이 연구는 NIH / NIAAA (AA11852과 AA007471)에서 보조금에 의해 지원되었다.

<ol>
	<li>Geiger, B. M., Frank, L. E., Caldera-Siu, A. D., Pothos, E. N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Survivable+stereotaxic+surgery+in+rodents.">Survivable stereotaxic surgery in rodents.</a> J. Vis. Exp. (20), e880 (2008).</li><li>Howard, E. C., Schier, C. J., Wetzel, J. S., Gonzales, R. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+dopamine+response+in+the+nucleus+accumbens+core-shell+border+differs+from+that+in+the+core+and+shell+during+operant+ethanol+self-administration.">The dopamine response in the nucleus accumbens core-shell border differs from that in the core and shell during operant ethanol self-administration.</a> Alcohol Clin. Exp. Res. 33, 1355-1365 (2009).</li><li>Doyon, W. M., York, J. L., Diaz, L. M., Samson, H. H., Czachowski, C. L., Gonzales, R. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Dopamine+activity+in+the+nucleus+accumbens+during+consummatory+phases+of+oral+ethanol+self-administration.">Dopamine activity in the nucleus accumbens during consummatory phases of oral ethanol self-administration.</a> Alcohol Clin. Exp. Res. 27, 1573-1582 (2003).</li><li>Pettit, H. O., Justice, J. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effect+of+dose+on+cocaine+self-administration+behavior+and+dopamine+levels+in+the+nucleus+accumbens.">Effect of dose on cocaine self-administration behavior and dopamine levels in the nucleus accumbens.</a> Brain Res. 539, 94-102 (1991).</li><li>Paxinos, G., Kus, L., Ashwell, K. W. S., Watson, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Chemoarchitectonic Atlas of The Rat Forebrain. , (1998).</li><li>Paxinos, G., Watson, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> The Rat Brain in Stereotaxic Coordinates. , (1986).</li><li>Skoog, D. A., Holler, F. J., Nieman, T. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Principles of Instrumental Analysis. , (1998).</li><li>Doyon, W. M., Anders, S. K., Ramachandra, V. S., Czachowski, C. L., Gonzales, R. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effect+of+operant+self-administration+of+10%+ethanol+plus+10%+sucrose+on+dopamine+and+ethanol+concentrations+in+the+nucleus+accumbens.">Effect of operant self-administration of 10% ethanol plus 10% sucrose on dopamine and ethanol concentrations in the nucleus accumbens.</a> J Neurochem. 93, 1469-1481 (2005).</li><li>Carrillo, J., Gonzales, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+single+exposure+to+voluntary+ethanol+self-administration+produces+adaptations+in+ethanol+consumption+and+accumbal+dopamine+signaling.">A single exposure to voluntary ethanol self-administration produces adaptations in ethanol consumption and accumbal dopamine signaling.</a> Alcohol. 45, 559-5566 (2011).</li><li>Howard, E. C., Schier, C. J., Wetzel, J. S., Duvauchelle, C. L., Gonzales, R. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+shell+of+the+nucleus+accumbens+has+a+higher+dopamine+response+compared+with+the+core+after+non-contingent+intravenous+ethanol+administration.">The shell of the nucleus accumbens has a higher dopamine response compared with the core after non-contingent intravenous ethanol administration.</a> Neuroscience. 154, 1042-1053 (2008).</li><li>Robinson, D. L., Lara, J. A., Brunner, L. J., Gonzales, R. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Quantification+of+Ethanol+Concentrations+in+the+Extracellular+Fluid+of+the+Rat+Brain.">Quantification of Ethanol Concentrations in the Extracellular Fluid of the Rat Brain.</a> J. Neurochem. 75, 1685-1693 (2000).</li><li>Gonzales, R. A., McNabb, J., Yim, H. J., Ripley, T., Bungay, P. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Quantitative+Microdialysis+of+Ethanol+in+Rat+Striatum.">Quantitative Microdialysis of Ethanol in Rat Striatum.</a> Alcohol Clin. Exp. Res. 22, 858-867 (1998).</li></ol>

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응용 프로그램 및 제한 약물 자기 관리는 모델 약물 중독과 동물에 사용됩니다. 이 방식으로 모델 아르 학대의 많은 약물은 약물이 중앙 구획에 직접 전달되는, intravenously 관리 할 수​​ 있습니다. 이 자체 관리 세션 이상 복용의 가까이에 모니터링 할 수 있습니다. 에탄올은 구두 보통 자기 관리이기 때문에, 그것은 흡수와 신진 대사의 개별 차이로 인해 마약 수준을 모니터링하기가 훨씬 어렵습니다. 관심의 두뇌 지역에서 샘플로 microdialysis를 사용하여, 우리는이 지역에 도달 에탄올의 패턴을 모니터 할 수 없지만, 우리는 또한 동시에 각 자체 관리 단계에서 시간이 지남에 같은 지역에서 신경 전달 물질의 변화를 모니터링 할 수 있습니다. 뇌 Neurochemical 변경 및 약물 유발 반응은 약물 남용과 의존과 연결되어, 따라서 동시에 MEA 할 수있는 능력을특정 자기 관리 단계 동안 확인 neurochemical 및 약물 농도는 매우 강력하고 독특한 도구를 제공합니다. 행동과 analytes의 dialysate 농도를 상호 연관에 대해 유의해야 할 하나의 문제는 microdialysis의 배관의 물리적 특성입니다. 특히,이 수집 관에 microdialysis 프로브의 루멘에서 전달되는 유체에 대해 걸리는 시간은 매우 중요하며, 짧은 시간이 더 있습니다. 시간이 90 초 정도입니다 수 있도록 우리가 실험실에서, 우리는 프로브를 구성합니다. 이러한 조건에 사용, 우리는 dialysate 에탄올 농도 증가로 음료 및 사후 음료 기간에 걸쳐 감소 에탄올 소비의 개시에서 accumbal 도파민 증가가 나타났습니다. 2,3,8,9 이러한 실험 , 약리 연구의 데이터와 결합 할 때, 우리 채널에서 에탄올과 자기 관리 관련 환경 단서의 엄격 약리 효과를 분석 할 수있다신경 전달 물질 농도에 anges. 그것은 결합 동작 - microdialysis 기술의 특정 응용 프로그램이 저희 연구실의 현재 연구 관심 분야에 적합한 것을 주목해야한다. 그것은 우리가 동기 자기 관리 행동에 이러한 조치를 관련 수 있도록, 같은 지역에서 신경 전달 물질의 변화 패턴에 비해 뇌에 도달 에탄올의 시간 패턴을 평가하기 위해 설계되었습니다. 파생 dialysate의 에탄올 농도는 생체 프로브 복구에 대한 수정 및 뇌 에탄올 조직 농도의 일부분 일 뿐인 걸까되지 않습니다. 에탄올의 정량 microdialysis가 필요한 경우 프로브에 세포 우주에서 diffuses하는 에탄올의 추출 분율은 실험적으로 결정해야합니다. 방법과 자세한 설명을 위해 저희 연구실에서 이전 출판물을 참조하십시오. 10,11,12 이 프로토콜 가스 채널의 사용을 보여줍니다 있지만microdialysis 샘플의 천정 공간에서 에탄올의 견고한 위상 microextraction와 함께 romatography는 microdialysis 샘플의 에탄올 내용의 분석을위한 다른 방법을 사용할 수 있습니다. 그러나, 다른 방법은 몇 가지 단점으로 고생 할 수 있습니다. 예를 들어, 덜 민감 분석 방법은 여기에 도시 된 5-7 분보다 샘플링 시간 더 필요로 큰 샘플 볼륨이 필요할 수 있습니다. 여기에 설명 시스템의 유형은 유리 병의 천정 공간에 배치 섬유에 흡수함으로써 밀봉 된 샘플 병에 증기 단계에서 에탄올을 집중 견고한 위상 microextraction을 사용합니다. 이것은 일반적으로 증기의 50-100 microliters을 주입 할 수 있습니다 직접 헤드 스페이스 샘플링에 비해 검출 한계를 향상시킵니다. 헤드 스페이스 방법의 또 다른 주요 장점은 분석을 위해 주입 된 샘플은 매우 깨끗하고 소금은 무료입니다 것입니다. 액체 microdialysis 샘플의 직접 분사도 높은 sensit와 함께 사용할 수 있습니다ivity하지만,이 주입 된 소금을 청소하는 데 필요한 정기적 인 유지 관리 작업으로 인해 시간 다운 더 많은 악기를 필요합니다. 문제 해결 및 기타 사항 <ol><li> 실험이 시작하기 전에 microdialysis 세트 업이 제대로 작동하는지 확인하고 문제를 고장 쏘 자신에게 충분한 시간을 제공합니다. 우리는 당신이 밖으로 전환 할 준비, ACSF와 perfused, 설정 추가를하는 것이 좋습니다 가장 operant 세션을 매일 특정 시간에 발생하는 경향이로, 자신에게 시간을 절약 할 설정 오작동. </li><li> microdialysis 프로브는 프로브의 뇌를 침투로 조직의 전단에 의해 생성 된 조직의 손상을 최소화하기 위해 약 5 분에 걸쳐 삽입되어 있는지 확인하십시오. </li><li> operant 챔버에 쥐를 전송하는 경우는 두 번째 사람이 operant 프로그램을 초기화 할 수 있도록 전송 라인 (microdialysis 설정의 일부) 얽힌되고하지 않도록 지원하고있는 것이 유용합니다시간에 iated. </li><li> 당신은 적절한 온도에 도달하는 시간의 autosampler 물 목욕을 충분히 제공해야합니다. Google 시스템은 일반적으로 두 시간 밖에 50 ° C.에 도달해야합니다 또한 샘플이 완전히 샘플을 분석하기 전에 기화 있으니 안심하십시오. 우리는 일반적으로 시각이 샘플 분석을 시작하기 전에 발생했음을 확인합니다. </li><li> 표준 좋은 분석 화학 관행을 준수해야합니다. 여기에는 있지만, 개인, 장비 및 절차의 검증에 제한되지 않습니다. 간단한에서, 우리는 다음과 같은 가이드 라인을 고용 : 개인 사용자는 여러 일에 걸쳐 표준 농도 및 선형 표준 곡선 (R2 ≥ 0.99)을 재현 피크 높이 값을 생성 할 수있는 능력을 입증해야합니다. GC-FID 시스템이 제대로 작동하는지 확인하기 위해, 표준은 dialysate 샘플을 주입하기 전에 분석 할 항상 있어야합니다. 생성 된 표준 곡선 R2 ≥ 0.99이 있어야합니다. </li><li> 우리가 사용하는 장비의 대부분은2010 년 애질런트 테크놀로지스 인수 한 Varian 주식회사를 통해 구입했습니다. 현재 Bruker 과학 인스트루먼트는 Varian의 실험실 가스 크로마토 그래피 기기 사업을 획득하였습니다. 향후 구매를 들어, 두 회사를 참조하십시오. </li><li> 현재 프로토콜은 쥐와 operant 동작을 보여 주지만 뷰어는 행동의 더 많은 혼란이 작은 마우스 모델에서 발생할 가능성이 있다는 것을 알고 있어야한다. 인식해야 할 또 다른 문제는 중간 전두엽 피질 또는 핵 accumbens는 여기에 표시된 것보다 더 큰 정도 operant 동작을 방해 할 수 있습니다 이외의 다른 뇌 영역에 microdialysis 프로브의 게재 위치가 있습니다. 그것은 밀접하게 프로브 배치로 인한 손상이 행동의 심각한 변화를 초래 여부를 결정하는 여러 행동 매개 변수를 검사하는 것이 중요합니다. </li></ol>

<table border="1"><tbody><tr><td> 시약의 이름 </td><td> 회사 </td><td> 카탈로그 번호 </td></tr><tr><td> 95 % 에탄올 </td><td> AAPER 알코올 및 화학 Shelbyville, KY </td><td> E190, 111,000,190 </td></tr><tr><td> 울트라 순수 자당 </td><td> MP Biomedicals, LLC, 솔론, OH </td><td> 821721 </td></tr><tr><td> 울트라 고순도 헬륨 </td><td> 공기 가스 </td><td> HE UHP300 </td></tr><tr><td> 공기 </td><td> 공기 가스 </td><td> AIZ300 </td></tr><tr><td> 수소 </td><td> 공기 가스 </td><td> AIZ300 </td></tr><tr><td> GC 2 ML 병 </td><td> 애질런트 </td><td> # 8010-0015 </td></tr><tr><td> PTFE / 실리콘 septa와 GC의 유리 병 캡 </td><td> 애질런트 </td><td> # 8010-0084 </td></tr></tbody></table> 표 1. 특정 시약

microdialysis of ethanol during operant ethanol self-administration and ethanol determination by gas chromatography

Operant 자기 관리 방법은 일반적으로 에탄올 등의 학대의 많은 약물의 행동 및 약리 효과를 연구하는 데 사용됩니다. 그러나 에탄올은 일반적으로 오히려 남용의 많은 다른 약물과 같은 intravenously보다 구두 자기 관리입니다. 구두 관리 약물의 pharmacokinetics은 intravenously 관리 마약보다 더 복잡합니다. 에탄올의 약리 및 행동 효과 사이의 관계를 이해하는 것은 음주 중이나 후에 뇌에 도달 에탄올의 시간 코스에 대한 지식이 필요하기 때문에, 우리는 시간이 지남에 따라 뇌 dialysate의 에탄올 농도를 모니터링하는 불꽃 이온화 감지 생체의 microdialysis 및 가스 크로마토 그래피에 사용합니다. 결합 microdialysis - 행동 실험은 여러 기술의 사용을 포함한다. 의 자기 관리 및 neu 수많은 테스트하도록 구성 할 수있다이 문서, stereotaxic 수술, 행동 교육 및 microdialysis,rochemical 중심 가설은 단지 그들이 샘플 수집 및 dialysate의 에탄올 분석의 후속 단계에 관한 방법을 설명하기위한 포함되어 있습니다. 에탄올 연구에 따라 달라집니다 불꽃 - 이온화 감지,와 가스 크로마토 그래피를 통해 에탄올 농도 분석을 Dialysate이 자세히 설명되어 있습니다. 이러한 방법에 의해 생산 데이터는 자체 관리 절차를 수행하는 동안 뇌에 도달 에탄올의 패턴을 보여, 같은 dialysate 샘플 neurochemical 분석과 결합하여, 결론은 에탄올의 약리 및 행동 효과에 관한 한 할 수 있습니다.

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

operant 에탄올 동안 쥐의 뇌에 에탄올 농도의 시간 코스를 결정하는 방법은 자기 관리가 설명되어 있습니다. 가 생성되는 작은 볼륨에 필요한 감도를 가지고 있기 때문에 불꽃 이온화 검출과 가스 크로마토 그래피는, dialysate 샘플에 에탄올을 수량화하는 데 사용됩니다.

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.  ...

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17.4K 조회수.  The University of Texas at Austin. operant 에탄올 동안 쥐의 뇌에 에탄올 농도의 시간 코스를 결정하는 방법은 자기 관리가 설명되어 있습니다. 가 생성되는 작은 볼륨에 필요한 감도를 가지고 있기 때문에 불꽃 이온화 검출과 가스 크로마토 그래피는, dialysate 샘플에 에탄올을 수량화하는 데 사용됩니다.

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

Assessment of Ultrasonic Vocalizations During Drug Self-administration in Rats

Drug self-administration procedures are commonly used to study behavioral and neurochemical changes associated with human drug abuse, addiction and relapse. Various types of behavioral activity are commonly utilized as measures of drug motivation in animals. However, a crucial component of drug abuse relapse in abstinent cocaine users is "drug craving", which is difficult to model in animals, as it often occurs in the absence of overt behaviors. Yet, it is possible that a class of ultrasonic vocalizations (USVs) in rats may be a useful marker for affective responses to drug administration, drug anticipation and even drug craving. Rats vocalize in ultrasonic frequencies that serve as a communicatory function and express subjective emotional states. Several studies have shown that different call frequency ranges are associated with negative and positive emotional states. For instance, high frequency calls ("50-kHz") are associated with positive affect, whereas low frequency calls ("22-kHz") represent a negative emotional state. This article describes a procedure to assess rat USVs associated with daily cocaine self-administration. For this procedure, we utilized standard single-lever operant chambers housed within sound-attenuating boxes for cocaine self-administration sessions and utilized ultrasonic microphones, multi-channel recording hardware and specialized software programs to detect and analyze USVs. USVs measurements reflect emotionality of rats before, during and after drug availability and can be correlated with commonly assessed drug self-administration behavioral data such lever responses, inter-response intervals and locomotor activity. Since USVs can be assessed during intervals prior to drug availability (e.g., anticipatory USVs) and during drug extinction trials, changes in affect associated with drug anticipation and drug abstinence can also be determined. In addition, determining USV changes over the course of short- and long-term drug exposure can provide a more detailed interpretation of drug exposure effects on affective functioning.

Drug self-administration and ultrasonic vocalizations (USV) are used as behavioral assessments in animal research, but rarely in combination. The purpose of this article is to describe the advantages of recording USVs during drug self-administration procedures to assess affective responses to drug experience.

쥐에서 의약품 자체 관리하는 동안 초음파 Vocalizations 평가

Drug self-administration procedures are commonly used to study behavioral and neurochemical changes associated with human drug abuse, addiction and ...

연구

신경과학

Operant Sensation Seeking in the Mouse

Operant methods are powerful behavioral tools for the study of motivated behavior. These 'self-administration' methods have been used extensively in drug addiction research due to their high construct validity. Operant studies provide researchers a tool for preclinical investigation of several aspects of the addiction process. For example, mechanisms of acute reinforcement (both drug and non-drug) can be tested using pharmacological or genetic tools to determine the ability of a molecular target to influence self-administration behavior1-6. Additionally, drug or food seeking behaviors can be studied in the absence of the primary reinforcer, and the ability of pharmacological compounds to disrupt this process is a preclinical model for discovery of molecular targets and compounds that may be useful for the treatment of addiction3,7-9. One problem with performing intravenous drug self-administration studies in the mouse is the technical difficulty of maintaining catheter patency. Attrition rates in these experiments are high and can reach 40% or higher10-15. Another general problem with drug self-administration is discerning which pharmacologically-induced effects of the reinforcer produce specific behaviors. For example, measurement of the reinforcing and neurological effects of psychostimulants can be confounded by their psychomotor effects. Operant methods using food reinforcement can avoid these pitfalls, although their utility in studying drug addiction is limited by the fact that some manipulations that alter drug self-administration have a minimal impact on food self-administration. For example, mesolimbic dopamine lesion or knockout of the D1 dopamine receptor reduce cocaine self-administration without having a significant impact on food self-administration 12,16. 
Sensory stimuli have been described for their ability to support operant responding as primary reinforcers (i.e. not conditioned reinforcers)17-22. Auditory and visual stimuli are self-administered by several species18,21,23, although surprisingly little is known about the neural mechanisms underlying this reinforcement. The operant sensation seeking (OSS) model is a robust model for obtaining sensory self-administration in the mouse, allowing the study of neural mechanisms important in sensory reinforcement24. An additional advantage of OSS is the ability to screen mutant mice for differences in operant behavior that may be relevant to addiction. We have reported that dopamine D1 receptor knockout mice, previously shown to be deficient in psychostimulant self-administration, also fail to acquire OSS24. This is a unique finding in that these mice are capable of learning an operant task when food is used as a reinforcer. While operant studies using food reinforcement can be useful in the study of general motivated behavior and the mechanisms underlying food reinforcement, as mentioned above, these studies are limited in their application to studying molecular mechanisms of drug addiction. Thus, there may be similar neural substrates mediating sensory and psychostimulant reinforcement that are distinct from food reinforcement, which would make OSS a particularly attractive model for the study of drug addiction processes. The degree of overlap between other molecular targets of OSS and drug reinforcers is unclear, but is a topic that we are currently pursuing. While some aspects of addiction such as resistance to extinction may be observed with OSS, we have found that escalation 25 is not observed in this model24. Interestingly, escalation of intake and some other aspects of addiction are observed with self-administration of sucrose26. Thus, when non-drug operant procedures are desired to study addiction-related processes, food or sensory reinforcers can be chosen to best fit the particular question being asked. 
In conclusion, both food self-administration and OSS in the mouse have the advantage of not requiring an intravenous catheter, which allows a higher throughput means to study the effects of pharmacological or genetic manipulation of neural targets involved in motivation. While operant testing using food as a reinforcer is particularly useful in the study of the regulation of food intake, OSS is particularly apt for studying reinforcement mechanisms of sensory stimuli and may have broad applicability to novelty seeking and addiction.

In this protocol we describe a method of operant learning using sensory stimuli as a reinforcer in the mouse. It requires no prior training or food restriction, and it allows the study of motivated behavior without the use of a pharmacological or natural reinforcer such as food.

마우스의 추구 Operant 감각

Operant methods are powerful behavioral tools for the study of motivated behavior.  These 'self-administration' methods have been used extensively in drug ...

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 ...

Methods for Intravenous Self Administration in a Mouse Model

Animal models have been developed to study the reinforcing effects of drugs, including the intravenous self-administration (IVSA) paradigm. The advantages of using an IVSA paradigm to study the reinforcing properties of drugs of abuse such as cocaine include the fact that the drug is self-administered instead of experimenter-administered, the schedule of reinforcement can be altered, and accurate measurement of the quantities of drug consumed as well as the timing and pattern of IV injections can be obtained. Furthermore, the intravenous route of administration avoids potential confounds related to first pass metabolism or taste, and produces rapid increases in blood and brain drug levels. As outlined in this video, intravenous self-administration can be obtained without prior food restriction or prior drug training following careful catheter placement during surgery and meticulous daily catheter flushing and maintenance. Experimental procedures outlined in this paper include a description of animal housing and acclimation methods, operant training using sweetened milk solutions, and catheter implantation surgery.

The intravenous self-administration (IVSA) paradigm is considered to be the gold standard in examining the reinforcing properties of drugs of abuse in rodents. This manuscript outlines the experimental procedures and surgical techniques necessary to obtain reliable IVSA data. In particular, meticulous catheter implantation and maintenance are highlighted.

마우스 모델에서 정맥 주사 자기 관리를위한 방법

Animal models have been developed to study the reinforcing effects of drugs, including the intravenous self-administration (IVSA) paradigm. The advantages ...

Identification of Olfactory Volatiles using Gas Chromatography-Multi-unit Recordings (GCMR) in the Insect Antennal Lobe

All organisms inhabit a world full of sensory stimuli that determine their behavioral and physiological response to their environment. Olfaction is especially important in insects, which use their olfactory systems to respond to, and discriminate amongst, complex odor stimuli. These odors elicit behaviors that mediate processes such as reproduction and habitat selection1-3. Additionally, chemical sensing by insects mediates behaviors that are highly significant for agriculture and human health, including pollination4-6, herbivory of food crops7, and transmission of disease8,9. Identification of olfactory signals and their role in insect behavior is thus important for understanding both ecological processes and human food resources and well-being.
	To date, the identification of volatiles that drive insect behavior has been difficult and often tedious. Current techniques include gas chromatography-coupled electroantennogram recording (GC-EAG), and gas chromatography-coupled single sensillum recordings (GC-SSR)10-12. These techniques proved to be vital in the identification of bioactive compounds. We have developed a method that uses gas chromatography coupled to multi-channel electrophysiological recordings (termed 'GCMR') from neurons in the antennal lobe (AL; the insect's primary olfactory center)13,14. This state-of-the-art technique allows us to probe how odor information is represented in the insect brain. Moreover, because neural responses to odors at this level of olfactory processing are highly sensitive owing to the degree of convergence of the antenna's receptor neurons into AL neurons, AL recordings will allow the detection of active constituents of natural odors efficiently and with high sensitivity. Here we describe GCMR and give an example of its use.
	Several general steps are involved in the detection of bioactive volatiles and insect response. Volatiles first need to be collected from sources of interest (in this example we use flowers from the genus Mimulus (Phyrmaceae)) and characterized as needed using standard GC-MS techniques14-16. Insects are prepared for study using minimal dissection, after which a recording electrode is inserted into the antennal lobe and multi-channel neural recording begins. Post-processing of the neural data then reveals which particular odorants cause significant neural responses by the insect nervous system.
	Although the example we present here is specific to pollination studies, GCMR can be expanded to a wide range of study organisms and volatile sources. For instance, this method can be used in the identification of odorants attracting or repelling vector insects and crop pests. Moreover, GCMR can also be used to identify attractants for beneficial insects, such as pollinators. The technique may be expanded to non-insect subjects as well.

Identification of Olfactory Volatiles using Gas Chromatography-Multi-unit Recordings GCMR in the Insect Antennal Lobe

Olfactory cues mediate many different behaviors in insects, and are often complex mixtures comprised of tens to hundreds of volatile compounds. Using gas chromatography with multi-channel recording in the insect antennal lobe, we describe a method for the identification of bioactive compounds.

곤충 Antennal 엽에 가스 크로마토 그래피 - 멀티 유닛 레코딩 (GCMR)를 사용하여 강한 휘발성 물질의 식별

All  organisms inhabit a world full of sensory stimuli that determine their  behavioral and physiological response to their environment. Olfaction is ...

Hippocampal Insulin Microinjection and In vivo Microdialysis During Spatial Memory Testing

Glucose metabolism is a useful marker for local neural activity, forming the basis of methods such as 2-deoxyglucose and functional magnetic resonance imaging. However, use of such methods in animal models requires anesthesia and hence both alters the brain state and prevents behavioral measures. An alternative method is the use of in vivo microdialysis to take continuous measurement of brain extracellular fluid concentrations of glucose, lactate, and related metabolites in awake, unrestrained animals. This technique is especially useful when combined with tasks designed to rely on specific brain regions and/or acute pharmacological manipulation; for example, hippocampal measurements during a spatial working memory task (spontaneous alternation) show a dip in extracellular glucose and rise in lactate that are suggestive of enhanced glycolysis1-3,4-5, and intrahippocampal insulin administration both improves memory and increases hippocampal glycolysis6. Substances such as insulin can be delivered to the hippocampus via the same microdialysis probe used to measure metabolites. The use of spontaneous alternation as a measure of hippocampal function is designed to avoid any confound from stressful motivators (e.g. footshock), restraint, or rewards (e.g. food), all of which can alter both task performance and metabolism; this task also provides a measure of motor activity that permits control for nonspecific effects of treatment. Combined, these methods permit direct measurement of the neurochemical and metabolic variables regulating behavior.

Hippocampal Insulin Microinjection and In vivo Microdialysis During Spatial Memory Testing

Modulation of hippocampally-dependent spatial working memory by direct intrahippocampal microinjection, accompanied and followed by in vivo microdialysis for metabolites in conscious, behaving animals.

Hippocampal 인슐린 Microinjection 및<em&gt; 생체에</em공간 메모리 테스트 기간 동안&gt; Microdialysis

Glucose metabolism is a useful marker for local neural activity, forming the basis of methods such as 2-deoxyglucose and functional magnetic resonance ...

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

Alcohol use disorder (AUD) is a serious health challenge. Despite a large hereditary component to AUD, few genes have been unambiguously implicated in their etiology. The fruit fly, Drosophila melanogaster, is a powerful model for exploring molecular-genetic mechanisms underlying alcohol-related behaviors and therefore holds great promise for identifying and understanding the function of genes that influence AUD. The use of the Drosophila model for these types of studies depends on the availability of assays that reliably measure behavioral responses to ethanol. This report describes an assay suitable for assessing ethanol sensitivity and rapid tolerance in flies. Ethanol sensitivity measured in this assay is influenced by the volume and concentration of ethanol used, a variety of previously reported genetic manipulations, and also the length of time the flies are housed without food immediately prior to testing. In contrast, ethanol sensitivity measured in this assay is not affected by the vigor of fly handling, sex of the flies, and supplementation of growth medium with antibiotics or live yeast. Three different methods for quantitating ethanol sensitivity are described, all leading to essentially indistinguishable ethanol sensitivity results. The scalable nature of this assay, combined with its overall simplicity to set-up and relatively low expense, make it suitable for small and large scale genetic analysis of ethanol sensitivity and rapid tolerance in Drosophila.

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 ...

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 ...

Custom-made Microdialysis Probe Design

Microdialysis is a commonly used technique in neuroscience research. Therefore commercial probes are in great demand to monitor physiological, pharmacological and pathological changes in cerebrospinal fluid. Unfortunately, commercial probes are expensive for research groups in public institutions. In this work, a probe assembly is explained in detail to build a reliable, concentric, custom-made microdialysis probe for less than $10. The microdialysis probe consists of a polysulfone membrane with a molecular cut-off of 30 kDa. Probe in vitro recoveries of substances with different molecular weight (in the range of 100-1,600 Da) and different physicochemical properties are compared. The probe yields an in vitro recovery of approximately 20% for the small compounds glucose, lactate, acetylcholine and ATP. In vitro recoveries for neuropeptides with a molecular weight between 1,000-1,600 Da amount to 2-6%. Thus, while the higher molecular weight of the neuropeptides lowered in vitro recovery values, dialysis of compounds in the lower range (up to 500 Da) of molecular weights has no great impact on the in vitro recovery rate. The present method allows utilization of a dialysis membrane with other cut-off value and membrane material. Therefore, this custom-made probe assembly has the advantage of sufficient flexibility to dialyze substances in a broad molecular weight range. Here, we introduce a microdialysis probe with an exchange length of 2 mm, which is applicable for microdialysis in mouse and rat brain regions. However, dimensions of the probe can easily be adapted for larger exchange lengths to be used in larger animals.

Microdialysis is a commonly used technique in neuroscience research. Therefore commercial probes are in great demand.In this work a probe assembly is explained in detail to build a reliable, concentric, custom-made microdialysis probe for less than $10.

주문 제작 미세 투석 프로브 디자인

Microdialysis is a commonly used technique in neuroscience research. Therefore commercial probes are in great demand to monitor physiological, ...

Assessment of Dopaminergic Homeostasis in Mice by Use of High-performance Liquid Chromatography Analysis and Synaptosomal Dopamine Uptake

Dopamine (DA) is a modulatory neurotransmitter controlling motor activity, reward processes and cognitive function. Impairment of dopaminergic (DAergic) neurotransmission is strongly associated with several central nervous system-associated diseases such as Parkinson&#39;s disease, attention-deficit-hyperactivity disorder and drug addiction1,2,3,4. Delineating disease mechanisms involving DA imbalance is critically dependent on animal models to mimic aspects of the diseases, and thus protocols that assess specific parts of the DA homeostasis are important to provide novel insights and possible therapeutic targets for these diseases.
Here, we present two useful experimental protocols that when combined provide a functional read-out of the DAergic system in mice. Biochemical and functional parameters on DA homeostasis are obtained through assessment of DA levels and dopamine transporter (DAT) functionality5. When investigating the DA system, the ability to reliably measure endogenous levels of DA from adult brain is essential. Therefore, we present how to perform high-performance liquid chromatography (HPLC) on brain tissue from mice to determine levels of DA. We perform the experiment on tissue from dorsal striatum (dStr) and nucleus accumbens (NAc), but the method is also suitable for other DA-innervated brain areas.
DAT is essential for reuptake of DA into the presynaptic terminal, thereby controlling the temporal and spatial activity of released DA. Knowing the levels and functionality of DAT in the striatum is of major importance when assessing DA homeostasis. Here, we provide a protocol that allows to simultaneously deduce information on surface levels and function using a synaptosomal6 DA uptake assay.
Current methods combined with standard immunoblotting protocols provide the researcher with relevant tools to characterize the DAergic system.

Synaptosomal dopamine uptake and high-performance liquid chromatography analysis represent experimental tools to investigate dopamine homeostasis in mice by assessing the function of the dopamine transporter and levels of dopamine in striatal tissue, respectively. Here we present protocols to measure dopamine tissue content and assess the functionality of the dopamine transporter.

고성능 액체 크로마토그래피 분석 및 Synaptosomal 도파민 통풍 관의 사용에 의해 쥐에서 Dopaminergic 항상성의 평가

Dopamine (DA) is a modulatory neurotransmitter controlling motor activity, reward processes and cognitive function. Impairment of dopaminergic (DAergic) ...