarded odor reflect associative learning - presented as a performance index (PI). The conclusion regarding the associative nature of the performance index is compelling, because apart from the contingency between odors and tastants, other parameters, such as odor and reward exposure, passage of time and handling do not differ between the two groups9.

Appetitive Associative Olfactory Learning in Drosophila Larvae

Drosophila larvae are able to associate odor stimuli with gustatory reward. Here we describe a simple behavioral paradigm that allows the analysis of appetitive associative olfactory learning.

에 Appetitive 연관 강한 학습<em&gt; Drosophila</em&gt; 애벌레

In the following we describe the methodological  details of appetitive associative olfactory learning in Drosophila larvae. The setup, in combination with ...

Light Preference Assay to Study Innate and Circadian Regulated Photobehavior in Drosophila Larvae

Light acts as environmental signal to control animal behavior at various levels. The Drosophila larval nervous system is used as a unique model to answer basic questions on how light information is processed and shared between rapid and circadian behaviors. Drosophila larvae display a stereotypical avoidance behavior when exposed to light. To investigate light dependent behaviors comparably simple light-dark preference tests can be applied. In vertebrates and arthropods the neural pathways involved in sensing and processing visual inputs partially overlap with those processing photic circadian information. The fascinating question of how the light sensing system and the circadian system interact to keep behavioral outputs coordinated remains largely unexplored. Drosophila is an impacting biological model to approach these questions, due to a small number of neurons in the brain and the availability of genetic tools for neuronal manipulation. The presented light-dark preference assay allows the investigation of a range of visual behaviors including circadian control of phototaxis.

Light Preference Assay to Study Innate and Circadian Regulated Photobehavior in Drosophila Larvae

Here we describe a light-dark preference test for Drosophila larva. This assay provides information about innate and circadian regulation of light sensing and processing photobehavior.

에 타고난 및 시간주기 규제 Photobehavior을 공부하는 빛 환경의 분석<em&gt; 초파리</em&gt; 애벌레

Light acts as environmental signal to control animal behavior at various levels. The Drosophila larval nervous system is used as a unique model to answer ...

Optogenetic Perturbation of Neural Activity with Laser Illumination in Semi-intact Drosophila Larvae in Motion

Drosophila larval locomotion is a splendid model system in developmental and physiological neuroscience, by virtue of the genetic accessibility of the underlying neuronal components in the circuits1-6. Application of optogenetics7,8 in the larval neural circuit allows us to manipulate neuronal activity in spatially and temporally patterned ways9-13. Typically, specimens are broadly illuminated with a mercury lamp or LED, so specificity of the target neurons is controlled by binary gene expression systems such as the Gal4-UAS system14,15. In this work, to improve the spatial resolution to "sub-genetic resolution", we locally illuminated a subset of neurons in the ventral nerve cord using lasers implemented in a conventional confocal microscope. While monitoring the motion of the body wall of the semi-intact larvae, we interactively activated or inhibited neural activity with channelrhodopsin16,17 or halorhodopsin18-20, respectively. By spatially and temporally restricted illumination of the neural tissue, we can manipulate the activity of specific neurons in the circuit at a specific phase of behavior. This method is useful for studying the relationship between the activities of a local neural assembly in the ventral nerve cord and the spatiotemporal pattern of motor output.

Optogenetic Perturbation of Neural Activity with Laser Illumination in Semi-intact Drosophila Larvae in Motion

Here we describe a protocol for optogenetic manipulation of motoneuronal activity while monitoring changes in motor output (muscle contraction) in semi-intact Drosophila larvae using lasers within a conventional confocal microscope. This technique enables researchers to achieve local perturbation of neural activity in a few neuromeres to elucidate the dynamics of motor circuits.

세미 그대로의 레이저 조명과 신경 활동의 Optogenetic 섭동<em&gt; 초파리</em모션&gt; 유충

Drosophila larval locomotion is a splendid model system in developmental and physiological neuroscience, by virtue of the genetic accessibility of the ...

Foraging Path-length Protocol for Drosophila melanogaster Larvae

The Drosophila melanogaster larval path-length phenotype is an established measure used to study the genetic and environmental contributions to behavioral variation. The larval path-length assay was developed to measure individual differences in foraging behavior that were later linked to the foraging gene. Larval path-length is an easily scored trait that facilitates the collection of large sample sizes, at minimal cost, for genetic screens. Here we provide a detailed description of the current protocol for the larval path-length assay first used by Sokolowski. The protocol details how to reproducibly handle test animals, perform the behavioral assay and analyze the data. An example of how the assay can be used to measure behavioral plasticity in response to environmental change, by manipulating feeding environment prior to performing the assay, is also provided. Finally, appropriate test design as well as environmental factors that can modify larval path-length such as food quality, developmental age and day effects are discussed.

Foraging Path-length Protocol for Drosophila melanogaster Larvae

We provide a detailed protocol for a Drosophila melanogaster foraging path-length assay. We discuss the preparation and handling of test animals, how to perform the assay and analyze the data.

대한 꼴 경로 길이 프로토콜<em&gt; 노랑 초파리</em&gt; 애벌레

The Drosophila melanogaster larval path-length phenotype is an established measure used to study the genetic and environmental contributions to behavioral ...

Novel Assay for Cold Nociception in Drosophila Larvae

How organisms sense and respond to noxious temperatures is still poorly understood. Further, the mechanisms underlying sensitization of the sensory machinery, such as in patients experiencing peripheral neuropathy or injury-induced sensitization, are not well characterized. The genetically tractable Drosophila model has been used to study the cells and genes required for noxious heat detection, which has yielded multiple conserved genes of interest. Little is known however about the cells and receptors important for noxious cold sensing. Although, Drosophila does not survive prolonged exposure to cold temperatures (&#8804;10 &#186;C), and will avoid cool, preferring warmer temperatures in behavioral preference assays, how they sense and possibly avoid noxious cold stimuli has only recently been investigated.
Here we describe and characterize the first noxious cold (&#8804;10 &#186;C) behavioral assay in Drosophila. Using this tool and assay, we show an investigator how to qualitatively and quantitatively assess cold nociceptive behaviors. This can be done under normal/healthy culture conditions, or presumably in the context of disease, injury or sensitization. Further, this assay can be applied to larvae selected for desired genotypes, which might impact thermosensation, pain, or nociceptive sensitization. Given that pain is a highly conserved process, using this assay to further study&#160;thermal nociception will likely glean important understanding of pain processes in other species, including vertebrates.

Novel Assay for Cold Nociception in Drosophila Larvae

Here we demonstrate a novel assay to study cold nociception in Drosophila larvae. This assay utilizes a custom-built Peltier probe capable of applying a focal noxious cold stimulus and results in quantifiable cold-specific behaviors. This technique will allow further cellular and molecular dissection of cold nociception.

콜드 통각에 대한 소설 분석<em&gt; 초파리</em&gt; 애벌레

How organisms sense and respond to noxious temperatures is still poorly understood. Further, the mechanisms underlying sensitization of the sensory ...

A Temperature Gradient Assay to Determine Thermal Preferences of Drosophila Larvae

Many animals, including the fruit fly, Drosophila melanogaster, are capable of discriminating minute differences in&#160;environmental temperature, which enables them to seek out their preferred thermal landscape. To define the temperature preferences of larvae over a defined linear range, we developed an assay using a temperature gradient. To establish a single-directional gradient, two aluminum blocks are connected to independent water baths, each of which controls the temperature of individual blocks. The two blocks set the lower and upper limits of the gradient. The temperature gradient is established by placing an agarose-coated aluminum plate over the two water-controlled blocks so that the plate spans the distance between them. The ends of the aluminum plate that is set on the top of the water blocks defines the minimum and maximum temperatures, and the regions in-between the two blocks form a linear temperature gradient. The gradient assay can be applied to&#160;larvae of different ages and can be used to identify mutants that exhibit phenotypes, such as those with mutations affecting genes encoding TRP channels and opsins, which are required for temperature discrimination.

A Temperature Gradient Assay to Determine Thermal Preferences of Drosophila Larvae

Here, we present a protocol to determine the preferred environmental temperature of Drosophila larvae using a continuous thermal gradient.

초파리 애벌레의 열 환경 설정을 결정 하는 온도 분석 결과

Many animals, including the fruit fly, Drosophila melanogaster, are capable of discriminating minute differences in&#160;environmental temperature, which ...

Electrophysiological Methods for Measuring Photopigment Levels in Drosophila Photoreceptors

The Drosophila G-protein-coupled photopigment rhodopsin (R) is composed of a protein (opsin) and a chromophore. The activation process of rhodopsin is initiated by photon absorption-inducing isomerization of the chromophore, promoting conformational changes of the opsin and resulting in a second dark-stable photopigment state (metarhodopsin, M). Investigation of this bi-stable photopigment using random mutagenesis requires simple and robust methods for screening mutant flies. Therefore, several methods for measuring reductions in functional photopigment levels have been designed. One such method exploits the charge displacements within the photopigment following photon absorption and the huge amounts of photopigment molecules expressed in the photoreceptors. This electrical signal, named the early receptor potential (or early receptor current), is measured by a variety of electrophysiological methods (e.g., electroretinogram and whole-cell recordings) and is linearly proportional to functional photopigment levels. The advantages of this method are the high signal-to-noise ratio, direct linear measurement of photopigment levels, and independence of phototransduction mechanisms downstream to rhodopsin or metarhodopsin activation. An additional electrophysiological method called prolonged depolarizing afterpotential (PDA) exploits the bi-stability of Drosophila photopigment and the absorption-spectral differences of fly R and M pigment states. The PDA is induced by intense blue light, converting saturating amounts of rhodopsin to metarhodopsin, resulting in the failure of light-response termination for an extended time in darkness, but it can be terminated by metarhodopsin to rhodopsin conversion using intense orange light. Since the PDA is a robust signal that requires massive photopigment conversion, even small defects in the biogenesis of the photopigment lead to readily detected abnormal PDA. Indeed, defective PDA mutants led to the identification of novel signaling proteins important for phototransduction.

Electrophysiological Methods for Measuring Photopigment Levels in Drosophila Photoreceptors

We present a protocol to electrophysiologically characterize bi-stable photopigments: (i) exploiting the charge displacements within the photopigment molecules following photon-absorption and their huge amount in the photoreceptors, and (ii) exploiting the absorption-spectra differences of rhodopsin and metarhodopsin photopigment states. These protocols are useful to screen for mutations affecting bi-stable photopigment systems.&#160;

초파리 광수용체에서 광안료 수준을 측정하기 위한 전기생리학적 방법

The Drosophila G-protein-coupled photopigment rhodopsin (R) is composed of a protein (opsin) and a chromophore. The activation process of rhodopsin is ...