우리는 비판적 원고를 읽기 위해 차가운 프로브 분석, 플라이 주식에 대한 블루밍턴 초파리 증권 센터 및 Galko 실험실 구성원의 초기 단계 개발을위한 사라 우 및 카밀 그레이엄합니다. 이 작품은 NIH NRSA HNT에 (NIH F31NS083306)에 의해 NIH R01NS069828, R21NS087360 및 MJG에 대한 기초 연구에서 텍사스 MD 앤더슨 클라크 원정대의 대학에 의해 지원되었다.

애벌레의 제조 1 <ol><li> 25 ºC 배양기에서 주식이나 유전 십자가를 올립니다. <ol><li> 십자가를 배양하는 경우, 20 ~ 25 명 처녀 여성과 일반 옥수수 가루 플라이 미디어를 포함하는 바이알 당 15 ~ 20 명의 남성을 사용합니다. 여성은 음식의 새로운 유리 병에 전송하기 전에 약 48 시간 동안 알을 낳기 할 수 있습니다. </li></ol></li><li> 4-5 일 분 계란 바닥 후, 부드럽게 약해서 식품 및 유충에 물 스트림을 분출하여 원하는 유전자형 3 차 령 유충을 수집하고, 중소 깨끗한 페트리 접시에 내용물을 부어 (60mm X 15 mm). </li><li> 다음 부드럽게 테스트되는 큰 &quot;방황 3 차 령&quot;또는 작은 병약 유충 크기 배제하여 집게 나 붓 유충 정렬. 원하지 않는 유전자 마커 또는 분산을 포함 폐기 유충. </li><li> 실험자 전에 &quot;블라인드&quot;가 될 수 있도록 분류 유충이 배치됩니다 동료 실험실 구성원 레이블 요리 또는 용기를 가지고perimental 조건 또는 해당되는 경우 유충의 유전자형. 실험자들은 실험실 부재에 의해 생성 된 키를 이용하여 상기 실험 후 수집 된 데이터를 복호화하여 라벨을 적용 할 수있다. </li><li> scoopula를 사용하여 (35mm X 10mm 또는 60mm X 15mm)을 실온에서 물과 깨끗한 라벨 페트리 접시 가득 반쯤로 신선 식품의 다임 크기 양 옮긴다. </li><li> 집게 또는 붓을 사용하여 조심스럽게 음식에 정렬 된 유충을 이동 (시험이 20 분을 예상하는 경우 기아 또는 탈수를 방지하기 위해). </li></ol> 2. 콜드 프로브 분석 <ol><li> 이 원하는 온도로 냉각 할 수있는 몇 분을 허용 차가운 프로브 유닛을 켭니다. 낮은 온도로 설정하는 경우, 결로 가능성이 프로브를 따라 형성됩니다. </li><li> 실험실로 초과 수분을 닦아은 유충에 적용하기 전에 닦아주십시오. 즉시 사용, 장소는 모두 보온에 프로브에 뚜껑을 절연하고 깨끗한 팁을 유지하고 손상을 방지하기 위해하지 않는 경우. </li><li> 어두운 가동 비닐의 박편 상으로 중간의 제 3 령 유충을 배치 명 시야 현미경 (일반적으로, 노트북 바인더로부터 절단 작은 조각을 사용). 콘트라스트 시각화하고, 프로브와 올바르게 정렬 건드리지 않고 유충을 이동시키는 검은 비닐 AIDS. </li><li> 대비를 너무 빨리 건조되는 유충을 방지하기 위해 현미경 (자재 표 참조)과 중간 휘도 (50-75 %의 최대 휘도)로 광 유닛 (자재 표 참조)를 조정한다. </li><li> 그들은 결과를 혼동 수 있기 때문에 첫 정상 연동 운동을 나타내지 않는 유충을 폐기하십시오. 물을 페트리 접시 달리 유충 정상적인 운동을 억제 비닐로 버티로부터 유충 습윤되어야한다. 물은 유충의 주위에 웅덩이 안된다. </li><li> 고전으로 유충을 이동 비닐을 사용하여, 전후 몸통 축에 90도 각도로 충 향해 손에 프로브를 사전RECT 위치 (도 1A 참조). </li><li> 동양은 프로브의 팁은 부드럽게 프로브 현미경 단계에 약 45도 각도 유충의 중간 지느러미 표면을 가로 질러 누워있다. </li><li> 프로브 접촉시, 실험실 타이머를 시작합니다. 계속 전진 또는 후진시키면서 약간 애벌레 표피의 표면을 들여 충분한 하향 압력을 적용한다. </li><li> 최대 10 초 동안 또는 냉간 유발 행동 반응이 관찰 될 때까지 자리에 프로브를 잡고 - 둘 중 먼저 발생. 그런 다음 프로브를 제거합니다. </li><li> 관찰 된 행동 응답 및 응답 대기 시간을 기록합니다. 10 초 내에 응답하지 않는 애벌레는 &quot;비 반응자&quot;로 간주됩니다. 지연은 또한 대응 대해 응답 내성 / 진폭의 변화를 측정하는데 사용될 수있다. </li><li> 유충을 취소하고 다음을 준비합니다. </li><li> 시험 유충의 원하는 개수 (N 3 개 세트에 도달 할 때까지 반복 단계 2.3-2.11 = 20-40 유충 U이었다) 여기에 나오지도. </li></ol>

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미국 특허 출원 (CL)은 차가운 프로브의 설계에 보류 중입니다. 차가운 프로브 부품 번호 및 건설의 세부 사항에 대한 추가 정보와 도구 디자인에 대한 자세한 상담은 요청에 따라 제공됩니다.

여기에 설명 된 분석은 정 성적 및 정량적으로 여러 유전 배경, 환경 적 영향들, 및 / 또는 손상을 유도 조건에서 통각 유충 또는 통각 과민을 평가하는데 사용될 수있다. 이 분석은이 도구 냉 자극의 초점 응용이 가능하기 때문에 하나의 특히 저온에 응답 말초 감각 뉴런의 서브셋의 기능을 평가할 수있다. 흥미롭게도,이 추위 유발 행동은 열 (7), (11)에 ?...

초파리는 새로운 보존 유전자와 복잡한 행동을 기초 신경 회로의 식별을위한 매우 유용한 것으로 입증되었습니다. 파리 정교한 유전 툴킷 통각 5, 6, 7의 세포 및 분자 염기 해부 정확한 유전 적 신경 조작 1,2,3,4 허용 간략화 신경계를 제공한다. 유충 부드러운 촉감 8, 9, 10, 유해한 열 11, 12, 13 및 유해 자극 (4)의 기계적 감각에 대한 행동 분석 주어진 이러한 분석에 특히 유용 </sup&gt; 11은 이미 설립 투명 유충 표피는 표피와 하부 감각 뉴런의 라이브 영상 또는 고정을 허용하고있다. 최근 유해 감기위한 분석은 또한 우리는 여기에서 더 상세히 설명 7 개발되었다. 벌금, 원뿔 스쳐 차가운 프로브를 사용하여, 우리는 표시하거나 거친 기계적 또는 고온 자극 (7), (8) 후, 11 초파리 애벌레 전시 부드러운 터치 다음 정상 운동 동안 관찰 행동, 구별 차가운 별 반응 일련의 행동, 그 . 차가운 특정 행동은 강력한 전신 수축 (CT), U 자형 (미국)에 후방 세그먼트 (PR) 및 전방의 동시 인상 및 후방 세그먼트의 45-90º 인상을 포함한다. 이러한 행동의 유병률은 감소 온도 만의 각 피크 증가살짝 다른 추운 온도. 최근 연구는 CT의 응답이 유해 열이나 강한 기계적 자극 (7)에 응답 것보다 다른 주변 감각 신경에 의해 매개되는 것을 의미한다. 많은 척추 통각 수용기 같은 초파리 다수의 돌기 (MD) 말초 감각 신경 세포는 표피 (1) 위에 arborize 수지상 복합 구조를 갖는다. MD 뉴런 복부 신경 코드 (14)에 자신의 축삭을 투영 모든 유충 바디 부분에 존재한다. MD 감각 신경 돌기의 형태에 기초하여 네 개의 서로 다른 클래스 (I-IV)로 분리하고, 감각 기능 4, 9, 10, 15, 16, 17을 변화 하였다. 클래스 IV 뉴런은 유충 횡 체 롤 응답 요구되지만고온 또는 열악한 기계적 자극 4 클래스 III 뉴런 부드러운 터치 응답 (9), (10)에 필요한 단지 초기 활성화되지 않고, 또한 냉간 유발 행동 반응 (7)에 요구한다. 모두 클래스 III 및 IV 클래스 신경 자극 9, 10, 17, 19 7, 11, 18, 유해하고 비에 유해한 행동 반응을 촉진하기 위해 이산 일시적 수용체 전위 (TRP) 채널을 사용한다. 또한, 애벌레 통각은 20 세포 및 행동 수준 12, 21, 다음과 같은 부상을 민감합니다. 여기에 설명 된 분석은 허용 quantificatio보통, 하나의 잠재적 인 N은 상온 (18-22 ºC)에 유해 냉기 (10 ≤ ºC)까지 저온, 무해한 차가운 (11-17 ºC)로 행동 반응을 변경. 이 분석에 사용되는 저온 직접 클래스 III 감각 뉴런을 활성화하고 질적 양적 7을 분석 할 강력한 재현성 칼슘 증가 냉간 유발 행동 반응을 유도 할 수있다. 이 분석은 초기 통각, 통각 과민 또는 통각 소성 영향 유전 적 및 환경 두 인자를 결정하는 다양한 환경 조건에 노출 된 유충 (변경 영양, 부상되는 약물)에뿐만 아니라 거의 모든 유전형의 유충에 적용될 수있다. thermosensation 여러 종에 걸쳐 유비쿼터스 것을 감안할 때,이 분석은 통각의 연구를위한 유용한 도구를 제공하며, 새로운 유전자 대상 또는 개선 신경 세포의 상호 작용을 발견 할 수있다척추 동물의 통각에 대한 우리의 이해. 맞춤 제작 된 콜드 프로브 (콜드 프로브, 소재의 표 참조)를 통한 열 전도 알루미늄 샤프트와 원추형 팁을 냉각시키는 펠티에 소자를 제어 폐쇄 루프 온도를 이용한다. 서미스터는 알루미늄 원추형 팁 내부에 내장되고, 제어부의 실시간 온도를보고한다. 히트 싱크 팬 (열 제어 유닛 자재 표 참조)를 얻을 수있는 펠티에 효과의 열 부하 (QC)이므로 (22-0 °의 C)의 바람직한 온도 범위를 조절하는 열전 모듈에 부착된다. 콜드 프로브 팁의 유해 냉 자극 전후방 단부로부터 등거리 세그먼트 (들), 등쪽 정중선에 손으로 적용되는 유충 (약 A4 세그먼트,도 1a 참조). 차가운 자극에 대응하여, 유충은 일반적으로 10 초 컷오프에서 세 냉간 유발 행동 중 하나를 생산 : 몸 전체를수축 (CT), U 자형 (US) 또는 (결과 설명) 후부 세그먼트 (PR)의 인상에 전방 및 후방 세그먼트의 45-90º 인상. 이러한 행동의 아무도는 정상적인 연동 운동이나 먹이를 찾아 다니는 행동 중에 수행되지 않습니다. 이러한 동작은 부드러운 촉감 응답 및 높은 온도 또는 유해한 기계적 자극에 대한 반응 혐오 압연에서 구별된다.

<table><tbody><tr><td>Cold Probe</td><td>Pro-Dev Engineering</td><td>Custom-built on demand</td><td>Part numbers and construction details can be provided on request</td></tr><tr><td>Thermal Control Unit</td><td>TE Technology</td><td>Custom Built enclosure</td><td>Part numbers and construction details can be provided on request</td></tr><tr><td>Zeiss Stemi 2000 microscope</td><td>Zeiss</td><td>NT55-605</td><td></td></tr><tr><td>Fiber-Lite MI-150 High Intensity Illuminator</td><td>Dolan-Jenner Industries.</td><td>A20500</td><td></td></tr><tr><td>Schott Dual Gooseneck 23 inch Fiber Optic Light Guide</td><td>Schott North America, Inc.</td><td>Schott A08575</td><td></td></tr><tr><td>Forceps</td><td>FST</td><td>FS-1670</td><td>Used to sort and handle larvae. Be sure to smooth and blunt forceps tips slightly to lower the risk of accidently puncturing or injuring the larvae</td></tr><tr><td>Paintbrush</td><td>Dick Blick Art Materials</td><td>06762-1002</td><td>Used to sort and handle larvae. It is helpful if the paintbrush is damp during use.</td></tr><tr><td>35 mm x 10 mm Polystyrene Petri dish</td><td>Falcon</td><td>351008</td><td></td></tr><tr><td>60mm x 10 mm Polystyrene Petri dish</td><td>Falcon</td><td>351007</td><td></td></tr><tr><td>Piece of black vinyl (at least 2 inches x 2 inches)</td><td></td><td></td><td>Used to provide contrast and orient larvae to the cold probe</td></tr><tr><td>Fisherbrand Scoopula Spatula</td><td>Fisher Scientific</td><td>14-357Q</td><td>Used to move food</td></tr><tr><td>Kimtech Science Kimwipes</td><td>Fisher Scientific</td><td>06-666A</td><td>Used to dry the larvae and cold probe if there is excess moisture</td></tr></tbody></table>

novel assay for cold nociception in drosophila larvae

어떻게 생물 감지하고 여전히 제대로 이해 유해 온도에 반응한다. 또한, 말초 신경 병증 또는 손상 유발 과민성 발생한 환자와 같은 감각 기계 과민성, 기본 메커니즘은 잘 특징 아니다. 유 전적으로 취급 용이 초파리 모델은, 유해한 열 감지에 필요한 세포의 유전자를 연구하는 데 사용 된 이는 관심의 여러 보존 유전자를 산출했다. 약간은 세포 및 유해 차가운 ​​감지에 대한 중요한 수용체에 대해 그러나 알려져있다. 초파리는 낮은 온도 (≤10 ° C)에 장시간 노출 살아남을하지 않으며, 유해 차가운 자극을가 감지 방법 행동 선호도 분석에 따뜻한 온도를 선호 멋진 피하고 가능한 피할 것,하지만 최근 조사되었다. 여기에 우리가 설명하고의 첫 번째 유해 감기 (≤10 ° C) 행동 분석의 특징초파리. 이 도구 및 분석을 사용하여, 우리는 질적 및 양적 차가운 침해 행동을 평가하는 방법 조사관을 보여줍니다. 이 질병, 부상 또는 감작의 맥락에서 아마도 일반 / 건강한 문화 조건에서 수행, 또는 할 수 있습니다. 또한,이 분석은 thermosensation 통증 또는 통각 과민에 영향을 줄 수있는 원하는 유전자형 선정 유충에 적용될 수있다. 고통이 더 가능성이 척추 동물을 포함한 다른 종, 통증 프로세스의 중요한 이해를 수집합니다 것입니다 열 통각을 연구하기 위해이 분석을 사용하여, 고도의 보존 과정 감안할.

가끔 일시, 머리 회전 및 방향 (22)의 변화를 포함하는 연동 모션 초파리 유충 이동. 그러나 초점 유해 냉 자극 프로그램, 유충 전시 유해한 열 및 기계적 자극에 혐오 횡 롤 달리 고유 일련의 행동에 응답. 이러한 동작은 부드러운 촉감 8, 9, 10 또는 실온 프로브의 접촉에 응답도 ?...

Watch this Scientific Journal Video about Novel Assay for Cold Nociception in Drosophila Larvae at JoVE.com

Novel Assay for Cold Nociception in Drosophila Larvae

여기에서 우리는 초파리 유충의 차가운 통각을 연구하는 새로운 분석을 보여줍니다. 이 분석은 초점 유해 냉 자극을인가 할 수있는 맞춤식 펠티에 프로브를 이용하여 정량화하고 초기 특정 행동을 초래한다. 이 기술은 차가운 통각의 추가 세포 및 분자 해부를 허용합니다.

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 ...

novel-assay-for-cold-nociception-in-drosophila-larvae

Research

JoVE Journal

Neuroscience

Novel Assay for Cold Nociception in Drosophila Larvae

7.7K 조회수.  UT MD Anderson Cancer Center.  여기에서 우리는 초파리 유충의 차가운 통각을 연구하는 새로운 분석을 보여줍니다. 이 분석은 초점 유해 냉 자극을인가 할 수있는 맞춤식 펠티에 프로브를 이용하여 정량화하고 초기 특정 행동을 초래한다. 이 기술은 차가운 통각의 추가 세포 및 분자 해부를 허용합니다. 

비디오: Novel Assay for Cold Nociception in Drosophila Larvae

Local and Global Methods of Assessing Thermal Nociception in Drosophila Larvae

In this article, we demonstrate assays to study thermal nociception in Drosophila larvae. One assay involves spatially-restricted (local) stimulation of thermal nociceptors1,2 while the second involves a wholesale (global) activation of most or all such neurons3. Together, these techniques allow visualization and quantification of the behavioral functions of Drosophila nociceptive sensory neurons.
The Drosophila larva is an established model system to study thermal nociception, a sensory response to potentially harmful temperatures that is evolutionarily conserved across species1,2. The advantages of Drosophila for such studies are the relative simplicity of its nervous system and the sophistication of the genetic techniques that can be used to dissect the molecular basis of the underlying biology4-6 In Drosophila, as in all metazoans, the response to noxious thermal stimuli generally involves a "nocifensive" aversive withdrawal to the presented stimulus7. Such stimuli are detected through free nerve endings or nociceptors and the amplitude of the organismal response depends on the number of nociceptors receiving the noxious stimulus8. In Drosophila, it is the class IV dendritic arborization sensory neurons that detect noxious thermal and mechanical stimuli9 in addition to their recently discovered role as photoreceptors10. These neurons, which have been very well studied at the developmental level, arborize over the barrier epidermal sheet and make contacts with nearly all epidermal cells11,12. The single axon of each class IV neuron projects into the ventral nerve cord of the central nervous system11 where they may connect to second-order neurons that project to the brain.
Under baseline conditions, nociceptive sensory neurons will not fire until a relatively high threshold is reached. The assays described here allow the investigator to quantify baseline behavioral responses or, presumably, the sensitization that ensues following tissue damage. Each assay provokes distinct but related locomotory behavioral responses to noxious thermal stimuli and permits the researcher to visualize and quantify various aspects of thermal nociception in Drosophila larvae. The assays can be applied to larvae of desired genotypes or to larvae raised under different environmental conditions that might impact nociception. Since thermal nociception is conserved across species, the findings gleaned from genetic dissection in Drosophila will likely inform our understanding of thermal nociception in other species, including vertebrates.

Local and Global Methods of Assessing Thermal Nociception in Drosophila Larvae

In this article, we demonstrate assays to study thermal nociception in Drosophila larvae. One assay involves spatially-restricted (local) stimulation of thermal nociceptors1,2 while the second involves a wholesale (global) activation of most or all such neurons3. Together, these techniques allow visualization and quantification of the behavioral functions of Drosophila nociceptive sensory neurons.

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 ...

연구

신경과학

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 ...

Novel Whole-tissue Quantitative Assay of Nitric Oxide Levels in Drosophila Neuroinflammatory Response

Neuroinflammation is a complex innate immune response vital to the healthy function of the central nervous system (CNS). Under normal conditions, an intricate network of inducers, detectors, and activators rapidly responds to neuron damage, infection or other immune infractions. This inflammation of immune cells is intimately associated with the pathology of neurodegenerative disorders, such as Parkinson&#39;s disease (PD), Alzheimer&#39;s disease and ALS. Under compromised disease states, chronic inflammation, intended to minimize neuron damage, may lead to an over-excitation of the immune cells, ultimately resulting in the exacerbation of disease progression. For example, loss of dopaminergic neurons in the midbrain, a hallmark of PD, is accelerated by the excessive activation of the inflammatory response. Though the cause of PD is largely unknown, exposure to environmental toxins has been implicated in the onset of sporadic cases. The herbicide paraquat, for example, has been shown to induce Parkinsonian-like pathology in several animal models, including Drosophila melanogaster. Here, we have used the conserved innate immune response in Drosophila to develop an assay capable of detecting varying levels of nitric oxide, a cell-signaling molecule critical to the activation of the inflammatory response cascade and targeted neuron death. Using paraquat-induced neuronal damage, we assess the impact of these immune insults on neuroinflammatory stimulation through the use of a novel, quantitative assay. Whole brains are fully extracted from flies either exposed to neurotoxins or of genotypes that elevate susceptibility to neurodegeneration then incubated in cell-culture media. Then, using the principles of the Griess reagent reaction, we are able to detect minor changes in the secretion of nitric oxide into cell-culture media, essentially creating a primary live-tissue model in a simple procedure. The utility of this model is amplified by the robust genetic and molecular complexity of Drosophila melanogaster, and this assay can be modified to be applicable to other Drosophila tissues or even other small, whole-organism inflammation models.

Novel Whole-tissue Quantitative Assay of Nitric Oxide Levels in Drosophila Neuroinflammatory Response

Levels of the inflammatory cell-signaling molecule nitric oxide (NO) are commonly assayed using Griess reagent. In this protocol, we have created a modified Griess assay utilizing live Drosophila brain tissue in order to detect the secretion of NO in a simple, quantifiable and highly repeatable method.

Neuroinflammation is a complex innate immune response vital to the healthy function of the central nervous system (CNS). Under normal conditions, an ...

면역학 및 감염병학

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 ...

A Single-fly Assay for Foraging Behavior in Drosophila

For many animals, hunger promotes changes in the olfactory system in a manner that facilitates the search for appropriate food sources. In this video article, we describe an automated assay to measure the effect of hunger or satiety on olfactory dependent food search behavior in the adult fruit fly Drosophila melanogaster. In a light-tight box illuminated by red light that is invisible to fruit flies, a camera linked to custom data acquisition software monitors the position of six flies simultaneously. Each fly is confined to walk in individual arenas containing a food odor at the center. The testing arenas rest on a porous floor that functions to prevent odor accumulation. Latency to locate the odor source, a metric that reflects olfactory sensitivity under different physiological states, is determined by software analysis. Here, we discuss the critical mechanics of running this behavioral paradigm and cover specific issues regarding fly loading, odor contamination, assay temperature, data quality, and statistical analysis.

A Single-fly Assay for Foraging Behavior in Drosophila

In this video article, we describe an automated assay to measure the effect of hunger or satiety on olfactory dependent food search behavior in the adult fruit fly Drosophila melanogaster.

먹이 찾아 돌아다 행동에 대한 단일 비행 분석에<em&gt; 초파리</em

For many animals, hunger promotes changes in the olfactory system in a manner that facilitates the search for appropriate food sources. In this video ...

An Improved Assay and Tools for Measuring Mechanical Nociception in Drosophila Larvae

Published assays for mechanical nociception in Drosophila have led to variable assessments of behavior. Here, we fabricated, for use with Drosophila larvae, customized metal nickel-titanium alloy (nitinol) filaments. These mechanical probes are similar to the von Frey filaments used in vertebrates to measure mechanical nociception. Here, we demonstrate how to make and calibrate these mechanical probes and how to generate a full behavioral dose-response from subthreshold (innocuous or non-noxious range) to suprathreshold (low to high noxious range) stimuli. To demonstrate the utility of the probes, we investigated tissue damage-induced hypersensitivity in Drosophila larvae. Mechanical allodynia (hypersensitivity to a normally innocuous mechanical stimulus) and hyperalgesia (exaggerated responsiveness to a noxious mechanical stimulus) have not yet been established in Drosophila larvae. Using mechanical probes that are normally innocuous or probes that typically elicit an aversive behavior, we found that Drosophila larvae develop mechanical hypersensitization (both allodynia and hyperalgesia) after tissue damage. Thus, the mechanical probes and assay that we illustrate here will likely be important tools to dissect the fundamental molecular/genetic mechanisms of mechanical hypersensitivity.

An Improved Assay and Tools for Measuring Mechanical Nociception in Drosophila Larvae

The goal of this protocol is to show how to perform an improved assay for mechanical nociception in Drosophila larvae. We use the assay here to demonstrate that mechanical hypersensitivity (allodynia and hyperalgesia) exists in Drosophila larvae.

드로소필라 애벌레의 기계적 고치절 측정을 위한 향상된 분석 및 도구

Published assays for mechanical nociception in Drosophila have led to variable assessments of behavior. Here, we fabricated, for use with Drosophila ...

행동분석학

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 ...

Behavioral Assays for Optogenetic Manipulation of Neural Circuits in Drosophila melanogaster

Optogenetics has become a fundamental technique in neuroscience, enabling precise control of neuronal activity through light stimulation. This study introduces easy-to-implement setups for applying optogenetic methods in Drosophila melanogaster. Two optogenetic tools, CsChrimson, a red-light-activated cation channel, and GtACR2, a blue-light-activated anion channel, were employed in four experimental approaches. Three of these approaches involve single-fly experiments: (1) a blue-light optogenetic thermotactic positional preference assay targeting temperature-sensitive heating cells, (2) a red-light optogenetic positional preference assay activating bitter sensing neurons, and (3) a proboscis extension response assay activating the sweet-sensing neurons. The fourth approach (4) is a fly maze setup to assess avoidance behaviors using multiple flies. The ability to manipulate neural activity temporally and spatially offers powerful insights into sensory processing and decision-making, underscoring the potential of optogenetics to advance our knowledge of neural function. These methods provide an accessible and robust framework for future research in neuroscience to enhance the understanding of specific neural pathways and their behavioral outcomes.

This paper presents methods for optogenetic manipulation in Drosophila melanogaster, utilizing CsChrimson and GtACR2 to activate and silence specific neurons. Four experiments are described to utilize optogenetics to explore thermotactic and gustatory behaviors, providing insights into the underlying neural mechanisms governing these processes.

Drosophila melanogaster에서 신경 회로의 광유전학적 조작을 위한 행동 분석

Optogenetics has become a fundamental technique in neuroscience, enabling precise control of neuronal activity through light stimulation. This study ...

Local Heat Probe Assay: A Method to Assess Nociception of High Temperatures in Drosophila Larvae

Source: Chattopadhyay, A., et al. <a href="https://www.jove.com/video/3837/" target="_blank">Local and Global Methods of Assessing Thermal Nociception in Drosophila Larvae</a>. J. Vis. Exp. (2012).
Drosophila larvae have dedicated sensory neurons&#8211;nociceptors&#8211;whose response to potentially harmful thermal stimuli triggers aversive&#8211;or so-called &#34;nocifensive&#34;&#8211;behaviors in the animal. This video describes a method to assess thermal nociception by recording larval behavior in response to local application of a heat probe onto the cuticle. This method is, therefore, called the local heat probe assay.

국소 열 프로브 분석: 초파리 유충의 고온 통각을 평가하는 방법

Source: Chattopadhyay, A., et al. Local and Global Methods of Assessing Thermal Nociception in Drosophila Larvae. J. Vis. Exp. (2012).
Drosophila larvae ...

콜드 통각에 대한 소설 분석

요약

더 많은 비디오 탐색

에서 열 Nociception를 평가 지역 및 글로벌 방법

의 Proprioceptors의 시각화

Novel Whole-tissue Quantitative Assay of Nitric Oxide Levels in Drosophila Neuroinflammatory Response

에 Appetitive 연관 강한 학습

에 타고난 및 시간주기 규제 Photobehavior을 공부하는 빛 환경의 분석

먹이 찾아 돌아다 행동에 대한 단일 비행 분석에

드로소필라 애벌레의 기계적 고치절 측정을 위한 향상된 분석 및 도구

초파리 애벌레의 열 환경 설정을 결정 하는 온도 분석 결과

Drosophila melanogaster에서 신경 회로의 광유전학적 조작을 위한 행동 분석

국소 열 프로브 분석: 초파리 유충의 고온 통각을 평가하는 방법