We would like to acknowledge Beth Levine for critically reviewing the manuscript, Valerie Tassinari for developing the genotyping protocol, and Matt Ferola and Carlos Maya for their exceptional animal care.

모든 실험은 관리 및 동물의 사용을위한 건강 가이드의 국립 연구소에 의해 기술 된 프로토콜에 따라 수행하고 ALS TDI의 기관 동물 관리 및 사용위원회 (IACUC)에 의해 승인되었다. 1. 동물, 주택 및 연구 설계 참고 :. SOD1-G93A 마우스 식민지가 원래 들것 등 7에 의해 생성 된 높은 사본 B6SJLTgN (SOD1G93A) 1Gur 균주에서 유래되었다. 식민지는 현재 야생 형 C57BL / 6 SJL F1 암컷과 유전자 변형 C57BL / 6 SJL 남성 교차에 의해 유지되고있다. F1의 동물 B6 여성과 SJL 남성 교차에 의해 생성됩니다. 마우스는 나이 35~45일에서 루게릭 병의 TDI 매주로 제공됩니다. <ol><li> 마우스 적어도 하나의 주 연구에 할당하기 전에 (18 ~ 23 ° C와 40 % -60 % 습도의 온도에서 12 시간의 빛 / 어둠주기) 동물 시설에 적응하도록 허용합니다. </li><li> 주택의 남성 SOD1-G93A 마이크그룹이 남성은 싸움을하는 경향이 보관되어 있기 때문에 케이지 당 전자 하나의 마우스. 케이지 당 두 개의 최대 하우스 여성 SOD1-G93A 마우스. 플라스틱 오두막의 형태로, 환경 농축을 제공합니다. 쥐가 물 / 음식을 마시는 약물 투여를 필요로하는 연구의 일부가 아닌, 음식과 물을 광고 무제한을 제공합니다. 이러한 경우, 배급 및 기록 음식과 물. </li><li> 연령 하루 약 50 약물 효능 연구를 시작하고 64 쓰레기 일치 성 균형 마우스 (코호트 당 32)를 포함한다. 마지막으로, 연구를 수행하는 연구자는 실험 조건에 눈을 멀게되어 있는지 확인합니다. 참고 : 일반적인 약물 효능 연구는 그러나 그것은 이전 또는 이후 연구를 시작하는 것이 바람직 수 있습니다 치료 개입의 유형에 따라 연령 하루 50에서 시작하는 것이 좋습니다. 참고 : 코호트 당 32 마우스 성별 특정 약물 효과를 검출하기 위해 전원과 연구. 성별 별 비교가 필요없는 경우, 코호트 당 24 마우스는 전반적으로 약물 효과를 검출하기 위해 충분하다.</li></ol> 2. 체중 <ol><li> 매일 그램의 10 분의 정밀도와 연구의 모든 마우스 녹음 체중. 참고 : 체중은 질병의 진행 및 만성 약물 치료에서 발생할 수있는 불안감의 민감한 지표이다. </li><li> 마우스 체중 따라서, 아침에 더 높은되고, 하루의 과정 동안 상당히 다양 매일 같은 시간에 주위를 기록한다. </li></ol> 3. 신경 학적 채점 시스템 (NeuroScore) <ol><li> 매일 다음 세 가지 조건 마우스를 관찰하여, 각 마우스 평가 신경 스코어 (NS)을 순차적으로 수행 : b) 마우스) 도보시키고, 마비 (c)의 개시 이후) 마우스 꼬리에 의해 일시 중지된다 마우스 측면에 배치됩니다. <ol><li> 꼬리 서스펜션 테스트를 위해, 식품 분지에서 멀리, 1-2 초 동안 자신의 홈 케이지의 와이어 상단을 통해 꼬리의 기지에서 마우스 약 1.5」를 개최 OBS 동안뒷다리를 어빙. 서스펜션 시험을 3 회 반복하고, 가장 일관된 결과를 기록한다. </li><li> 보행 테스트를 들어, 일부 견인과 25cm의 도보 거리를 제공하는 깨끗한 표면에 마우스를 위치 (예 :., 종이 타월 미끄럼 방지하기 위해서 테이프). 그 걸음 걸이를 관찰하면서 마우스가 75cm 총 (종이 타월의 3 배 길이)을 돌​​아 다닐 수있다. </li><li> &quot;바로 잡고 반사&quot;시험을 위해, 그것의 왼쪽 또는 오른쪽에 마우스를 위치시키고 정지 시계가 양측으로부터 도움없이 자체를 오른쪽으로 소요 시간을 측정하여. 참고 :이 테스트의 하나의 시험이 필요합니다. </li></ol></li><li> 0 ~ 4 주에 독립적 규모에 (왼쪽 또는 오른쪽) 각 사지의 NS를 결정 오른쪽 뒷다리 NS 1 동안 예를 들어 0의 점수, 1, 왼쪽 뒷다리 NS 0을 나타냅니다. </li><li> (표 1에서 제공하는 요약) 관찰의 다음 세트에 해당하는 신경 학적 점수 (0-4)를 지정합니다 :<ol><li> 다음이 관찰되는 경우 NS 0 (프리 증상) 할당 : 마우스 꼬리에 의해 중단되는 경우, 뒷다리는 정상 스플레이 예를 제시이를 충분히 멀리 측 방향 중심선으로부터 연장하고 2 초 동안이 위치에서 유지되거나 이상. 마우스가 걸어 허용되는 경우, 정상 보용은 관찰된다. </li><li> 마우스가 꼬리에 의해 중단되는 경우, 뒷다리가 이상 스플레이, 예를 제시,이 붕괴 또는 부분적 측면 중간 선으로 축소하거나 꼬리 서스펜션 동안 떠는 또는이 후퇴되어 관찰되는 다음과 같은 경우 NS 1 (첫 번째 증상) 지정 / 쥐는. 마우스가 걸어 허용되는 경우, 정상 또는 약간 느린 보용이 관찰된다. </li><li> 다음이 관찰된다면 (2) NS (마비의 발병) 할당 : 마우스 꼬리에 의해 중단되는 경우, 뒷다리 훨씬 연장하지 않는 부분적으로 또는 완전히 붕괴이다. (여전히 관절 운동이있을 수 있습니다). 마우스가 걸어 허용되면 뒷다리 앞으로 MOT에 사용이온 그러나 발가락은 90cm의 산책하는 동안 두 배 이상 아래로 말려 있거나 다리의 어떤 부분은 케이지 바닥 / 테이블을 따라 드래그합니다. 마우스의 좌우 측에 배치 될 때, 양측에서 10 초 이내에 스스로를 잘 할 수있다. </li><li> 다음이 관찰되는 경우 (3) NS (마비)를 할당 : 마우스 꼬리에 의해 중단되는 경우, 최소의 뒷다리 OR 관절 운동 마비 강성이있다. 마우스가 걸어 허용하면 앞으로 그러나 사지가 전진 운동을 위해 사용하지 않는 움직임이있다. 마우스의 좌우 측에 배치 될 때, 양측에서 10 초 이내에 스스로를 잘 할 수있다. 참고 : 드물게 마비의 발병 후, 소변 습기가 뒷다리에 표시되지 않을 수 있습니다. 왼쪽 치료 소변 수분이 소변에 &quot;굽기&quot;피부 병변이 발생할 수 있습니다. 머리를 클리핑하여 소변의 수분을 치료, 따뜻한 물이 소변을 제거하기 위해 젖은 적용하고 부드럽게 건조시킨다. 피부 병변이있는 경우에 적용 antibio틱 연고. </li><li> 다음이 관찰되는 경우 (4) NS (애호 종점)를 할당 : 마우스 꼬리에 의해 중단되는 경우, 경질의 뒷다리가 마비. 마우스가 걸어 허용되면, 더 전진 운동이 없다. 마우스의 왼쪽과 오른쪽에 배치 될 때 어느 측에서 10 초 이내에 자체를 잘 할 수 없습니다. 즉, 반사를 바로 잡고의 부재. </li></ol></li></ol> 4. 데이터 입력 및 분석 <ol><li> 약물 효능 연구의 과정에서, 실험실 정보 관리 시스템 (LIMS)에 각 마우스 또는 시판 스프레드 시트 소프트웨어에 수집 된 데이터를 입력합니다. 주 : ALS TDI 개발 LIMS 소프트웨어는 내부 연구에서 수집 된 모든 데이터에 대한 중앙 저장소입니다, ALS TDI 조사관에 대한 시간에이 데이터를 편리하게 접근 할 수 있도록. LIMS는 모든 연구 일에 다시 혈통, 체중 또는 신경 학적 점수, 고유 ID로, 개인 마우스를 추적그 수명 기간 동안 언급 한 다른 관측을 포함한다. LIMS 궁극적 후속 분석을 위해 사용되는 스프레드 시트를 생성한다. </li><li> 약효 조사, 즉, 모든 마우스는 인간 종점에 도달 할 때, 사용 된 스프레드 시트 소프트웨어 (자재 표 참조) 좌우 뒷다리 NS을 평균하여 매일 각 마우스 결합 NS를 계산의 완료시 . 다음, 각각의 마우스와 각 마우스가 특정 NS에서 보낸다 일 수에 대한 각 NS의 평균 연령을 계산한다. </li><li> 통계 소프트웨어를 스프레드 시트 소프트웨어에서 데이터를 가져 와서 시간이 지남에 NS 전반적인 변화를 분석 (재료의 표 참조). 회귀의 영향과 중간 NS 연령과 약물 치료, 예상 NS 추정 순서 로지스틱 회귀 모델을 사용하여 이러한 데이터를 맞 춥니 다. 로그 - 랭크 카이 스퀘어 테스트를 사용 효과 우도 확률을 추정하고 NS 진행에 약물 치료 효과가 있는지 여부를 결정하는 통계LY 중요한 (단계별 지침에 대한 보충 코드 파일을 참조하십시오). 참고 : p- 값 &lt;0.05는 통계적으로 유의 한 것으로 간주됩니다. 주 : 통계 분석이 방법은 NS 데이터와 개별 마우스에 대한 NS 궤도의 변수 자연의 서수 본질을 존중합니다. 각각의 NS 궤도 충분히 NS 응답을 반영하지 않은 NS 시간 연관된 그룹 평균 결과 현저히 상이 할 수 있기 때문에 바람직하다. 또한, 경우에 따라서는 NS 특히 영 - 일 범위에서 변동될 수있다. </li><li> 마지막으로, 사용 그래프 소프트웨어는 약물 치료 및 제어 동물 그래프 &quot;서수는 NS에서 중간 나이 대 NS를 예상&quot;생성 (재료의 표 참조) (그림 2, 보충 코드 단계별 지침에 대한 파일 참조). 편리 y 축에서 질병 발병 (I의 평균 연령을 보간하여 약물 처리 군과 대조군 사이의 변화를 추정하기 위해이 그래프를 사용합니다.E., X 축상의 2 NS). 주 : 두 그룹 간의 평균 연령의 차이는 약물 - 유도 변화를 나타내고, 효과의 크기의 실제 측정 값으로 취급된다. 참고 : 증상이 질병의 진행을 둔화 나타내는 예상 NS 대 평균 연령 곡선에서 오른쪽으로 이동합니다. 반대로, 예상 NS 대 평균 연령 곡선의 좌측 이동은 증상이 질병의 진행을 가속 나타냅니다. </li></ol>

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A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+effect+of+probiotic+therapy+on+development+of+experimental+obesity+in+rats+caused+by+monosodium+glutamate.">The effect of probiotic therapy on development of experimental obesity in rats caused by monosodium glutamate.</a> Fiziol. Zh. 60 (2), 63-69 (2014).</li><li>Smittkamp, S. E., Brown, J. W., Stanford, J. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Time-course+and+characterization+of+orolingual+motor+deficits+in+B6SJL-Tg(SOD1-G93A)1Gur/J+mice.">Time-course and characterization of orolingual motor deficits in B6SJL-Tg(SOD1-G93A)1Gur/J mice.</a> Neuroscience. 151 (2), 613-621 (2008).</li><li>C, M., Gurney, M. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+low+expressor+line+of+transgenic+mice+carrying+a+mutant+human+Cu,Zn+superoxide+dismutase+(SOD1)+gene+develops+pathological+changes+that+most+closely+resemble+those+in+human+amyotrophic+lateral+sclerosis.">A low expressor line of transgenic mice carrying a mutant human Cu,Zn superoxide dismutase (SOD1) gene develops pathological changes that most closely resemble those in human amyotrophic lateral sclerosis.</a> Acta Neuropathol. 93 (6), 537-550 (1997).</li><li>Wegorzewska, I., Baloh, R. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=TDP-43-based+animal+models+of+neurodegeneration:+new+insights+into+ALS+pathology+and+pathophysiology.">TDP-43-based animal models of neurodegeneration: new insights into ALS pathology and pathophysiology.</a> Neurodegener. Dis. 8 (4), 262-274 (2011).</li><li>Hatzipetros, T., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=C57BL/6J+congenic+Prp-TDP43A315T+mice+develop+progressive+neurodegeneration+in+the+myenteric+plexus+of+the+colon+without+exhibiting+key+features+of+ALS.">C57BL/6J congenic Prp-TDP43A315T mice develop progressive neurodegeneration in the myenteric plexus of the colon without exhibiting key features of ALS.</a> Brain Res. 1584, 59-72 (2014).</li></ol>

The authors have nothing to disclose.

이 보고서에서 우리는 제대로 적용하는 경우, 안정적으로 SOD1-G93A 마우스에서 질병의 진행을 평가하고 약물 유발 변경을 식별 할 수있는, 빠르고 간단하게 비디오 프로토콜을 설명합니다. 다른 그룹은 SOD1-G93A 마우스 8-10에 대한 표현형 점수 시스템을 개발하는 동안, 그들은 종종 절차에 대한 충분한 정보를 제공하고 복제 부적절하지 않습니다. 따라서, 채점 시스템은 거의들을 개발하고 ?...

1990 년대 중반에 개발 SOD1-G93A 형질 전환 마우스 모델은 근 위축성 측삭 경화증 (ALS) 1의 가장 널리 사용되는 동물 모델되었습니다 이래. 이 형질 전환 마우스 모델은 유 전적으로 아미노산 93 (G93A)에 알라닌 돌연변이 ALS 관련 글리신을 품고 인간의 구리 / 아연 슈퍼 옥사이드 디스 뮤 타제 1 (SOD1) 유전자의 돌연변이 형태를 과발현하도록 설계되었다. G93A 돌연변이 일괄 메이크업 가족 ALS 사례 2의 약 5 분의 1 SOD1 유전자에 많은 변이 중 하나입니다. SOD1-G93A 모델, 그것은 궁극적으로, 이러한 운동 신경 손실, 점진적 근육 약화 및 위축 등 인간 루게릭 병의 병리학 적 특징의 많은 되풀이되었습니다 때문에 루게릭 병에 그것의 명백한 유전 링크에 추가 루게릭 병 치료제 개발 연구의 주력이되었다 마비와 사망 3. 종종 트랜스 제닉 동물 모델의 경우에서와 같이 비록, 고유 BI 존재SOD1-G93A 마우스와 관련된 학적 변화. 스콧 등. 최소 24 쓰레기 일치 성 균형 잡힌 쥐의 동료가 4 생물 다양성에 의해 생성 된 소음을 극복하기 위해 약물 효능 연구 설계에 필요하다고 결정했다. 공격적인 질환 진행과 매일 모니터링에 대한 필요성과 함께 이러한 연구에 필요한 동물의 다수의 신경 근육 강도 및 기능을 측정하기위한 정교한 시간 소모적 잠재적 스트레스 기법의 사용을 금지한다 (예를 들면, 근전도, 악력, 로토 등). 대신에, 확실하게 빠른 SOD1-G93A 마우스의 신경 근육 기능을 평가 생체 내 선별 도구의 필요성을 강조한다. ALS TDI에서 프로토콜은 평균에 마우스 미만 30 초에 SOD1-G93A 마우스의 신경 근육 기능의 신뢰성 평가를 허용하는 개발되었다. 이 프로토콜 encompa언론 및 비디오에서이 보고서에 설명되어있는 간단한 신경 학적 평가 시스템 (NeuroScore)를 사용하여 사지 기능의 일일 평가를 SSES. 뒷다리 적자는 초기가 SOD1-G93A 마우스 5,6에서 질병의 신경 학적 기호를보고 있기 때문에 NeuroScore는 뒷다리의 기능에 초점을 맞추고 있습니다. 또한, ALS TDI에 의해 개발 된 프로토콜은 마비 (약간 또는 부분 마비), 진행과 마비의 정도 발병의 공정한 평가를 제공하며 질병의 진행에 약물 유발 변경 사항을 식별 할 정도로 민감하다.

<table border="0" cellpadding="0" cellspacing="0">
	<tbody>
		<tr>
			<td>B6SJLTgN(SOD1G93A)1Gur strain</td>
			<td>The Jackson Laboratory, Bar Harbor, ME</td>
			<td>002726</td>
			<td>Strain of mice used in this study</td>
		</tr>
		<tr>
			<td>Breeders</td>
			<td>Biomedical Research Models, Inc., Worcester, MA</td>
			<td></td>
			<td>Colony maintainance</td>
		</tr>
		<tr>
			<td>Scale</td>
			<td>Navigator OHAUS, Parsippany, NJ</td>
			<td>Model N14120</td>
			<td>Used to weigh mice</td>
		</tr>
		<tr>
			<td>Nutra-GelH</td>
			<td>Bio-Serv, Flemington, NJ</td>
			<td>#S4798</td>
			<td>Wet food provided to sick mice</td>
		</tr>
		<tr>
			<td>Irradiated Lab Animal Diet</td>
			<td>Harlan, Indianapolis, IN</td>
			<td>2918-111914M</td>
			<td>Chow diet</td>
		</tr>
		<tr>
			<td>Lab-grade Sani-chips</td>
			<td>Harlan Teklad, Indianapolis, IN</td>
			<td>7090</td>
			<td>Bedding</td>
		</tr>
		<tr>
			<td>JMPH&#160;</td>
			<td>SAS Institute, Inc., SAS Campus Drive, Cary, NC</td>
			<td>&#160;v10.0.2</td>
			<td>Statistical software</td>
		</tr>
		<tr>
			<td>Microsoft Excel</td>
			<td>Microsoft, One Microsoft Way, Redmond, WA</td>
			<td>Excel 2013 (v 15.0)</td>
			<td>Spreadsheet software</td>
		</tr>
		<tr>
			<td>GraphPad Prism 5</td>
			<td>GraphPad software Inc., La Jolla, CA</td>
			<td>v5.4</td>
			<td>Graphing software</td>
		</tr>
		<tr>
			<td>Mouse Huts</td>
			<td>Bio-Serv, Flemington, NJ</td>
			<td>K3272</td>
			<td>For environmental enrichment</td>
		</tr>
	</tbody>
</table>

a quick phenotypic neurological scoring system for evaluating disease progression in the sod1-g93a mouse model of als

The SOD1-G93A transgenic mouse is the most widely used animal model of amyotrophic lateral sclerosis (ALS). At ALS TDI we developed a phenotypic screening protocol, demonstrated in video herein, which reliably assesses the neuromuscular function of SOD1-G93A mice in a quick manner. This protocol encompasses a simple neurological scoring system (NeuroScore) designed to assess hindlimb function. NeuroScore is focused on hindlimb function because hindlimb deficits are the earliest reported neurological sign of disease in SOD1-G93A mice. The protocol developed by ALS TDI provides an unbiased assessment of onset of paresis (slight or partial paralysis), progression and severity of paralysis and it is sensitive enough to identify drug-induced changes in disease progression. In this report, the combination of a detailed manuscript with video minimizes scoring ambiguities and inter-experimenter variability thus allowing for the protocol to be adopted by other laboratories and enabling comparisons between studies taking place at different settings. We believe that this video protocol can serve as an excellent training tool for present and future ALS researchers.

2014 평가 하였다 동안 90 남성과 94 여성 비 처리 SOD1-G93A 마우스의 신경 학적 점수 데이터는 ALS TDI에서 공부했다. 결과는 여성에 비하여 NS 1에서 그들의 수명의 큰 비율에 의해 입증 수컷 마우스는 일반적으로, 암컷 마우스보다 더 공격적인 질환 진행을 보여준다. NS 2, NS 3 성별에 걸쳐 거의 동일한 빈도로 발생합니다. 이 &quot;정상&quot;표현형 간주되기 때문에 NS 0은 분석에서 제외 하였다. NS (4) 또한 ...

Watch this Scientific Journal Video about A Quick Phenotypic Neurological Scoring System for Evaluating Disease Progression in the SOD1-G93A Mouse Model of ALS at JoVE.com

A Quick Phenotypic Neurological Scoring System for Evaluating Disease Progression in the SOD1-G93A Mouse Model of ALS

This video protocol describes a sensitive, reliable, and quick method for evaluating the neuromuscular deficits in a transgenic mouse model of amyotrophic lateral sclerosis.

ALS의 SOD1-G93A 마우스 모델에서 질병의 진행을 평가하기위한 빠른 표현형 신경 채점 시스템

The SOD1-G93A transgenic mouse is the most widely used animal model of amyotrophic lateral sclerosis (ALS). At ALS TDI we developed a phenotypic screening ...

a-quick-phenotypic-neurological-scoring-system-for-evaluating-disease

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19.2K 조회수.  ALS Therapy Development Institute. This video protocol describes a sensitive, reliable, and quick method for evaluating the neuromuscular deficits in a transgenic mouse model of amyotrophic lateral sclerosis.

비디오: A Quick Phenotypic Neurological Scoring System for Evaluating Disease Progression in the SOD1-G93A Mouse Model of ALS

Development of a Unilaterally-lesioned 6-OHDA Mouse Model of Parkinson's Disease

The unilaterally lesioned 6-hyroxydopamine (6-OHDA)-lesioned rat model of Parkinson's disease (PD) has proved to be invaluable in advancing our understanding of the mechanisms underlying parkinsonian symptoms, since it recapitulates the changes in basal ganglia circuitry and pharmacology observed in parkinsonian patients1-4. However, the precise cellular and molecular changes occurring at cortico-striatal synapses of the output pathways within the striatum, which is the major input region of the basal ganglia remain elusive, and this is believed to be site where pathological abnormalities underlying parkinsonian symptoms arise3,5.
In PD, understanding the mechanisms underlying changes in basal ganglia circuitry following degeneration of the nigro-striatal pathway has been greatly advanced by the development of bacterial artificial chromosome (BAC) mice over-expressing green fluorescent proteins driven by promoters specific for the two striatal output pathways (direct pathway: eGFP-D1; indirect pathway: eGFP-D2 and eGFP-A2a)8, allowing them to be studied in isolation. For example, recent studies have suggested that there are pathological changes in synaptic plasticity in parkinsonian mice9,10. However, these studies utilised juvenile mice and acute models of parkinsonism. It is unclear whether the changes described in adult rats with stable 6-OHDA lesions also occur in these models. Other groups have attempted to generate a stable unilaterally-lesioned 6-OHDA adult mouse model of PD by lesioning the medial forebrain bundle (MFB), unfortunately, the mortality rate in this study was extremely high, with only 14% surviving the surgery for 21 days or longer11. More recent studies have generated intra-nigral lesions with both a low mortality rate &gt;80% loss of dopaminergic neurons, however expression of L-DOPA induced dyskinesia11,12,13,14 was variable in these studies. Another well established mouse model of PD is the MPTP-lesioned mouse15. Whilst this model has proven useful in the assessment of potential neuroprotective agents16, it is less suitable for understanding mechanisms underlying symptoms of PD, as this model often fails to induce motor deficits, and shows a wide variability in the extent of lesion17, 18.
Here we have developed a stable unilateral 6-OHDA-lesioned mouse model of PD by direct administration of 6-OHDA into the MFB, which consistently causes &gt;95% loss of striatal dopamine (as measured by HPLC), as well as producing the behavioural imbalances observed in the well characterised unilateral 6-OHDA-lesioned rat model of PD. This newly developed mouse model of PD will prove a valuable tool in understanding the mechanisms underlying generation of parkinsonian symptoms.

Development of a Unilaterally-lesioned 6-OHDA Mouse Model of Parkinson&#039;s Disease

A protocol for performing unilateral 6-OHDA lesions of the medial forebrain bundle in mice is described. This method has a low mortality rate (13.3 %) with 89% of the surviving animals showing &#62;95% loss of striatal dopamine and 90.63&#177;-4.02 % ipsiversive rotational bias towards the side of the lesion.

파킨슨병의 일방적-6-OHDA lesioned 마우스 모델의 개발

The unilaterally lesioned 6-hyroxydopamine (6-OHDA)-lesioned rat model of Parkinson's disease (PD) has proved to be invaluable in advancing our ...

연구

의학

Utilizing Transcranial Magnetic Stimulation to Study the Human Neuromuscular System

Transcranial magnetic stimulation (TMS) has been in use for more than 20 years 1, and has grown exponentially in popularity over the past decade. While the use of TMS has expanded to the study of many systems and processes during this time, the original application and perhaps one of the most common uses of TMS involves studying the physiology, plasticity and function of the human neuromuscular system. Single pulse TMS applied to the motor cortex excites pyramidal neurons transsynaptically 2 (Figure 1) and results in a measurable electromyographic response that can be used to study and evaluate the integrity and excitability of the corticospinal tract in humans 3. Additionally, recent advances in magnetic stimulation now allows for partitioning of cortical versus spinal excitability 4,5. For example, paired-pulse TMS can be used to assess intracortical facilitatory and inhibitory properties by combining a conditioning stimulus and a test stimulus at different interstimulus intervals 3,4,6-8. In this video article we will demonstrate the methodological and technical aspects of these techniques. Specifically, we will demonstrate single-pulse and paired-pulse TMS techniques as applied to the flexor carpi radialis (FCR) muscle as well as the erector spinae (ES) musculature. Our laboratory studies the FCR muscle as it is of interest to our research on the effects of wrist-hand cast immobilization on reduced muscle performance6,9, and we study the ES muscles due to these muscles clinical relevance as it relates to low back pain8. With this stated, we should note that TMS has been used to study many muscles of the hand, arm and legs, and should iterate that our demonstrations in the FCR and ES muscle groups are only selected examples of TMS being used to study the human neuromuscular system.

Transcranial magnetic stimulation (TMS) is a non-invasive tool to gain insight on the physiology and function of the human nervous system. Here, we present our TMS techniques to study cortical excitability of the upper limb and lumbar musculature.

인간 신경근육학 시스템을 공부하기 Transcranial 자기 자극을 활용

Transcranial magnetic stimulation (TMS) has been in use for more than 20 years 1, and has grown exponentially in popularity over the past decade.  While ...

Use of Animal Model of Sepsis to Evaluate Novel Herbal Therapies

Sepsis refers to a systemic inflammatory response syndrome resulting from a microbial infection. It has been routinely simulated in animals by several techniques, including infusion of exogenous bacterial toxin (endotoxemia) or bacteria (bacteremia), as well as surgical perforation of the cecum by cecal ligation and puncture (CLP)1-3. CLP allows bacteria spillage and fecal contamination of the peritoneal cavity, mimicking the human clinical disease of perforated appendicitis or diverticulitis. The severity of sepsis, as reflected by the eventual mortality rates, can be controlled surgically by varying the size of the needle used for cecal puncture2. In animals, CLP induces similar, biphasic hemodynamic cardiovascular, metabolic, and immunological responses as observed during the clinical course of human sepsis3. Thus, the CLP model is considered as one of the most clinically relevant models for experimental sepsis1-3.
Various animal models have been used to elucidate the intricate mechanisms underlying the pathogenesis of experimental sepsis. The lethal consequence of sepsis is attributable partly to an excessive accumulation of early cytokines (such as TNF, IL-1 and IFN-&gamma;)4-6 and late proinflammatory mediators (e.g., HMGB1)7. Compared with early proinflammatory cytokines, late-acting mediators have a wider therapeutic window for clinical applications. For instance, delayed administration of HMGB1-neutralizing antibodies beginning 24 hours after CLP, still rescued mice from lethality8,9, establishing HMGB1 as a late mediator of lethal sepsis. The discovery of HMGB1 as a late-acting mediator has initiated a new field of investigation for the development of sepsis therapies using Traditional Chinese Herbal Medicine. In this paper, we describe a procedure of CLP-induced sepsis, and its usage in screening herbal medicine for HMGB1-targeting therapies.

Sepsis refers to a systemic inflammatory response syndrome resulting from a microbial infection, and can be simulated by a surgical technique termed cecal ligation and puncture (CLP). Here we describe a method to use CLP-induced animal model to screen medicinal herbs for therapeutic agents.

소설 허벌 요법을 평가하는 패혈증의 동물 모델의 사용

Sepsis refers to a systemic inflammatory response syndrome resulting from a microbial infection. It has been routinely simulated in animals by several ...

Clinical Testing and Spinal Cord Removal in a Mouse Model for Amyotrophic Lateral Sclerosis (ALS)

Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disorder resulting in progressive degeneration of motoneurons. Peak of onset is around 60 years for the sporadic disease and around 50 years for the familial disease. Due to its progressive course, 50% of the patients die within 30 months of symptom onset. In order to evaluate novel treatment options for this disease, genetic mouse models of ALS have been generated based on human familial mutations in the SOD gene, such as the SOD1 (G93A) mutation. Most important aspects that have to be evaluated in the model are overall survival, clinical course and motor function. Here, we demonstrate the clinical evaluation, show the conduction of two behavioural motor tests and provide quantitative scoring systems for all parameters. Because an in depth analysis of the ALS mouse model usually requires an immunohistochemical examination of the spinal cord, we demonstrate its preparation in detail applying the dorsal laminectomy method. Exemplary histological findings are demonstrated. The comprehensive application of the depicted examination methods in studies on the mouse model of ALS will enable the researcher to reliably test future therapeutic options which can provide a basis for later human clinical trials.

Clinical Testing and Spinal Cord Removal in a Mouse Model for Amyotrophic Lateral Sclerosis ALS

A mouse model for amyotrophic lateral sclerosis (ALS) is examined clinically and behaviorally. As a prerequisite for an accompanying immunohistological analysis the preparation of the spinal cord is depicted in detail.

루게릭병에 대한 마우스 모델 (ALS)의 임상 시험 및 척수 제거

Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disorder resulting in progressive degeneration of motoneurons. Peak of onset is around 60 ...

A Simple Method of Mouse Lung Intubation

A simple procedure to intubate mice for pulmonary function measurements would have several advantages in longitudinal studies with limited numbers or expensive animal. One of the reasons that this is not done more routinely is that it is relatively difficult, despite there being several published studies that describe ways to achieve it. In this paper we demonstrate a procedure that eliminates one of the major hurdles associated with this intubation, that of visualizing the trachea during the entire time of intubation. The approach uses a 0.5 mm fiberoptic light source that serves as an introducer to direct the intubation cannula into the mouse trachea. We show that it is possible to use this procedure to measure lung mechanics in individual mice over a time course of at least several weeks. The technique can be set up with relatively little expense and expertise, and it can be routinely accomplished with relatively little training. This should make it possible for any laboratory to routinely carry out this intubation, thereby allowing longitudinal studies in individual mice, thereby minimizing the number of mice needed and increasing the statistical power by using each mouse as its own control.

This paper describes a striaghforward and efficient method of intubating mice for pulmonary function measurements or pulmonary instillation, that allows the mice to recover and be studied at later times. The procedure involves an inexpensive fiberoptic light source that directly illuminates the trachea.

마우스 폐 삽관 법의 간단한 방법

A  simple procedure to intubate mice for pulmonary function measurements would  have several advantages in longitudinal studies with limited numbers or  ...

Bioluminescent Orthotopic Model of Pancreatic Cancer Progression

Pancreatic cancer has an extremely poor five-year survival rate of 4-6%. New therapeutic options are critically needed and depend on improved understanding of pancreatic cancer biology. To better understand the interaction of cancer cells with the pancreatic microenvironment, we demonstrate an orthotopic model of pancreatic cancer that permits non-invasive monitoring of cancer progression. Luciferase-tagged pancreatic cancer cells are resuspended in Matrigel and delivered into the pancreatic tail during laparotomy. Matrigel solidifies at body temperature to prevent leakage of cancer cells during injection. Primary tumor growth and metastasis to distant organs are monitored following injection of the luciferase substrate luciferin, using in vivo imaging of bioluminescence emission from the cancer cells. In vivo imaging also may be used to track primary tumor recurrence after resection. This orthotopic model is suited to both syngeneic and xenograft models and may be used in pre-clinical trials to investigate the impact of novel anti-cancer therapeutics on the growth of the primary pancreatic tumor and metastasis.

Improved understanding of pancreatic cancer biology is critically needed to enable the development of better therapeutic options to treat pancreatic cancer. To address this need, we demonstrate an orthotopic model of pancreatic cancer that permits non-invasive monitoring of cancer progression using in vivo bioluminescence imaging.

췌장암의 진행 발광 동소 모델

Pancreatic cancer has an extremely poor five-year survival rate of 4-6%.  New therapeutic options are critically needed and depend on improved ...

Flexible Colonoscopy in Mice to Evaluate the Severity of Colitis and Colorectal Tumors Using a Validated Endoscopic Scoring System

The use of modern endoscopy for research purposes has greatly facilitated our understanding of gastrointestinal pathologies. In particular, experimental endoscopy has been highly useful for studies that require repeated assessments in a single laboratory animal, such as those evaluating mechanisms of chronic inflammatory bowel disease and the progression of colorectal cancer. However, the methods used across studies are highly variable. At least three endoscopic scoring systems have been published for murine colitis and published protocols for the assessment of colorectal tumors fail to address the presence of concomitant colonic inflammation. This study develops and validates a reproducible endoscopic scoring system that integrates evaluation of both inflammation and tumors simultaneously. This novel scoring system has three major components: 1) assessment of the extent and severity of colorectal inflammation (based on perianal findings, transparency of the wall, mucosal bleeding, and focal lesions), 2) quantitative recording of tumor lesions (grid map and bar graph), and 3) numerical sorting of clinical cases by their pathological and research relevance based on decimal units with assigned categories of observed lesions and endoscopic complications (decimal identifiers). The video and manuscript presented herein were prepared, following IACUC-approved protocols, to allow investigators to score their own experimental mice using a well-validated and highly reproducible endoscopic methodology, with the system option to differentiate distal from proximal endoscopic colitis (D-PECS).

Over the past few years, new generation endoscopes have emerged as important diagnostic research aids for evaluating murine colitis and colorectal tumors. We present herein a detailed protocol for endoscopic assessment of inflammation and colorectal tumors in mice, as well as a novel scoring system that uses decimal identifiers to document the endoscopic severity of colitis and colorectal tumors.

검증 된 내시경 채점 시스템을 사용하여 대장염과 대장 종양의 위험을 평가하기 위해 마우스의 유연한 대장 내시경 검사

The use of modern endoscopy for research purposes has greatly facilitated our understanding of gastrointestinal pathologies. In particular, experimental ...

Establishment and Characterization of UTI and CAUTI in a Mouse Model

Urinary tract infections (UTI) are highly prevalent, a significant cause of morbidity and are increasingly resistant to treatment with antibiotics. Females are disproportionately afflicted by UTI: 50% of all women will have a UTI in their lifetime. Additionally, 20-40% of these women who have an initial UTI will suffer a recurrence with some suffering frequent recurrences with serious deterioration in the quality of life, pain and discomfort, disruption of daily activities, increased healthcare costs, and few treatment options other than long-term antibiotic prophylaxis. Uropathogenic Escherichia coli (UPEC) is the primary causative agent of community acquired UTI. Catheter-associated UTI (CAUTI) is the most common hospital acquired infection accounting for a million occurrences in the US annually and dramatic healthcare costs. While UPEC is also the primary cause of CAUTI, other causative agents are of increased significance including Enterococcus faecalis. Here we utilize two well-established mouse models that recapitulate many of the clinical characteristics of these human diseases. For UTI, a C3H/HeN model recapitulates many of the features of UPEC virulence observed in humans including host responses, IBC formation and filamentation. For CAUTI, a model using C57BL/6 mice, which retain catheter bladder implants, has been shown to be susceptible to E. faecalis bladder infection. These representative models are being used to gain striking new insights into the pathogenesis of UTI disease, which is leading to the development of novel therapeutics and management or prevention strategies.

The ability to model urinary tract infections (UTI) is crucial in order to be able to understand bacterial pathogenesis and spawn the development of novel therapeutics. This work&#8217;s goal is to demonstrate mouse models of experimental UTI and catheter associated UTI that recapitulate and predict findings seen in humans.

마우스 모델의 설립 및 요로 감염 및 CAUTI의 특성

Urinary tract infections (UTI) are highly prevalent, a significant cause of morbidity and are increasingly resistant to treatment with antibiotics. ...

Echocardiographic Approaches and Protocols for Comprehensive Phenotypic Characterization of Valvular Heart Disease in Mice

The aim of this manuscript and accompanying video is to provide an overview of the methods and approaches used for imaging heart valve function in rodents, with detailed descriptions of the appropriate methods for anesthesia, the echocardiographic windows used, the imaging planes and probe orientations for image acquisition, the methods for data analysis, and the limitations of emerging technologies for the evaluation of cardiac and valvular function. Importantly, we also highlight several future areas of research in cardiac and heart valve imaging that may be leveraged to gain insights into the pathogenesis of valve disease in preclinical animal models. We propose that using a systematic approach to evaluating cardiac and heart valve function in mice can result in more robust and reproducible data, as well as facilitate the discovery of previously underappreciated phenotypes in genetically-altered and/or physiologically-stressed mice.

This protocol provides a detailed description of the echocardiographic approach for comprehensive phenotyping of heart and heart valve function in mice.

마우스의 심장 판막 질환의 종합 표현형 특성에 대한 심 초음파 접근 및 프로토콜

The aim of this manuscript and accompanying video is to provide an overview of the methods and approaches used for imaging heart valve function in ...

Evaluating the Procedure for Performing Awake Cystometry in a Mouse Model

Awake filling cystometry has been used for a long time to evaluate bladder function in freely moving mice, however, the specific methods used, vary among laboratories. The goal of this study was to describe the microsurgical procedure used to implant an intravesical tube and the experimental technique for recording urinary bladder pressure in an awake, freely moving mouse. In addition, experimental data is presented to show how surgery, as well as tubing type and size, affect lower urinary tract function and recording sensitivity. The effect of tube diameter on pressure recording was assessed in both polyethylene and polyurethane tubing with different internal diameters. Subsequently, the best performing tube from both materials was surgically implanted into the dome of the urinary bladder of male C57BL/6 mice. Twelve-hour, overnight micturition frequency was recorded in healthy, intact animals and animals 2, 3, 5, and 7 days post-surgery. At harvest, bladders were assessed for signs of swelling using gross observation and were subsequently processed for pathological analysis. The greatest extent of bladder swelling was observed on day 2 and 3, which correlated with behavioral voiding data showing significantly impaired bladder function. By day 5, bladder histology and voiding frequency had normalized. Based on the literature and evidence provided by our studies, we propose the following steps for in vivo recording of intravesical pressure and voided volume in an awake mouse: 1) Perform the surgery using an operating microscope and microsurgical tools, 2) Use polyethylene-10 tubing to minimize movement artifacts, and 3) Perform cystometry on post-operative day 5, when bladder swelling resolves.

This study describes the surgical procedures and experimental techniques for performing awake cystometry in a freely moving mouse. In addition, it provides experimental evidence to support its optimization and standardization.

마우스 모델에서 깨어나는 방광 측정을 수행하기위한 절차 평가

Awake filling cystometry has been used for a long time to evaluate bladder function in freely moving mice, however, the specific methods used, vary among ...