연구는 Linköping 대학교 Östergötland의 군 의회에 의해 재정 지원되었다.

1. 지느러미 루트를 통해 Ovariectomy <ol><li> 산소 / 질소 산화물 30% / 70 %의 혼합에서 isoflurane 1.5 %를 (유도 4 %)를 사용하여 예를 들어, 동물을 마취. 직장 온도계에 연결된 열판에서 발생하기 쉬운 위치에있는 동물을 놓습니다. 그것은 동물과 플레이트 사이에 고립의 얇은 레이어를 가지고하는 것이 중요하다, 그래서 온도계 피드백 시스템은 37 이내에 체온을 유지 ± 0.5 ° C. 눈 보호를위한 젤을 적용하고 수술 후 통증 완화를위한 동물의 목 측면에는 subcutaneously 식염수에 희석 5 MG의 Rimadyl / kg bodyweight를 관리합니다. </li><li> 장골의의 문장에서 cephally 4 * 4cm (3 생쥐에 대한 * 3 ㎝) 영역을 면도. 철저하게 4 * 4cm의 조리개 수술 시트와 동물을 취재하기 전에 예제 Jodopax 위해 함께 음모 지역을 씻는다. </li><li> 2~3센티미터 중간선 절개를 만들어 퉁명스럽게 기본 근막으로부터 피부를 해부. 중간선의 측면 하나 CM은, 다른 절개를 죽하다근막 윽. 퉁명스럽게 복강에 도달하기 전까지 같은 피상적 가능한이 근막 아래 laterally 해부. </li><li> 족집게 사용하여 복강에 난소를 둘러싸는 지방 조직을 잡고 부드럽게 빼낸다. 난소를 확인하고 자궁 뿔을하고이 구조의 proximally 혈관 0.5-1 센티미터 ligate. 난소와 출혈도 잡았 지방 조직을 잘라 복강에 남아있는 조직 입어요. contralateral 근막에 다른 절개를 확인하고 절차를 반복합니다. </li><li> 선호 monofilament의 봉합사를 이용하여 상처를 닫고 마취에서 깨어 동물 봅시다. 수술 후 최소 2 시간 동안 열띤 케이지 (25-27 ℃)에서 동물을 놓습니다. 동물은 처음 며칠 postoperatively 별도로 지내게됩니다. 그것은 새장가 자주 복구 단계에서 청소 / 교체되는 중추이다. 스물 네 시간 수술 후에 희석 앞으로 5 MG의 Rimadyl / kg bodyweight으로 subcutaneously 동물을 주입염분. </li></ol> 2. Silastic 캡슐을 통해 17β-estradiol의 피하 관리 <ol><li> 완전히 깨 기름에 17β-estradiol의 원하는 금액을 섞는다. 쥐 및 생쥐의 생리적 농도의 경우, 우리는 각각 180 μg / ML 및 μg / ML 18-36을 추천합니다. 위약 캡슐은 순수 참기름으로 채워진다. </li><li> 3cm (생쥐 2 ㎝) silastic 튜브의 길이와 5mm (쥐 3 ㎜) 도전을위한 작은 주걱 나무 스틱의 길이를 잘라. 포셉 사용하여 전체 길이를 채울 17β-estradiol/sesame 오일 솔루션을 주입하기 전에 silastic 캡슐로 한 목조 플러그를 조인다. 부드럽게 다른 나무 플러그 (플러그는 캡슐에 영구적으로 유지하기로하고)으로 열린 끝에 silastic 캡슐를 쏠. 이식하기 전에 하룻밤 나머지 17β-estradiol 솔루션 캡슐을 품어. </li><li> 캡슐의 주입은 쥐의 ovariectomy에서 사용 마취 중에 수행할 수 있습니다. 마일을 위해CE 그러나, ovariectomy 후 14 일간의 복구 기간이 강력하게 연약한 피부를 통해 튀어나와 캡슐의 위험을 최소화하는 것이 좋습니다. </li><li> 1.1에서와 같이, 또는 ovariectomy의 절개를 닫은 후 수술 준비 후, 면도하고 동물의 목 지느러미 부분에 2 * 2cm 영역을 씻는다. 1cm의 절개를 만들어 퉁명스럽게 캡슐을위한 충분한 공간과 함께 caudolaterally 피하 주머니 해부. silastic 캡슐 (이 시간 시점까지 17β-estradiol 용액에 incubated되었는지) 닦아 부드럽게 이후 봉합사에 의해 닫혀 절개를 통해 그것을 설치합니다. 그것은 확실히 중추입니다 특히 상처 가까운 지역에서 가능한 거의 같은 캡슐 변종은 피부. 이식에 대한 캡슐의 생산에서 그것은 오염은 피할 수있다는 것이 중요합니다. </li></ol> 3. 너트 - 크림 Nutella 통해 17β-estradiol의 Peroral 관리 <ol><li> 완전히 용해 17β-그리고 참깨 기름 estradiol,하고 Nutella와 참깨 오일을 섞는다. 우리는 각각의 매일 부분은 17β-estradiol 28 μg, 5 μL 참기름과 kg 바디 쥐에 대한 무게와 17β-estradiol 1.12 μg, 0.312 μL 참기름과 각 30 g 마우스 60 MG의 Nutella 당 1g의 Nutella를 포함하는 것이 좋습니다. 위약 너트 - 크림은 17β-estradiol이없는 경우를 제외하고 동일하게 준비가되어 있습니다. </li><li> 실험 전에 5 일 동안 호르몬없는 너트 - 크림을 먹는 동물을 훈련. 연수의 첫 번째 3 일, 쥐가 최적의 자택 연습장에서 (약 0.5 g에 Nutella 2-5 동물) 그룹에서 훈련을해야합니다. 훈련의 네번째와 다섯째 날에는 동물 실험 상황을 닮은 매일 부분을 수신하기 전에 별도의 케이지에 부착 할 것입니다. </li><li> 실험을 진행하는 동안 동물 Nutella 부분은 작은 세라믹 타일에 제공되는 이후, 모든 먹이 때를 위해 별도의 케이지에 부착 할 것입니다. 교육은 위에서, O로 수행되었을 경우UR 경험 쥐 및 생쥐 이상의 95 % 너트-크림을 수락하고, 한번에 완전히 길들여지지, 동물의 대부분은 초 내에 그것을 소비한다는 것입니다. </li></ol> 4. 대표 결과 현재의 방법을 재현하는 것을 목표로 생리 혈청 17β-estradiol 농도가 있으며, 약 5-140 PG / 쥐 및 생쥐 4,6를위한 5-35 PG / ML위한 ML. 피하 silastic 캡슐은 위의 지시에 따라, 300g 암컷 Sprague-Dawley의 쥐에 투여했을 때, 천천히 PG / ML 일 40 2-10 PG / ML 일 35에서 17β-estradiol 혈청 농도를 감소 생산하고 있습니다. 다만, 즉시 행정 후에, 1000 PG / ML 주변의 혈청 농도 피크는 4,5 (그림 1A) 8 시간 이내에 생리적 범위로 떨어지는, 기대할 수 있습니다. 25g 암컷 C57BL / 6 마​​우스에 투여 36 μg의 17β-estradiol/mL로 위에서 설명한 생쥐에 대한 silastic 캡슐은 혈청 conc을 세우다천천히 약 90 PG / ML 하루 2-25 PG / ML 일 35 감소한 entrations. 이것은 (그림 1B) 6을 조사하는 것은 아니지만, 쥐에서 발견되었다대로 이것은 아마도 유사한 피크 앞에있다. 복용량과 peroral Nutella 방식은 300g 암컷 Sprague-Dawley의 쥐에 투여했을 때, 위에 설명된 10-70 PG / ML (그림 2A) 4 사이의 일일 변동과 17β-estradiol의 생리적 수준을 렌더링. 위 복용량을 사용하는 마우스의 경우 일일 수준은 약 20 120 PG / ML (그림 2B) 6 사이의 변동. <img alt="그림 1" src="/files/ftp_upload/4013/4013fig1.jpg" /> <st<html> . 쥐의 17β-estradiol 농도 제시 룽&gt; 그림 1화한 그래프, 4,5와 쥐 (나중 포인트에서 N까지 시간 포인트에서 = 10 최대 24 H, N = 15) (B, N = 10) 캡슐 이식 후 6. silastic 캡슐은 1000 혈청 농도의 짧은 초기 피크를 생산 PG / ML지만, 100 PG / ML 아래 수준까지 시간 이내에 감소. 꾸준히 감소하고 있지만 오는 4~5주 들어, 캡슐은, 혈청 농도를 안정적으로 얻을 수 있습니다. 그늘진 지역은 각각 온전 여성 쥐 및 생쥐의 생리 17β-estradiol 혈청 농도에 해당합니다. 시각적으로 그래프에서 시연되지하더라도, 남북 각각의 변화는 (매우 낮은 평균 27 일부터 쥐의 1백83퍼센트로 28에서 silastic 캡슐 방식으로 마우스의 1백15%에 변이 계수 렌더링, 17β-estradiol 농도에있다 농도는 종종) 유사 높은 계수를 생성합니다. iles/ftp_upload/4013/4013fig2.jpg &quot;/&gt; 그림 2 쥐의 17β-estradiol 농도를 제시화한 그래프 (; N = 15). 4와 생쥐 (B, N = 10) Nutella에서 매일 호르몬 관리의 결과로서 6. peroral Nutella 방법은 피하 또는 정맥 주사 방법으로 생산되는만큼 높은 그러나 안하 시니 지금까지 매일 봉우리 렌더링. 이것은 인간의 호르몬 대체 요법의 pharmacokinetics을 모방한 것이었하고 peroral 치료는 위의 예제는 초기 96시간을 제시하는 경우에도 한 필요를 위해 진행할 수 있습니다. 그늘진 지역은 각각 온전 여성 쥐 및 생쥐의 생리 17β-estradiol 혈청 농도에 해당합니다. 비록 시각적으로 그래프에서 보여준 아니라 남북 각각의 변화는 (매우 낮은 평균 농도 Nutella 방법과 마우스로 쥐의 40~132%에서 그리고 13 일부터 2백19퍼센트로 변이 계수 렌더링, 17β-estradiol 농도에있다 자주 높이를 생성변동 계수).

<ol>
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저자가 공개하는 게 없다.

이전 연구에서 경험 위에 표시 관리 모드는 17β-estradiol의 생리 혈청 농도를 생산의 관점에서 상업 천천히 릴리스 알약과 주사 우수이었다. 느린 - 릴리스 알약이 매우 높고 장시간 혈청 농도 봉우리를 생산하는 동안 주사는 극단 매일 호르몬 변동의 원인과 동물 4-6의 반복 스트레스 처리를 필요로합니다. 적절한 관리 모드를 선택의 중요성은 낮은 복용량 17β-estradiol 관리 regimens (예 : 현재 ...

<table border="1"><tbody><tr><td> 시약의 이름 </td><td> 회사 </td><td> 카탈로그 번호 </td><td> 댓글 </td></tr><tr><td> Rimadyl, 50 밀리그램 / ML </td><td> 화이자 </td><td> 462,986 </td><td> ovariectomy 후 수술 후 진통제. </td></tr><tr><td> Lubrithal 아이 젤 </td><td> 레오 연구소 </td><td></td><td> 수술 중 안구 보호. </td></tr><tr><td> Jodopax의 수의사 </td><td> Pharmaxim </td><td></td><td> 수술 소독. </td></tr><tr><td> Vicryl 4-0 봉합사 </td><td> 존슨 앤 존슨 </td><td></td><td> 봉합과 난소를 내고 상처를주는 군. </td></tr><tr><td> 17β-estradiol,&gt; 98% </td><td> 시그마 </td><td> E8875-1G </td><td> silastic 캡슐 및 Nutella위한 참깨 기름합니다. </td></tr><tr><td> 향유 </td><td> 시그마 </td><td> S3547-1L </td><td> 17β-estradiol을 해소. </td></tr><tr><td> Silastic 튜빙, 아이디 / OD : 1.575/3.175 mm </td><td> 다우 코닝 </td><td> 508-008 </td><td> silastic 캡슐 생산. </td></tr><tr><td> 나무 작은 주걱 스틱 </td><td> Selefatrade </td><td> 12-0783 </td><td> silastic 캡슐의 상한. </td></tr><tr><td> Nutella </td><td> Ferrero </td><td></td><td> peroral 17β-estradiol에 대한 중간. </td></tr></tbody></table>

ovariectomy and 17&beta;-estradiol replacement in rats and mice: a visual demonstration

Estrogens는 효과의 매우 넓은 스펙트럼과 여성의 성 호르몬의 가족입니다. 쥐 및 생쥐는 에스 트로겐 연구에 사용하는 경우 이들은 일반적으로 17β-estradiol exogenously을 교체, 빠르게 사이클링 호르몬 생산을 ablate하기 위해 ovariectomized됩니다. 호르몬은 생리 혈청 농도를 얻기 위해 관리되는 방식에 관한 합의 부족 그러나,있다. 17β-estradiol 수준 / 관리 방법 뿌리깊은 실험 결과는 1-3 미치는 영향 때문에 이것은 중요합니다. 우리는 피하 silastic 캡슐과 당 구두 너트 - 크림 Nutella는 상용 느리게 출시 알약 (미국의 기업 혁신 연구에서 생산)로 우수하다고 찾는 17β-estradiol 행정의 다양한 모드를 특징 연구 시리즈가 있고 17β-estradiol 4-6의 생리 혈청 농도를 생산의 관점에서 매일 주사. 교수실 사이이전 buprenorphine 행정 7 사용되었습니다 너트 - 크림 방법의 antages은 에스 ​​트로겐 관리에 사용하면 그것이 peroral 호르몬 대체 요법과 유사하고 비침습가된다. 피하 silastic 캡슐이 편리하며 가장 안정적인 혈청 농도를 생산하고 있습니다. 이 비디오 문서 ovariectomy의 단계별 데모 및 silastic 캡슐 및 행정 절차의 중요한 측면에 대한 논의 뒤에 쥐 및 생쥐에서 peroral Nutella 의한 17β-estradiol 호르몬 대체가 포함되어 있습니다.

Watch this Scientific Journal Video about Ovariectomy and 17Î²-estradiol Replacement in Rats and Mice: A Visual Demonstration at JoVE.com

Ovariectomy and 17&amp;#946;-estradiol Replacement in Rats and Mice: A Visual Demonstration

silastic 캡슐과 peroral Nutella 의한 Ovariectomy 및 17β-estradiol의 후속 교체는 쥐 및 생쥐에서 증명하고 있습니다.

고양이와 쥐의 Ovariectomy 및 17β-estradiol 대체 : 비주얼 데모

Estrogens are a family of female sexual hormones with an exceptionally wide spectrum of effects. When rats and mice are used in estrogen research they are ...

ovariectomy-17-estradiol-replacement-rats-mice-visual

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JoVE Journal

Biology

Ovariectomy and 17&beta;-estradiol Replacement in Rats and Mice: A Visual Demonstration

56.7K 조회수.  Linköping University. silastic 캡슐과 peroral Nutella 의한 Ovariectomy 및 17β-estradiol의 후속 교체는 쥐 및 생쥐에서 증명하고 있습니다.

비디오: Ovariectomy and 17&#946;-estradiol Replacement in Rats and Mice: A Visual Demonstration

A Magnetic Tether System to Investigate Visual and Olfactory Mediated Flight Control in Drosophila

It has been clear for many years that insects use visual cues to stabilize their heading in a wind stream. Many animals track odors carried in the wind. As such, visual stabilization of upwind tracking directly aids in odor tracking. But do olfactory signals directly influence visual tracking behavior independently from wind cues? Also, the recent deluge of research on the neurophysiology and neurobehavioral genetics of olfaction in Drosophila has motivated ever more technically sophisticated and quantitative behavioral assays. Here, we modified a magnetic tether system originally devised for vision experiments by equipping the arena with narrow laminar flow odor plumes. A fly is glued to a small steel pin and suspended in a magnetic field that enables it to yaw freely. Small diameter food odor plumes are directed downward over the fly s head, eliciting stable tracking by a hungry fly. Here we focus on the critical mechanics of tethering, aligning the magnets, devising the odor plume, and confirming stable odor tracking.

Here we describe how to tether a fly in an olfactory magnetic-tether (OMT) apparatus. We describe how to align the rare-earth magnets and odor ports, and how to set mass flow rates for both the stimulus delivery and vacuum suction to achieve optimal odor tracking.

Drosophila에서 Visual과 강한 중재 플라이트 컨트롤을 맡은 이후로 자기 테더 시스템

It has been clear for many years that insects use visual cues to stabilize their heading in a wind stream. Many animals track odors carried in the wind. ...

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생물학

Demonstration of Cutaneous Allodynia in Association with Chronic Pelvic Pain

Pelvic pain is a common condition that is associated with dysmenorrhea and endometriosis. In some women the severe episodes of cyclic pain change and the resultant pain becomes continuous and this condition becomes known as Chronic Pelvic Pain. This state can be present even after the appropriate medical or surgical therapy has been instituted. It can be associated with pain and tenderness in the muscles of the abdomen wall and intra-pelvic muscles leading to severe dyspareunia. Additional symptoms of irritable bowel and interstitial cystitis are common. A common sign of the development of this state is the emergence of cutaneous allodynia which emerges from the so-called viscero-somatic reflex. A simple bedside test for the presence of cutaneous allodynia is presented that does not require excessive time or special equipment. This test builds on previous work associated with changes in sensation related to gall bladder function and the viscera-somatic reflex(1;2).
The test is undertaken with the subject s permission after an explanation of how the test will be performed. Allodynia refers to a condition in which a stimulus that is not normally painful is interpreted by the subject as painful. In this instance the light touch associated with a cotton-tipped applicator would not be expected to be painful. A positive test is however noted by the woman as suddenly painful or suddenly sharp. The patterns of this sensation are usually in a discrete pattern of a dermatome of the nerves that innervate the pelvis.
The underlying pathology is now interpreted as evidence of neuroplasticity as a consequence of severe and repeating pain with changes in the functions of the dorsal horns of the spinal cord that results in altered function of visceral tissues and resultant somatic symptoms(3). 
The importance of recognizing the condition lies in an awareness that this process may present coincidentally with the initiating condition or after it has been treated. It also permits the clinician to evaluate the situation from the perspective that alternative explanations for the pain may be present that may not require additional surgery.

Demonstration of Cutaneous Allodynia in Association with Chronic Pelvic Pain

A demonstration of the bedside test for cutaneous allodynia and its clinical implications.

실연 피부 Allodynia 만성 골반 통증과 공동으로

Pelvic pain is a common condition that is associated with dysmenorrhea and endometriosis.  In some women the severe episodes of cyclic pain change and the ...

Single-cell Suction Recordings from Mouse Cone Photoreceptors

Rod and cone photoreceptors in the retina are responsible for light detection. In darkness, cyclic nucleotide-gated (CNG) channels in the outer segment are open and allow cations to flow steadily inwards across the membrane, depolarizing the cell. Light exposure triggers the closure of the CNG channels, blocks the inward cation current flow, and thus results in cell hyperpolarization. Based on the polarity of photoreceptors, a suction recording method was developed in 1970s that, unlike the classic patch-clamp technique, does not require penetrating the plasma membrane 1. Drawing the outer segment into a tightly-fitting glass pipette filled with extracellular solution allows recording the current changes in individual cells upon test-flash exposure. However, this well-established "outer-segment-in (OS-in)" suction recording is not suitable for mouse cone recordings, because of the low percentage of cones in the mouse retina (3%) and the difficulties in identifying the cone outer segments. Recently, an inner-segment-in (IS-in) recording configuration was developed to draw the inner segment/nuclear region of the photoreceptor into the recording pipette 2,3. In this video, we will show how to record from individual mouse cone photoresponses using single-cell suction electrode.

We will show how to record flash responses from single mouse cones using a suction electrode.

마우스 콘 Photoreceptors에서 단일 셀 석션 레코딩

Rod and cone photoreceptors in the retina are responsible for light detection. In darkness, cyclic nucleotide-gated (CNG) channels in the outer segment ...

A Practical Approach to Genetic Inducible Fate Mapping: A Visual Guide to Mark and Track Cells In Vivo

Fate maps are generated by marking and tracking cells in vivo to determine how progenitors contribute to specific structures and cell types in developing and adult tissue. An advance in this concept is Genetic Inducible Fate Mapping (GIFM), linking gene expression, cell fate, and cell behaviors in vivo, to create fate maps based on genetic lineage.
GIFM exploits X-CreER lines where X is a gene or set of gene regulatory elements that confers spatial expression of a modified bacteriophage protein, Cre recombinase (CreERT). CreERT contains a modified estrogen receptor ligand binding domain which renders CreERT sequestered in the cytoplasm in the absence of the drug tamoxifen. The binding of tamoxifen releases CreERT, which translocates to the nucleus and mediates recombination between DNA sequences flanked by loxP sites. In GIFM, recombination typically occurs between a loxP flanked Stop cassette preceding a reporter gene such as GFP.
Mice are bred to contain either a region- or cell type-specific CreER and a conditional reporter allele. Untreated mice will not have marking because the Stop cassette in the reporter prevents further transcription of the reporter gene. We administer tamoxifen by oral gavage to timed-pregnant females, which provides temporal control of CreERT release and subsequent translocation to the nucleus removing the Stop cassette from the reporter. Following recombination, the reporter allele is constitutively and heritably expressed. This series of events marks cells such that their genetic history is indelibly recorded. The recombined reporter thus serves as a high fidelity genetic lineage tracer that, once on, is uncoupled from the gene expression initially used to drive CreERT.
We apply GIFM in mouse to study normal development and ascertain the contribution of genetic lineages to adult cell types and tissues. We also use GIFM to follow cells on mutant genetic backgrounds to better understand complex phenotypes that mimic salient features of human genetic disorders.
This video article guides researchers through experimental methods to successfully apply GIFM. We demonstrate the method using our well characterized Wnt1-CreERT;mGFP mice by administering tamoxifen at embryonic day (E)8.5 via oral gavage followed by dissection at E12.5 and analysis by epifluorescence stereomicroscopy. We also demonstrate how to micro-dissect fate mapped domains for explant preparation or FACS analysis and dissect adult fate-mapped brains for whole mount fluorescent imaging. Collectively, these procedures allow researchers to address critical questions in developmental biology and disease models.

A Practical Approach to Genetic Inducible Fate Mapping: A Visual Guide to Mark and Track Cells In Vivo

Genetic Inducible Fate Mapping (GIFM) marks and tracks cells with fine spatial and temporal control in vivo and elucidates how cells from a specific genetic lineage contribute to developing and adult tissues. Demonstrated here are the techniques required to fate map E12.5 mouse embryos for epifluorescent and explant analysis.

유전자 Inducible의 운명 매핑에 대한 실용적인 접근법 : 셀을 선택하고 추적하는 비주얼 가이드 VIVO에서</em

Fate maps are generated by marking and tracking cells in vivo to determine how progenitors contribute to specific structures and cell types in developing ...

Manual Restraint and Common Compound Administration Routes in Mice and Rats

Being able to safely and effectively restrain mice and rats is an important part of conducting research. Working confidently and humanely with mice and rats requires a basic competency in handling and restraint methods. This article will present the basic principles required to safely handle animals. One-handed, two-handed, and restraint with specially designed restraint objects will be illustrated. Often, another part of the research or testing use of animals is the effective administration of compounds to mice and rats. Although there are a large number of possible administration routes (limited only by the size and organs of the animal), most are not used regularly in research. This video will illustrate several of the more common routes, including intravenous, intramuscular, subcutaneous, and oral gavage. The goal of this article is to expose a viewer unfamiliar with these techniques to basic restraint and substance administration routes. This video does not replace required hands-on training at your facility, but is meant to augment and supplement that training.

Working safely and humanely with research rodents requires a core competency in handling and restraint methods. This article will present the basic principles required to safely handle and effectively administer compounds to mice and rats.

수동 규제와 마우스 및 쥐 일반적인 합성 관리 경로

Being able to safely and effectively restrain mice and rats is an important part of conducting research.  Working confidently and humanely with mice and ...

Diagnostic Necropsy and Selected Tissue and Sample Collection in Rats and Mice

There are multiple sample types that may be collected from a euthanized animal in order to help diagnose or discover infectious agents in an animal colony. Proper collection of tissues for further histological processing can impact the quality of testing results. This article describes the conduct of a basic gross examination including identification of heart, liver, lungs, kidneys, and spleen, as well as how to collect those organs. Additionally four of the more difficult tissue/sample collection techniques are demonstrated. Lung collection and perfusion can be particularly challenging as the tissue needs to be properly inflated with a fixative in order for inside of the tissue to fix properly and to enable thorough histologic evaluation. This article demonstrates the step by step technique to remove the lung and inflate it with fixative in order to achieve optimal fixation of the tissue within 24 hours. Brain collection can be similarly challenging as the tissue is soft and easily damaged. This article demonstrates the step by step technique to expose and remove the brain from the skull with minimal damage to the tissue. The mesenteric lymph node is a good sample type in which to detect many common infectious agents as enteric viruses persist longer in the lymph node than they are shed in feces. This article demonstrates the step by step procedure for locating and aseptically removing the mesenteric lymph node. Finally, identification of infectious agents of the respiratory tract may be performed by bacterial culture or PCR testing of nasal and/or bronchial fluid aspirates taken at necropsy. This procedure describes obtaining and preparing the respiratory aspirate sample for bacterial culture and PCR testing.

This article describes the procedures for conducting a basic postmortem examination of a mouse or rat, and the collection of basic organs, as well as more challenging sample types from for histological, microbiological, and PCR evaluation.

진단 검시와 고양이와 마우스의 선택 조직 및 샘플 수집

There are multiple sample types that may be collected from a euthanized animal in order to help diagnose or discover infectious agents in an animal ...

Design and Analysis of Temperature Preference Behavior and its Circadian Rhythm in Drosophila

The circadian clock regulates many aspects of life, including sleep, locomotor activity, and body temperature (BTR) rhythms1,2. We recently identified a novel Drosophila circadian output, called the temperature preference rhythm (TPR), in which the preferred temperature in flies rises during the day and falls during the night 3. Surprisingly, the TPR and locomotor activity are controlled through distinct circadian neurons3. Drosophila locomotor activity is a well&#160;known circadian behavioral output and has provided strong contributions to the discovery of many conserved mammalian circadian clock genes and mechanisms4. Therefore, understanding TPR will lead to the identification of hitherto unknown molecular and cellular circadian mechanisms. Here, we describe how to perform and analyze the TPR assay. This technique not only allows for dissecting the molecular and neural mechanisms of TPR, but also provides new insights into the fundamental mechanisms of the brain functions that integrate different environmental signals and regulate animal behaviors. Furthermore, our recently published data suggest that the fly TPR shares features with the mammalian BTR3. Drosophila are ectotherms, in which the body temperature is typically behaviorally regulated. Therefore, TPR is a strategy used to generate a rhythmic body temperature in these flies5-8. We believe that further exploration of Drosophila TPR will facilitate the characterization of the mechanisms underlying body temperature control in animals.

Design and Analysis of Temperature Preference Behavior and its Circadian Rhythm in Drosophila

We recently identified a novel Drosophila circadian output, temperature preference rhythm (TPR), in which the preferred temperature in flies rises during the day and falls during the night. TPR is regulated independently from another circadian output, locomotor activity. Here&#160;we describe the design and analysis of TPR in Drosophila.

드로소필라의 온도 선호도 행동과 Circadian 리듬의 설계 및 분석

The circadian clock regulates many aspects of life, including sleep, locomotor activity, and body temperature (BTR) rhythms1,2. We recently identified a ...

Physiology Lab Demonstration: Glomerular Filtration Rate in a Rat

Measurements of glomerular filtration rate (GFR), and the fractional excretion of sodium (Na) and potassium (K) are critical in assessing renal function in health and disease. GFR is measured as the steady state renal clearance of inulin which is filtered at the glomerulus, but not secreted or reabsorbed along the nephron. The fractional excretion of Na and K can be determined from the concentration of Na and K in plasma and urine. The renal clearance of inulin can be demonstrated in an anesthetized animal which has catheters in the femoral artery, femoral vein and bladder. The equipment and supplies used for this procedure are those commonly available in a research core facility, and thus makes this procedure a practical means for measuring renal function. The purpose of this video is to demonstrate the procedures required to perform a lab demonstration in which renal function is assessed before and after a diuretic drug. The presented technique can be utilized to assess renal function in rat models of renal disease.

The purpose of this protocol is to demonstrate the principles and techniques for measuring and calculating glomerular filtration rate, urine flow rate, and excretion of sodium and potassium in a rat. This demonstration can be used to provide students with an overall conceptual understanding of how to measure renal function.

생리학 실험실 데모 : 쥐의 사구체 여과 속도

Measurements of glomerular filtration rate (GFR), and the fractional excretion of sodium (Na) and potassium (K) are critical in assessing renal function ...

Validation of a Mouse Model to Disrupt LINC Complexes in a Cell-specific Manner

Nuclear migration and anchorage within developing and adult tissues relies heavily upon large macromolecular protein assemblies called LInkers of the Nucleoskeleton and Cytoskeleton (LINC complexes). These protein scaffolds span the nuclear envelope and connect the interior of the nucleus to components of the surrounding cytoplasmic cytoskeleton. LINC complexes consist of two evolutionary-conserved protein families, Sun proteins and Nesprins that harbor C-terminal molecular signature motifs called the SUN and KASH domains, respectively. Sun proteins are transmembrane proteins of the inner nuclear membrane whose N-terminal nucleoplasmic domain interacts with the nuclear lamina while their C-terminal SUN domains protrudes into the perinuclear space and interacts with the KASH domain of Nesprins. Canonical Nesprin isoforms have a variable sized N-terminus that projects into the cytoplasm and interacts with components of the cytoskeleton. This protocol describes the validation of a dominant-negative transgenic mouse strategy that disrupts endogenous SUN/KASH interactions in a cell-type specific manner. Our approach is based on the Cre/Lox system that bypasses many drawbacks such as perinatal lethality and cell nonautonomous phenotypes that are associated with germline models of LINC complex inactivation. For this reason, this model provides a useful tool to understand the role of LINC complexes during development and homeostasis in a wide array of tissues.

Nuclear envelope proteins play a central role in many basic biological processes and have been implicated in a variety of human diseases. This protocol describes a new Cre/Lox-based mouse model that allows for the spatiotemporal control of LINC complexes disruption.

마우스 모델의 유효성 검사는 세포 고유의 방식으로 LINC 단지를 방해

Nuclear migration and anchorage within developing and adult tissues relies heavily upon large macromolecular protein assemblies called LInkers of the ...

In Vivo Detection and Analysis of Rb Protein SUMOylation in Human Cells

The post-translational modifications of proteins are critical for the proper regulation of intracellular signal transduction. Among these modifications, small ubiquitin-related modifier (SUMO) is a ubiquitin-like protein that is covalently attached to the lysine residues of a variety of target proteins to regulate cellular processes, such as gene transcription, DNA repair, protein interaction and degradation, subcellular transport, and signal transduction. The most common approach to detecting protein SUMOylation is based on the expression and purification of recombinant tagged proteins in bacteria, allowing for an in vitro biochemical reaction which is simple and suitable for addressing mechanistic questions. However, due to the complexity of the process of SUMOylation in vivo, it is more challenging to detect and analyze protein SUMOylation in cells, especially when under endogenous conditions. A recent study by the authors of this paper revealed that endogenous retinoblastoma (Rb) protein, a tumor suppressor that is vital to the negative regulation of the cell cycle progression, is specifically SUMOylated at the early G1 phase. This paper describes a protocol for the detection and analysis of Rb SUMOylation under both endogenous and exogenous conditions in human cells. This protocol is appropriate for the phenotypical and functional investigation of the SUMO-modification of Rb, as well as many other SUMO-targeted proteins, in human cells.

In Vivo Detection and Analysis of Rb Protein SUMOylation in Human Cells

Small ubiquitin-related modifier (SUMO) family proteins are conjugated to the lysine residues of target proteins to regulate various cellular processes. This paper describes a protocol for the detection of retinoblastoma (Rb) protein SUMOylation under endogenous and exogenous conditions in human cells.

Vivo에서 검색 및 Rb 단백질 SUMOylation 인간 세포에서의 분석

The post-translational modifications of proteins are critical for the proper regulation of intracellular signal transduction. Among these modifications, ...