These studies were supported by grants from the National Institutes of Health, National Institute for Alcoholism and Alcohol Abuse to M.G. (P20AA017828, R01AA020634, P50 AA022537). The authors thank Jill Bettinger for helpful discussions and Jacqueline DeLoyht for technical assistance.

분석 전에 1 일 <ol><li> 짧은 (1-5 분) CO 2에서 11 (단일 섹스)의 그룹 신선한 식품 유리 병에 파리를 수집합니다. </li><li> 파리는 환경 제어 공간에서 음식을 유리 병에 (일반적으로 25 ° C, 상대 습도 60 %, 12 시간 빛 / 어둠 사이클) O / N을 복구 할 수 있습니다. </li><li> 계획된 실험에 적합한 최종 농도 (들)에 정제 (≥18 MΩ) 물에 순수 (100 %) 에탄올을 희석하여 에탄올 용액 (들)을 준비합니다. 솔루션 (들) RT O / N로 복귀 할 수 있습니다. 참고 : 에탄올의 희석은 발열이다. </li></ol> 분석 2. 일 <ol><li> 파리의 각 바이알을 테스트하려면, 깨끗하고 빈 식품 유리 병을 (i)를 준비; 즉, 검사 바이알 (ⅱ) 새로운 셀룰로오스 아세테이트 플러그, (ⅲ) 실리콘 마개 및 (IV)의 에탄올 용액 (표 3 참조)를 1 ㎖. </li><li> 55-65%에 20 ~ 25 ° C의 온도와 상대 습도를 조절하여 테스트 룸을 준비합니다. </li><li> 다른 노동자 되세요튜브의 각 그룹에 고유 코드를 할당하고 이후의 코드를 기록합니다. 몇 분 동안 적응 시험 방에 파리 코드화 된 튜브를 놓습니다. </li><li> 2.3에서 비행 병에 코드를 일치하도록 빈 테스트 튜브 레이블. </li><li> 표 1과 유사한 스프레드 시트의 열로 코드 (하나의 열 / 병)을 입력하여 하드 카피 테스트 로그를 구축합니다. </li><li> 가이드로 테스트 로그를 사용하여 테스트 룸에서 배열을 일치로 파리와 빈 테스트 튜브 코드화 된 식품 유리 병을 준비. 참고 : 테스트하는 튜브의 합리적인 최대 수 (24) (즉, 4 병 각 6 세트). </li><li> 전송 일치 / 표시 빈 테스트 튜브로 음식 바이알에서 날아 셀룰로오스 아세테이트 플러그 유리 병 상판 아래 2cm가 될 때까지 즉시 테스트 튜브에 셀룰로오스 아세테이트 플러그를 삽입합니다. </li><li> 이하, 엇갈린 1 분 간격으로 집합으로 네 병의 각 행을 처리합니다. </li><li> 시간 0 평가의 경우, 개별적으로 w 각각의 유리 병을 파악i 번째 엄지 손가락과 집게 손가락은 유리 병의 바닥에 파리를 노크 30 초를 기다린 후 죽은 / 움직입니다 파리의 수를 계산하기 위해 테이블​​에 부드럽게 세 번 누릅니다. 시간에 하드 카피 테스트 로그에 0 분 각각의 병에 대한 부동 / 죽은 파리의 수를 기록한다. </li><li> 타이머는 시간 0에서 연속적으로 카운트 업을 시작하고, 즉시 4 튜브의 첫 번째 행 / 세트 바이알 셀룰로스 아세테이트 플러그에 에탄올 1 ㎖을 첨가 시작한다. 그들은 테스트 할 순서에 5 초 간격으로 바이알에 셀룰로오스 아세테이트 플러그에 에탄올 1 ㎖을 추가한다. 에탄올은 셀룰로오스 아세테이트 플러그 골고루 흡수되도록 원을 그리 셀룰로오스 아세테이트 플러그에 에탄올을 추가합니다. 에탄올은 세트의 모든 4 테스트 튜브에 첨가되었을 때, 그것을 밀봉하는 바이 얼 실리콘 플러그를 삽입한다. </li><li> 회 1, 2, 3, 4, 5 분에서, 각각의 튜브 (4) 두 번째, 세 번째, 네 번째, 다섯 번째 및 여섯 번째 세트에 에탄올 1 ㎖을 추가한다. 후 실리콘 마개를 삽입 계속4 튜브의 각 세트에 에탄올을 추가. </li><li> 시간 6 분에서, 엄지와 집게 손가락으로 각 유리 병을 잡고 유리 병의 바닥에 파리를 노크 테이블에 부드럽게 세 번을 눌러, 30 초 대기 후 계산과의 총 수를 기록하여 4 병의 첫 번째 세트를 테스트 진정시켰다 파리. 점수는 (i)는 유리 병의 바닥에 서 있지만, 또는 그들의 날개를 펄럭없이 자신의 허리에 도보 또는 (ⅱ) 거짓말하지 않는 경우 진정으로 날아갑니다. </li><li> 표 2의 스케줄을 사용하여 5 초 간격으로 셋 내의 각각의 바이알 핸들. </li><li> 제 세트에 대해 수행 배 7, 8, 9, 10 및 11 분에서 각각 튜브의, 제 3, 4, 5, 6 세트를 테스트한다. </li><li> 시간이 12 분으로 2.12에 기재된 바와 같이, 튜브 (4)를 다시 상기 제 1 세트의 테스트를 각각 13, 14, 15, 16 및 17 분에서의 바이알 제, 3, 4, 5, 6 세트의 연속 시험. </li><li> 모든 파리가들 때까지 2.12에 설명 된대로 테스트 파리 계속edated. </li><li> 하드 카피 테스트 로그의 각 유리 병에 파리의 총 수를 입력합니다. 파리의 총 수에서 시간 0에서 움직이지 / 죽은 파리를 검열. </li><li> 시간 (x 축) 대 % 활성 파리 (y 축)와 같은 각각의 평가 지점에서의 시간 퍼센트 활성 파리와 플롯 데이터를 계산한다. 다항식 또는 S 자형 곡선이 맞는 3 차에서 진정 작용 시간 (50) 값 (ST50, 50 %의 진정 작용에 시간) 보간 또는 곡선 아래의 면적을 계산하여 에탄올 진정 작용을 정량. </li><li> 디코딩 유리 병에서 데이터를 컴파일하고 통계 분석을 수행 (예를 들면, 일원 분산 분석 페로 니 여러 비교 테스트와) 실험 설계에 대한 적절한. </li></ol>

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The authors have nothing to disclose.

재현성 의미있는 표현형을 정량 간단 분석은 행동의 분석을위한 큰 가치입니다. 여기에 설명 된 작품은 초파리에서 에탄올 진정 감도와 빠른 내성을 측정하기위한 분석의 몇 가지 실용적인 측면을 해결합니다. 이 작품이 아니 긴하지만, 행동 분석 연구에서 피험자에 대한 환경 및 유전 적 배경을 일정하게 유지에 의해 촉진된다. 또한, 비교는 일반적으로 사육과 나란히 테스트 파리의 그룹...

알코올 사용 장애 (AUD)는 전 세계적으로 (1 검토) 거대한 건강 문제입니다. AUD의 개발 구동 메커니즘 복잡하지만, 이러한 장애는 유전 주요 구성 요소를 가지고 (예를 들어, 2). AUD 및 보존 행동 반응의 대형 유전은 더 나은의 분자 기초를 이해하는 방향으로 에탄올과 관련된 행동의 특정 유전자의 참여를 조사하기 위해 유전자 모델 생물을 사용하여 강한 관심을 생성 한 (3,4 검토) 많은 종에서 에탄올 AUD. 초파리, 초파리 melanogaster의는 (3,4 검토) 에탄올과 관련된 행동의 분자 유전 학적 메커니즘을 탐험을위한 선도적 인 모델 생물로 떠오르고있다. 파리의 연구 (5 검토) 에탄올 행동 반응의 여러 신호 전달 경로에 대한 역할을 강조했다. 흥미롭게도, 유전자 및 경로의 일부 행동 응 답에 영향을 미치는 그파리에서 에탄올 ES는 또한 설치류 에탄올과 관련된 행동 및 / 또는 인간 AUD (예 : 6-14)에 연루되어있다. 초파리 모델 시스템에서 사용할 수있는 유전 적 도구의 스위트 룸과 함께 종에서 에탄올과 관련된 행동을, 구동 메커니즘의 보존, 에탄올 행동 반응의 유전학을 조사를 위해 초파리 모델의 유틸리티를 강조한다. 감도 (15, 16)과 인간에 에탄올 (17에서 검토) 허용 오차는 AUD의 발전에 연결되어 있습니다. 에탄올 이러한 행동 반응은 모두 (3,4 검토) 실험실 분석의 다양한 통해 파리에서 모델링 할 수있다. 저자에게 공지 플라이 분석 모두에서 진정 에탄올 중 시간에 따른 에탄올 - 유도 진정 / 운동 실조 또는 경시에 기초하여 복구된다. 에탄올 감도 및 R의 유전학에 우리 그룹에서 이전 기사에서초파리 APID 공차는, 파리의 에탄올 증기 유도 진정 작용에 기초하여 행동 분석은 (18)을 사용 하였다. 이 분석에서 시험이 라이브 성인 전송함으로써 개시 하였다는 셀룰로오스 아세테이트 플러그의 상부 (즉, 비 - 플라이 측)에 에탄올을 첨가, 셀룰로오스 아세테이트 플러그 바이알에서 파리 트래핑 음식 바이알 비우기 마취없이 날아간 및 실리콘 스토퍼 파리, 셀룰로오스 아세테이트 플러그 및 에탄올을 함유하는 바이알을 밀봉 (18 참조,도에서 개략적 S3 참조). 파리의 상이한 그룹을 나타내는 여러 바이알이 분석의 처리량을 증가 병렬 평가 하였다. 유리 병은 익명의 코드를 부여하고 실험자은 진정 작용의 평가에 의도하지 않은 편견을 방지하기 위해 치료 그룹에 눈을 멀게했다. 표준 실험에서, 바이알에 30 초 복구 후 각 바이알에 진정 파리의 수를 계수하고 converte시키고, 6 분 간격으로 부드럽게 탭핑 하였다 파리%의 활성 파리에 D. 파리 내부 에탄올 18 진정제 (이 보고서의 참조 (18) 및 CF도 1a 및도 1b)에서 점진적인 증가를 일으키는 시간 의존적 방식으로 셀룰로스 아세테이트 플러그에서 에탄올 증기를 흡수. 이 분석에서 진정 운영 체제는 산책을하거나 (II) 또는 날개를 펄럭없이 뒤에 거짓말의 부재에서 파리 (I) 서로 정의 하였다. 여기서, 에탄올이 진정 작용 분석법 상세히 설명에는, 사용과 관련된 추가의 최적화 동작이 제공되며, 분석은 비행 진정 감도 음식 보충 옵션 기여를 해결하기 위해 사용된다.

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an inexpensive, scalable behavioral assay for measuring ethanol sedation sensitivity and rapid tolerance in drosophila

알코올 사용 장애 (AUD)는 건강에 심각한 도전이다. AUD로 큰 유전 요소에도 불구하고, 몇 가지 유전자는 분명하게 자신의 원인에 연루되어있다. 초파리, 초파리 melanogaster의는 알코올과 관련된 행동의 기초 분자 유전 메커니즘을 탐구하기위한 강력한 모델이므로 확인하고 AUD에 영향을 미치는 유전자의 기능을 이해하는 데 큰 약속을 보유하고있다. 이러한 유형의 연구를위한 모델 초파리의 사용은 안정적 에탄올 행동 반응을 측정하는 분석법의 가용성에 의존한다. 이 보고서는 파리에서 에탄올 감도와 빠른 내성을 평가하기위한 적절한 분석을 설명합니다. 이 분석에서 측정 에탄올 감도가 초파리 시험 직전 음식없이 수납되어 사용되는 부피의 에탄올 농도, 이전에보고 된 유전자 조작의 다양하고, 또한 시간의 길이에 의해 영향을 받는다. 한편, 에탄올 S이 분석에서 측정 ensitivity 플라이 처리, 파리의 성별, 항생제 또는 살아있는 효모와 성장 매체의 보충의 활력에 영향을받지 않습니다. 정량 에탄올 감도 세 가지 방법은 모두 본질적으로 구별 에탄올 감도 결과에 선도적 인 기술되어있다. 전반적인 단순 결합이 분석의 확장 성 성격, 그리고 상대적으로 낮은 비용으로 업을 설정 초파리 에탄올 감도와 빠른 관용의 크고 작은 규모의 유전 적 분석에 적합하게합니다.

이 에탄올 진정 작용 분석에서 원시 데이터는 에탄올 증기 노출 시간의 함수로서 진정시켰다 파리의 수이다. 원시 데이터는 시간의 함수로서 파리 퍼센트 활성으로 전환된다 (primary 데이터를도 1A, B, D - F). 곡선 적합로부터 기본 데이터에서 에탄올에 진정 감도 보간을 통해, 진정 작용 시간 50 (ST50)로 곡선 (AUC) 아래 진정제 또는 AEA가 날아간의 50 %에 필요한 시간을 정량화 ...

Watch this Scientific Journal Video about An Inexpensive, Scalable Behavioral Assay for Measuring Ethanol Sedation Sensitivity and Rapid Tolerance in Drosophila at JoVE.com

An Inexpensive, Scalable Behavioral Assay for Measuring Ethanol Sedation Sensitivity and Rapid Tolerance in Drosophila

Straightforward assays for measuring ethanol sensitivity and rapid tolerance in Drosophila facilitate the use of this model organism for investigating these important ethanol-related behaviors. Here, a relatively simple, scalable assay for measuring ethanol sensitivity and rapid tolerance in flies is described.

에탄올 진정 감도 및 신속한 공차 측정을위한 저렴하고 확장 가능한 행동 분석<em&gt; 초파리</em

Alcohol use disorder (AUD) is a serious health challenge. Despite a large hereditary component to AUD, few genes have been unambiguously implicated in ...

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JoVE Journal

Neuroscience

An Inexpensive, Scalable Behavioral Assay for Measuring Ethanol Sedation Sensitivity and Rapid Tolerance in Drosophila

10.2K 조회수.  Virginia Commonwealth University. Straightforward assays for measuring ethanol sensitivity and rapid tolerance in Drosophila facilitate the use of this model organism for investigating these important ethanol-related behaviors. Here, a relatively simple, scalable assay for measuring ethanol sensitivity and rapid tolerance in flies is described.

비디오: An Inexpensive, Scalable Behavioral Assay for Measuring Ethanol Sedation Sensitivity and Rapid Tolerance in Drosophila

Combining Behavioral Endocrinology and Experimental Economics: Testosterone and Social Decision Making

Behavioral endocrinological research in humans as well as in animals suggests that testosterone plays a key role in social interactions. Studies in rodents have shown a direct link between testosterone and aggressive behavior1 and folk wisdom adapts these findings to humans, suggesting that testosterone induces antisocial, egoistic or even aggressive behavior2. However, many researchers doubt a direct testosterone-aggression link in humans, arguing instead that testosterone is primarily involved in status-related behavior3,4. As a high status can also be achieved by aggressive and antisocial means it can be difficult to distinguish between anti-social and status seeking behavior.
We therefore set up an experimental environment, in which status can only be achieved by prosocial means. In a double-blind and placebo-controlled experiment, we administered a single sublingual dose of 0.5 mg of testosterone (with a hydroxypropyl-&beta;-cyclodextrin carrier) to 121 women and investigated their social interaction behavior in an economic bargaining paradigm. Real monetary incentives are at stake in this paradigm; every player A receives a certain amount of money and has to make an offer to another player B on how to share the money. If B accepts, she gets what was offered and player A keeps the rest. If B refuses the offer, nobody gets anything. A status seeking player A is expected to avoid being rejected by behaving in a prosocial way, i.e. by making higher offers.
The results show that if expectations about the hormone are controlled for, testosterone administration leads to a significant increase in fair bargaining offers compared to placebo. The role of expectations is reflected in the fact that subjects who report that they believe to have received testosterone make lower offers than those who say they believe that they were treated with a placebo. These findings suggest that the experimental economics approach is sensitive for detecting neurobiological effects as subtle as those achieved by administration of hormones. Moreover, the findings point towards the importance of both psychosocial as well as neuroendocrine factors in determining the influence of testosterone on human social behavior.

The procedure described in this protocol shows that testosterone administration and folk beliefs about testosterone may be associated with directly opposed social behaviors.

행동 내분비학 및 실험 경제학을 결합 : 테스토 스 테론과 사회 의사 결정

Behavioral endocrinological research in humans as well as in animals suggests that testosterone plays a key role in social interactions. Studies in ...

연구

신경과학

A Simple Way to Measure Ethanol Sensitivity in Flies

Low doses of ethanol cause flies to become hyperactive, while high doses are sedating. The sensitivity to ethanol-induced sedation of a given fly strain is correlated with that strain s ethanol preference, and therefore sedation is a highly relevant measure to study the genetics of alcohol responses and drinking. We demonstrate a simple way to expose flies to ethanol and measure its intoxicating effects. The assay we describe can determine acute sensitivity, as well as ethanol tolerance induced by repeat exposure. It does not require a technically involved setup, and can therefore be applied in any laboratory with basic fly culture tools.

A simple assay to measure the sedating effects of ethanol on Drosophila flies, based on the loss of righting reflex, is described.

파리에서 에탄올 민감도를 측정하는 간단한 방법

Low doses of ethanol cause flies to become hyperactive, while high doses are sedating. The sensitivity to ethanol-induced sedation of a given fly strain ...

Microdialysis of Ethanol During Operant Ethanol Self-administration and Ethanol Determination by Gas Chromatography

Operant self-administration methods are commonly used to study the behavioral and pharmacological effects of many drugs of abuse, including ethanol. However, ethanol is typically self-administered orally, rather than intravenously like many other drugs of abuse. The pharmacokinetics of orally administered drugs are more complex than intravenously administered drugs. Because understanding the relationship between the pharmacological and behavioral effects of ethanol requires knowledge of the time course of ethanol reaching the brain during and after drinking, we use in vivo microdialysis and gas chromatography with flame ionization detection to monitor brain dialysate ethanol concentrations over time. 
Combined microdialysis-behavioral experiments involve the use of several techniques. In this article, stereotaxic surgery, behavioral training and microdialysis, which can be adapted to test a multitude of self-administration and neurochemical centered hypotheses, are included only to illustrate how they relate to the subsequent phases of sample collection and dialysate ethanol analysis. Dialysate ethanol concentration analysis via gas chromatography with flame-ionization detection, which is specific to ethanol studies, is described in detail. Data produced by these methods reveal the pattern of ethanol reaching the brain during the self-administration procedure, and when paired with neurochemical analysis of the same dialysate samples, allows conclusions to be made regarding the pharmacological and behavioral effects of ethanol.

A method to determine the time course of ethanol concentration in the brains of rats during operant ethanol self-administration is described. Gas chromatography with flame ionization detection is used to quantify ethanol in the dialysate samples, because it has the sensitivity required for the small volumes that are generated.

가스 크로마토 그래피에 의한 operant 에탄올 자기 관리 및 에탄올 결정시 에탄올의 Microdialysis

Operant self-administration methods are commonly used to study the behavioral and pharmacological effects of many drugs of abuse, including ethanol.  ...

Measuring Changes in Tactile Sensitivity in the Hind Paw of Mice Using an Electronic von Frey Apparatus

Measuring inflammation-induced changes in thresholds of hind paw withdrawal from mechanical pressure is a useful technique to assess changes in pain perception in rodents. Withdrawal thresholds can be measured first at baseline and then following drug, venom, injury, allergen,&#160;or otherwise evoked inflammation by applying an accurate force on very specific areas of the skin. An electronic von Frey apparatus allows precise assessment of mouse hind paw withdrawal thresholds that are not limited by the available filament sizes in contrast to classical von Frey measurements. The ease and rapidity of measurements allow for incorporation of assessment of tactile sensitivity outcomes in diverse models of rapid-onset inflammatory and neuropathic pain as multiple measurements can be taken within a short time period. Experimental measurements for individual rodent subjects can be internally controlled against individual baseline responses and exclusion criteria easily established to standardize baseline responses within and across experimental groups. Thus, measurements using an electronic von Frey apparatus represent a useful modification of the well-established classical von Frey filament-based assays for rodent mechanical allodynia that may also be applied to other nonhuman mammalian models.

Electronic von Frey measurements are an effective and improved alternative to classical von Frey filaments, as they allow rapid and precise measurement of changes in rodent mechanical withdrawal thresholds to continuously applied pressure stimuli before and after an inflammatory event.

Measuring inflammation-induced changes in thresholds of hind paw withdrawal from mechanical pressure is a useful technique to assess changes in pain ...

A Simple and Inexpensive Method for Determining Cold Sensitivity and Adaptation in Mice

Cold hypersensitivity is a serious clinical problem, affecting a broad subset of patients and causing significant decreases in quality of life. The cold plantar assay allows the objective and inexpensive assessment of cold sensitivity in mice, and can quantify both analgesia and hypersensitivity. Mice are acclimated on a glass plate, and a compressed dry ice pellet is held against the glass surface underneath the hindpaw. The latency to withdrawal from the cooling glass is used as a measure of cold sensitivity.
Cold sensation is also important for survival in regions with seasonal temperature shifts, and in order to maintain sensitivity animals must be able to adjust their thermal response thresholds to match the ambient temperature. The Cold Plantar Assay (CPA) also allows the study of adaptation to changes in ambient temperature by testing the cold sensitivity of mice at temperatures ranging from 30 &#176;C to 5 &#176;C. Mice are acclimated as described above, but the glass plate is cooled to the desired starting temperature using aluminum boxes (or aluminum foil packets) filled with hot water, wet ice, or dry ice. The temperature of the plate is measured at the center using a filament T-type thermocouple probe. Once the plate has reached the desired starting temperature, the animals are tested as described above.
This assay allows testing of mice at temperatures ranging from innocuous to noxious. The CPA yields unambiguous and consistent behavioral responses in uninjured mice and can be used to quantify both hypersensitivity and analgesia. This protocol describes how to use the CPA to measure cold hypersensitivity, analgesia, and adaptation in mice.

The Cold Plantar Assay (CPA) measures cold responsiveness between 30 &#176;C and 5 &#176;C, and can also measure cold adaptation. This protocol describes how to use the CPA to measure cold hypersensitivity, analgesia, and adaptation in mice.

마우스에 콜드 민감도 및 적응을 결정하기위한 간단하고 저렴한 방법

Cold hypersensitivity is a serious clinical problem, affecting a broad subset of patients and causing significant decreases in quality of life.  The cold ...

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 ...

A Behavioral Assay for Mechanosensation of MARCM-based Clones in Drosophila melanogaster

Because of the structural and functional homology to the hair cells of the mammalian inner ear, the neurons that innervate the Drosophila external sense organs provide an excellent model system for the study of mechanosensation. This protocol describes a simple touch behavior in fruit flies which can be used to identify mutations that interfere with mechanosensation. The tactile stimulation of a macrochaete bristle on the thorax of flies elicits a grooming reflex from either the first or third leg. Mutations that interfere with mechanotransduction (such as NOMPC), or with other aspects of the reflex arc, can inhibit the grooming response. A traditional screen of adult behaviors would have missed mutants that have essential roles during development. Instead, this protocol combines the touch screen with mosaic analysis with a repressible cell marker (MARCM) to allow for only limited regions of homozygous mutant cells to be generated and marked by the expression of green fluorescent protein (GFP). By testing MARCM clones for abnormal behavioral responses, it is possible to screen a collection of lethal p-element mutations to search for new genes involved in mechanosensation that would have been missed by more traditional methods.

A Behavioral Assay for Mechanosensation of MARCM-based Clones in Drosophila melanogaster

In order to identify novel mutations affecting mechanosensation, we designed an assay that measures the behavioral response to tactile stimulation of fly bristles in mutant clones generated by the MARCM method. The combination of techniques allows for the identification of mechanosensitive mutations that would otherwise be lethal.

의 Mechanosensation에 대한 행동 분석 클론의 MARCM은 기반<em&gt; 초파리</em

Because of the structural and functional homology to the hair cells of the mammalian inner ear, the neurons that innervate the Drosophila external sense ...

Electrophysiological Method for Recording Intracellular Voltage Responses of Drosophila Photoreceptors and Interneurons to Light Stimuli In Vivo

Voltage responses of insect photoreceptors and visual interneurons can be accurately recorded with conventional sharp microelectrodes. The method described here enables the investigator to measure long-lasting (from minutes to hours) high-quality intracellular responses from single Drosophila R1-R6 photoreceptors and Large Monopolar Cells (LMCs) to light stimuli. Because the recording system has low noise, it can be used to study variability among individual cells in the fly eye, and how their outputs reflect the physical properties of the visual environment. We outline all key steps in performing this technique. The basic steps in constructing an appropriate electrophysiology set-up for recording, such as design and selection of the experimental equipment are described. We also explain how to prepare for recording by making appropriate (sharp) recording and (blunt) reference electrodes. Details are given on how to fix an intact fly in a bespoke fly-holder, prepare a small window in its eye and insert a recording electrode through this hole with minimal damage. We explain how to localize the center of a cell's receptive field, dark- or light-adapt the studied cell, and to record its voltage responses to dynamic light stimuli. Finally, we describe the criteria for stable normal recordings, show characteristic high-quality voltage responses of individual cells to different light stimuli, and briefly define how to quantify their signaling performance. Many aspects of the method are technically challenging and require practice and patience to master. But once learned and optimized for the investigator's experimental objectives, it grants outstanding in vivo neurophysiological data.

Electrophysiological Method for Recording Intracellular Voltage Responses of Drosophila Photoreceptors and Interneurons to Light Stimuli In Vivo

Sharp microelectrodes enable accurate electrophysiological characterization of photoreceptor and visual interneuron output in living Drosophila. Here we show how to use this method to record high-quality voltage responses of individual cells to controlled light stimulation. This method is ideal for studying neural information processing in insect compound eyes.

의 기록 세포 내 전압 응답에 대한 전기 생리 방법<em&gt; 초파리</em&gt; 빛 자극에 광수와의 interneurons<em&gt; 생체</em

Voltage responses of insect photoreceptors and visual interneurons can be accurately recorded with conventional sharp microelectrodes. The method ...

Measuring and Altering Mating Drive in Male Drosophila melanogaster

Despite decades of investigation, the neuronal and molecular bases of motivational states remain mysterious. We have recently developed a novel, reductionist, and scalable system for in-depth investigation of motivation using the mating drive of male Drosophila&#160;melanogaster (Drosophila), the methods for which we detail here. The behavioral paradigm centers on the finding that male mating drive decreases alongside fertility over the course of repeated copulations and recovers over ~3 d. In this system, the powerful neurogenetic tools available in the fly converge with the genetic accessibility and putative wiring diagram available for sexual behavior. This convergence allows rapid isolation and interrogation of small neuronal populations with specific motivational functions. Here we detail the design and execution of the satiety assay that is used to measure and alter courtship motivation in the male fly. Using this assay, we also demonstrate that low male mating drive can be overcome by stimulating dopaminergic neurons. The satiety assay is simple, affordable, and robust to influences of genetic background. We expect the satiety assay to generate many new insights into the neurobiology of motivational states.

Measuring and Altering Mating Drive in Male Drosophila melanogaster

This article describes a behavioral assay that uses male mating drive in Drosophila melanogaster to study motivation. Using this method, researchers can utilize advanced fly neurogenetic techniques to uncover the genetic, molecular, and cellular mechanisms that underlie this motivation.

측정과 남성의 변경 결합 드라이브<em&gt; 노랑 초파리</em

Despite decades of investigation, the neuronal and molecular bases of motivational states remain mysterious. We have recently developed a novel, ...

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 ...