Name: mass histology to quantify neurodegeneration in drosophila
Uploaded: 2016-12-15T12:30:00
Description: Watch this Scientific Journal Video about Mass Histology to Quantify Neurodegeneration in Drosophila at JoVE.com

This work was supported by grants to D.K. from the Medical Research Foundation of Oregon and from NIH/NINDS (NS047663). E.S. was supported by a training grant from the NIH (T32AG023477).

1. 칼라의 머리를 고정하고 파라핀에 임베드 주 : 정착 프로세스의 모든 단계는 흄 후드에서 수행되어야한다. 건강 위험 포즈를하지 않으면 서 메틸 벤조 에이트는, 흄 후드에서 처리되지 않는 경우에 압도 될 수있는 매우 독특한 냄새를 가지고있다. <ol><li> 파리를 마취 전에 클로로포름 15 ㎖의 99 % 에탄올 30 ㎖에 빙초산을 5 ㎖ (클로로포름 및 아세트산을 혼합하지 않음)에 추가하여 Carnoy ?...

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설명 된 방법은 초파리의 뇌 퇴행을 정량하는 방법을 제공한다. 특정 세포 유형을 계산하는 등의 다른 방법이, 신경 퇴행을 식별하는 데 사용될 수 있지만,이 방법의 장점은,보다 일반적으로 적용 할 수 있다는 것이다. 세포를 계수하는 것은 이들 세포를 확실하게 특정 항체 또는 항상 가능하지 않은 세포 특이 마커의 발현을 사용하여 식별 될 수있는 것을 요구한다. 또한,이 극적으로 다른 ...

기대 수명의 증가와 함께, 알츠하이머 나 파킨슨 병 같은 신경 퇴행성 질환은 일반 인구 증가 건강에 위협이되고있다. 국립 보건원에 따르면 115 만명 전세계 기본 상당한 진전이 많은 분야에서 이들 질환의 적어도 일부에 관여하는 유전자와 위험 요인을 식별 하였지만 2050 치매에 의해 영향을받을 것으로 예상되는 분자 메커니즘은 여전히 ​​알 수없는 여부를 잘 이해한다. 예쁜 꼬마 선충과 초파리 melanogaster의 같은 간단한 무척추 동물 모델 생물은 짧은 라이프 사이클, 자손의 많은 수의, 잘 설립 때로는 독특한 유전 및 분자 방법 1의 가용성을 포함한 신경 퇴행성 질환의 메커니즘을 연구하는 실험 다양한 장점을 제공합니다 -12. 또한, 이러한 유기체는 편견 의무가 있습니다신경 퇴행성 표현형에 자신의 악화 또는 의한 개선 효과에 의해 이러한 질병에 기여하는 요인을 식별 할 수있는 상호 작용 화면. 이러한 유전자의 상호 작용을 분석하고 노화 효과를 평가하는 신경 퇴행을 검출하고 그 심각도를 측정하는 정량적 프로토콜을 필요로한다. 숫자 성능 값 13-21을 제공하는 등 후각 학습, 부정적인 geotaxis, 또는 빠른 phototaxis으로 초파리의 행동 측면을 측정 할 때이 평가는 비교적 쉽게 수행 할 수 있습니다. 이는 뉴런을 카운트함으로써 신경 세포의 생존에 대한 효과를 결정하는 것도 가능하다. PD의 영향, 그리고 심지어 그 후, 결과는 22-24 논란 한 도파민 뉴런 같이 명확하게 식별되는 특정 인구 집중 그러나이 경우에만 가능하다. 여기에 설명 된 프로토콜 파라핀 직렬 섹션을 수행 칼라 방법을 사용하는 방법즉 원래 초파리 25 해부학 뇌 돌연변이를 분리하여 사용하고 Böhl 하이젠 베르크에 의해 개발되었다. 칼라 방법의 사용은 다음에 저온부, vibratome 부, 비닐 부 26-28에 포함하도록 구성되었다. 여기서,이 방법은 퇴행성 신경 표현형 16,21,29-32에 파리 발전 액포를 측정하는 데 사용할 수있는 전체 플라이 헤드의 직렬 부분을 얻기 위해 사용된다. 이러한 측정들은 특정 뇌 영역에서 수행 될 수 있거나 뇌 전체를 커버 할 수있다; 후자의 방법은 노화 동안 관찰 한도 약한 퇴행성 표현형을 식별 할 수 있습니다. 칼라를 사용하는 경우 마지막으로, 20 파리에는 하나 준비로 처리 할 수 ​​있습니다뿐만 아니라 적은 시간이 소요되지만 인해 약간의 변화에 ​​아티팩트를 최소화 제어와 같은 준비 실험 파리의 분석 가능 준비.

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>Name of the Reagent/Equipment</td>
			<td>Company</td>
			<td>Catalog Number</td>
			<td></td>
		</tr>
		<tr>
			<td>Collar</td>
			<td>Genesee Scientific</td>
			<td>TS 48-100</td>
			<td>We are using custom made collars that are made from one piece of metal instead of layers as the ones by Genesee. A discription to make collars can be found at http://flybrain.neurobio.arizona.edu/Flybrain/html/atlas/fluorescent/index.html&#160;</td>
		</tr>
		<tr>
			<td>Rubber ice cube tray for embedding</td>
			<td>Household store</td>
			<td></td>
			<td>The size can be made to fit by glueing in additional walls&#160;</td>
		</tr>
		<tr>
			<td>Crystallizing dish</td>
			<td>Fisher Scientific company</td>
			<td>08-762-3</td>
			<td></td>
		</tr>
		<tr>
			<td>Ether</td>
			<td>Fisher Scientific Company</td>
			<td>E138-1</td>
			<td></td>
		</tr>
		<tr>
			<td>Ethanol</td>
			<td>Decon Laboratories Inc.</td>
			<td>2701</td>
			<td></td>
		</tr>
		<tr>
			<td>Choloroform</td>
			<td>Fisher Scientific Company</td>
			<td>C298-500</td>
			<td></td>
		</tr>
		<tr>
			<td>Glacial Acetic Acid</td>
			<td>Fisher Scientific Company</td>
			<td>A38-212</td>
			<td></td>
		</tr>
		<tr>
			<td>Methylbenzoate</td>
			<td>Fisher Scientific Company</td>
			<td>M205-500</td>
			<td>Distinct Odor</td>
		</tr>
		<tr>
			<td>Use in fume hood</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Low Melting Point Paraffin Wax</td>
			<td>Fisher Scientific Company</td>
			<td>T565</td>
			<td>Make sure to keep extra melted in a 65&#176;C waterbath</td>
		</tr>
		<tr>
			<td>Microtome</td>
			<td>Leica Biosystems</td>
			<td>Reichert Jung 2040 Autocut</td>
			<td></td>
		</tr>
		<tr>
			<td>Microscope Slide</td>
			<td>Fisher Scientific Company</td>
			<td>12-550D</td>
			<td></td>
		</tr>
		<tr>
			<td>Microscope Cover Glass</td>
			<td>Fisher Scientific Company</td>
			<td>12-545-M</td>
			<td></td>
		</tr>
		<tr>
			<td>SafeClear</td>
			<td>Fisher Scientific Company</td>
			<td>314-629</td>
			<td>Three different containers for washes</td>
		</tr>
		<tr>
			<td>Vertical Staining Jar with Cover</td>
			<td>Ted Pella Inc.&#160;</td>
			<td>432-1</td>
			<td></td>
		</tr>
		<tr>
			<td>Permount</td>
			<td>Fisher Scientific Company</td>
			<td>SP15-500</td>
			<td></td>
		</tr>
		<tr>
			<td>Poly-L-lysine Solution</td>
			<td>Sigma Life Science</td>
			<td>P8290-500</td>
			<td></td>
		</tr>
	</tbody>
</table>

mass histology to quantify neurodegeneration in drosophila

알츠하이머 병 (AD) 또는 파킨슨 병 (PD)와 같은 진보적 인 신경 퇴행성 질환은 전 세계적으로 사람의 건강에 위협이 증가한다. 포유 동물 모델은 병원성의 기전에 중요한 통찰력을 제공하고 있지만, 함께 높은 비용 포유류 시스템의 복잡성은 자신의 사용을 제한하고 있습니다. 따라서, 간단하지만 잘 확립 된 초파리 모델 시스템은 이러한 질병에 영향을 분자 경로를 조사하기위한 대안을 제공합니다. 행동 적자 외에도, 신경 퇴행성 질환 등 신경 죽음과 axonopathy 등의 조직 학적 표현형을 특징으로한다. 신경 세포의 변성을 정량화하고는 유전 적, 환경 적 요인에 의해 영향을받는 방법을 결정하기 위해, 우리는 성인 플라이 뇌에서 공포 측정을 기반으로하는 조직 학적 접근 방법을 사용합니다. 계통 오차의 영향을 최소화하기 위해 직접적으로 제어 및 경험치의 섹션을 비교하나 준비 erimental 파리, 우리는 파라핀 섹션의 &#39;칼라&#39;방법을 사용합니다. 신경 퇴행은 플라이 뇌 개발 액포의 크기 및 / 또는 개수를 측정하여 평가한다. 이것은 하나의 관심의 특정 영역에 집중하거나 완전한 헤드 걸쳐 직렬 부분을 취득하여 전체 뇌를 분석함으로써 수행 될 수있다. 정상적인 노화 중에 발생 따라서,이 방법은, 하나는 심한 변성뿐만 아니라 몇 부분 만 감지 상대적으로 약한 표현형뿐만 아니라 측정 할 수 있습니다.

전체 플라이 헤드를 포함 눈 안료 (33)에 의해 스테인드 시리얼 섹션에 설명 된 방법의 결과를 사용. 이것의 일부는 각각의 헤드의 부분이 위로부터 아래로 도시되어도 1b에 도시된다. 다른 파리에서 섹션이 예에서 왼쪽에서 오른쪽으로 볼 수 있습니다. 방향과 파리의 식별을 용이하게하기 위해, 눈이없는 비행 (사인 oculis)의 위치 3 (화살표, <stron...

Watch this Scientific Journal Video about Mass Histology to Quantify Neurodegeneration in Drosophila at JoVE.com

Mass Histology to Quantify Neurodegeneration in Drosophila

초파리 널리 신경 퇴행을 연구하기위한 모델 시스템으로 사용된다. 이 프로토콜은, 뇌의 공포 형성에 의해 결정된 바와 같이, 정량 할 수있는 변성하는 방법을 설명한다. 또한 인해 하나의 샘플로 처리 및 절편 제어 및 실험 파리에 의해 실험 절차에 효과를 최소화 할 수 있습니다.

질량 조직학은에 신경 퇴행을 정량화하기<em&gt; 초파리</em

Progressive neurodegenerative diseases like Alzheimer&#39;s disease (AD) or Parkinson&#39;s disease (PD) are an increasing threat to human health ...

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Mass Histology to Quantify Neurodegeneration in Drosophila

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One of the most important goals in neuroscience is to understand the molecular cues that instruct early stages of synapse formation. As such it has become ...

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Going beyond single gene function to cut deeper into gene regulatory networks requires multiple mutations combined in a single animal.  Such analysis of ...

Assaying Locomotor, Learning, and Memory Deficits in Drosophila Models of Neurodegeneration

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

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Magnetic resonance imaging (MRI) allows for the delineation between normal and abnormal tissue on a macroscopic scale, sampling an entire tissue volume ...

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