이 작품은 척수 및 건강의 인디애나 국무부에서 뇌 손상 연구 기금에서 NIH / NINDS 부여 4R00 NS 057,940, 그리고 부여 SCBI 200-12에 의해 지원되었다.

1. 아래 부분을 잘라냄 수술을위한 간단한 장치를 만들기 <ol><li> 우리가 만든 아래 부분을 잘라냄 장치는 세 부분으로 (그림 1)으로 구성되어 있습니다 : 유도 관의 부착 및 바늘을 수 있으며, 수술 중에 두개골 창을 가로질러 앉아 스테인레스 스틸 또는 플라스틱으로 만들어진 (1) 지원 플레이트 (2) 위치에 바늘을 보유하고 있으며 바늘의 회전을 허용하는지도 튜브 및 끝에서 ~ 3mm에서 90도 구​​부린 및 회전 삽입로 이동됩니다 (3) 바늘. </li><li> 두께 1-1.5 mm, 7 조각 ~ 10 X 30mm 직사각형 스테인리스 스틸 또는 투명 플라스틱 (1) 지원 플레이트를 만들기 위해 잘라. 1.5 인치 22 게이지 (BD 회사, # 305156)와 1.5 인치 25 게이지 (BD 회사, # 305127) 주사 바늘을 준비합니다. 지도 튜브하려면, 플라스틱 종료하고 그 총 길이가 31-32mm (2)에 관한 있도록 22 게이지 주사 바늘의 낮은 바늘 끝을 잘라. 모래 금속 끝을은 평평하고 부드럽게 만들 수 있습니다. 이 유도 관의 마지막 길이는 25 게이지 주사 바늘보다 짧은 5~6mm에 대해해야합니다. </li><li> 바늘이 금속의 가장자리에 수직되어 있는지 확인하고, 지원 접시에지도 튜브를 해결하기 위해 cyanoacrylate 접착제를 사용하십시오. </li><li> 지도 튜브에 25 게이지 주사 바늘을 삽입하고, 팁 (그림 1) 2.5-3 mm에서 바늘을 90도 구​​부. </li><li> 바늘의 결국 (그림 1, 바늘 정지)에 gluing 작은 플라스틱 튜브로 바늘의 수직 이동 범위를 조정합니다. 바늘의 마지막 이동 범위는 바늘이 대뇌 피질에 삽입 할 수있는 방법을 깊이 결정, 및 P21 쥐에 대한 1.6-1.8 mm와 P21 생쥐에 대한 1.3-1.5 mm해야합니다. 이 길이는 동물의 종 및 연령에 맞게 변경해야합니다. </li><li> 구부러진 끝 (Fig.1 바늘 표시기)와 같은 방향으로 바늘의 위쪽 끝에 작은 구리 와이어 또는 테이프의 작은 조각을 첨부합니다. 와이어 또는 테이프는 아래 부분을 잘라냄 수술 중 바늘의 회전 각도 (그림 1)을 나타냅니다. </li></ol> 2. 동물 준비 <ol><li> 모든 수술 장비 유리 비드 살균기, 또는 70 % 에탄올로 소독과 멸균, autoclaved해야합니다. </li><li> 22일 (P20 - 22) 또는 같은 나이에 CD1 쥐들이 - 우리는 출생 후의 20 세에서 스프 / Dawley 쥐를 사용합니다. 마우스 또는 쥐의 다른 변종은 특정 프로젝트에 사용할 수 있습니다. </li><li> 300ml의 IP 주사와 마우스를 마취 80 MG / kg와 xylazine 8 MG / kg. 동물이 꼬리 핀치에 응답하지 않습니다 후 수술이 시작됩니다. 수술하는 동안, 필요한 경우 한 3의 부스터 선량은 마취 칵테일의 원래 복용량은 원래 마취 상태를 복원하기 위해 부여하실 수 있습니다. </li><li> 전기 헤어 트리머와 함께 동물의 두피에서 머리카락을 자르고. 마취 동안 건조로부터 보호하기 위해 동물의 눈을에 안과 연고의 작은 금액을 적용합니다. 70 % 에탄올에 이어 10 % povidone - 요오드 솔루션을 사용하여 두피를 소독. </li><li> 안정적인 고정 위치에 머리를 유지하는 stereotaxic기구에 동물을 탑재합니다. 수술 전반에 걸쳐, 우리는 저체온증을 방지하기 위해 가열 패드에있는 동물을 유지. </li><li> 람다의 눈 사이에 연장 메스를 사용하여 두피에 중간선 앞쪽에 - 후부 절개를합니다. 이외에도 피부를 뽑아 충분히 왼쪽 두개골을 노출 hemostats를 사용합니다. </li><li> 왼쪽 두개골에 사각형 상처를 확인하고, 골막을 모으 메스를 사용합니다. 이 단계는 출혈을 줄이고 두개골에 드릴 용이합니다. </li></ol> 3. undercuts 만들기 <ol><li> 노출된 두개골 지역에 무균 식염수의 작은 금액을 추가하고 혈액을 청소하고 지역을 건조하는 데 몇 Q - 팁을 사용합니다. </li><li> 수술 현미경, 왼쪽 두개골의 중앙에 사각형 홈을 (쥐에 ~ 5 X 7mm, 4 마우스의 X 5mm) (약 왼쪽 sensorimotor 피질 이상) 드릴링 시작합니다. 두개골에 염분의 드롭을 적용하면 드릴링 및 없어져요 열을 용이하게합니다. 뚫고있는 뼈 약 삼분의이 깊이 후, 여분의 염분을 제거하고 면봉을 넣었로 드릴링 지역을 청소하십시오. 뼈의 중앙 조각은 포셉의 부드러운 터치에 따라 움직일 수있는 때까지 천천히 조심스럽게 깊이 드릴. </li><li> 조심스럽게 뼈의 가장자리에 포셉의 날카로운 팁을 삽입하고 천천히 왼쪽 반구를 노출 포셉 운동으로 뼈의 중앙 조각을 제거합니다. </li><li> 수술 현미경, 아래 부분을 잘라냄 장치를 보유하고 동양 parasagittal 방향으로 바늘과 바늘 끝과 대상 피​​질 영역의 시각화를 유지하는 등 약간 caudally 있도​​록 중간선, 틸팅 장치의 지원 플레이트 직각. 두개골 윈도우의 중앙에, 우수한 화살 치료 측면 지역 1~2mm으로 바늘의 끝을 겨냥해서 대형 선박의 직접적인 침투를 방지. </li><li> 혈관을 여분을 줄 수있는만큼 경질 통해 PIA 아래 수평 방향으로 바늘을 삽입합니다. 아래 부분을 잘라냄 장치의 바닥을 앉아두개골 창의 양쪽 가장자리에 너무 바늘은 대뇌 피질의 표면 (그림 1) 정상적인 위치입니다. 손으로 두개골 창의 가장자리 모두에서 장치를 휴식하는 것은 크게 감소하거나 다음 절차를 수행하는 동안 흔들어 손을 제거합니다. PIA 아래에 바늘을 올려 후 천천히 바늘이 깊이 갈 수있을 때까지 그것이 transcortical 상처를 만들 수 낮추십시오. ° 떨어진 VI는 아래 부분을 잘라냄 반 원을 흰색 물질 / 깊은 레이어를 만들 수 중간선에서 ~ 135를 회전합니다. 다음 PIA 아래 다시 바늘을 올립니다. 역방향 장치를 기울이기와 바늘을 철회. </li><li> 선택, 하나는 위의 절차 (단계 2.5) 반복하지만 윈도우의 측면 가장자리에서 추가 transcortical 상처를 만들만큼 바늘을 회전하지 않고 있습니다. 이것은보다 완벽한 대뇌 피질의 격리를 생성합니다. </li><li> 보호하기 위해 두개골 윈도우에 플라스틱 필름의 조각 (6 X 6mm)을 장소, 두피 치료. 완전히 마취에서 회복까지 온수 패드에있는 동물을 놓으십시오. </li></ol> 4. 대표 결과 : 코로나 피질 조각은 아래 부분을 잘라냄 수술의 성공을 확인하는 준비하실 수 있습니다. &gt; 이주 수술 후 준비 슬라이스에서 transcortical과 아래 부분을 잘라냄 인하는 현미경의 저전력 목표 (그림 2)에 따라보고 알 수있는 있습니다. 부분적으로 고립된 피질은 보통 약간 얇은되고, epileptiform 활동 분야 잠재적인 기록 (그림 3)과 대뇌 피질의 조각의 대부분에서 발견하실 수 있습니다. 흰 문제 또는 깊은 피질 층에 큰 구멍 대비 형성에서 lesioned 피질, 또는 위 또는 아래 하얀 문제되고 잘못된 컷의 밝은 극적인 추가 실험을위한 두뇌를 사용할 수 없게 될 것입니다. <img src="/files/ftp_upload/2840/2840fig1.jpg" alt="그림 1"/> 그림 1. 아래 부분을 잘라냄 장치의 구조 및 응용 프로그램. 지침 튜브 (2) 스테인레스 스틸 또는 투명 플라스틱으로되어 지원 플레이트 (1)에 붙어있다. 주사 바늘 (3) 팁로 ~ 3mm의지도 튜브 및 구부러진를 통해 삽입됩니다. 니들은 플라스틱 튜브로 만들어진 정지하면 바늘의 수직 이동 범위가 ~ 1.2 mm 각각 P21 마우스 및 쥐에 사용 ~ 1.5 mm로 제한되도록 바늘의 위쪽 끝에 붙어있다. 작은 구리 와이어의 세그먼트는 구부러진 바늘의 방향에 대해 바늘 지표로 핸들 아래에 고정됩니다. 장치가 기울어져 있으며, 두개골 윈도우의 양쪽 가장자리와 접촉 상처가 pial 표면에 평행하게 만들 수 있도록합니다. <img src="/files/ftp_upload/2840/2840fig2.jpg" alt="그림 2"/> 그림 2. 허리의 연한 고기 슬라이스의 대표 이미지. P48 쥐에서 아래 부분을 잘라냄 병변 이후 두 주를 준비했습니다 슬라이스의 A. 형광 이미지. 절삭 상처는 형광 염료 DiI에 의해 분류되었다. 오른쪽에 흰색 화살표는 transcortical 컷을 표시하고, 하단에있는 화살표는 계층 IV과 흰색 물질의 경계하지만 합격 아래 부분을 잘라냄 나타냅니다. <img src="/files/ftp_upload/2840/2840fig3.jpg" alt="그림 3"/> 그림 3. 아래 부분을 잘라냄 뇌 슬라이스에서 아래 부분을 잘라냄 슬라이스. 필드 잠재 녹화에서 필드 가능성이 기록은 부상 대뇌 피질 조직 hyperexcitability을 제안, epileptiform 활동을 전시.

<ol>
	<li>Hoffman, S. N., Salin, P. A., Prince, D. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Chronic+neocortical+epileptogenesis+in+vitro.">Chronic neocortical epileptogenesis in vitro.</a> J Neurophysiol. 71, 1762-1773 (1994).</li><li>Topolnik, L., Steriade, M., Timofeev, I. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Hyperexcitability+of+intact+neurons+underlies+acute+development+of+trauma-related+electrographic+seizures+in+cats+in+vivo.">Hyperexcitability of intact neurons underlies acute development of trauma-related electrographic seizures in cats in vivo.</a> Eur J Neurosci. 18, 486-496 (2003).</li><li>Graber, K., Prince, D. 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> Models of Seizures and Epilepsy. , 477-493 (2005).</li><li>Nita, D. A., Cisse, Y., Timofeev, I., Steriade, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Increased+propensity+to+seizures+after+chronic+cortical+deafferentation+in+vivo.">Increased propensity to seizures after chronic cortical deafferentation in vivo.</a> J Neurophysiol. 95, 902-913 (2006).</li><li>Sharpless, S. K., Halpern, L. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+electrical+excitability+of+chronically+isolated+cortex+studied+by+means+of+permanently+implanted+electrodes.">The electrical excitability of chronically isolated cortex studied by means of permanently implanted electrodes.</a> Electroencephalogr Clin Neurophysiol. 14, 244-255 (1962).</li><li>Echlin, F. A., Battista, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Epileptiform+Seizures+from+Chronic+Isolated+Cortex.">Epileptiform Seizures from Chronic Isolated Cortex.</a> Arch Neurol. 9, 154-170 (1963).</li><li>Prince, D. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Epileptogenic+neurons+and+circuits.">Epileptogenic neurons and circuits.</a> Adv Neurol. 79, 665-684 (1999).</li><li>Marin-Padilla, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Developmental+neuropathology+and+impact+of+perinatal+brain+damage.+II:+white+matter+lesions+of+the+neocortex.">Developmental neuropathology and impact of perinatal brain damage. II: white matter lesions of the neocortex.</a> J Neuropathol Exp Neurol. 56, 219-235 (1997).</li><li>Jin, X., Prince, D. A., Huguenard, J. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Enhanced+excitatory+synaptic+connectivity+in+layer+v+pyramidal+neurons+of+chronically+injured+epileptogenic+neocortex+in+rats.">Enhanced excitatory synaptic connectivity in layer v pyramidal neurons of chronically injured epileptogenic neocortex in rats.</a> J Neurosci. 26, 4891-4900 (2006).</li><li>Li, H., Prince, D. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Synaptic+activity+in+chronically+injured,+epileptogenic+sensory-motor+neocortex.">Synaptic activity in chronically injured, epileptogenic sensory-motor neocortex.</a> J Neurophysiol. 88, 2-12 (2002).</li><li>Salin, P., Tseng, G. F., Hoffman, S., Parada, I., Prince, D. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Axonal+sprouting+in+layer+V+pyramidal+neurons+of+chronically+injured+cerebral+cortex.">Axonal sprouting in layer V pyramidal neurons of chronically injured cerebral cortex.</a> J Neurosci. 15, 8234-8245 (1995).</li><li>Avramescu, S., Nita, D. A., Timofeev, I. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Neocortical+post-traumatic+epileptogenesis+is+associated+with+loss+of+GABAergic+neurons.">Neocortical post-traumatic epileptogenesis is associated with loss of GABAergic neurons.</a> J Neurotrauma. 26, 799-812 (2009).</li><li>Avramescu, S., Timofeev, I. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Synaptic+strength+modulation+after+cortical+trauma:+a+role+in+epileptogenesis.">Synaptic strength modulation after cortical trauma: a role in epileptogenesis.</a> J Neurosci. 28, 6760-6772 (2008).</li></ol>

관심 없음 충돌 선언하지 않습니다.

아래 부분을 잘라냄 모델은 외상후 epileptogenesis를 공부를위한 고효율 시스템입니다. 전형적인 수술을 완료하는 데 약 20-30분 소요하고, evoked 또는 자연 epileptiform 활동이 이주 수술 후 1-2 대부분의 동물에서 슬라이스에 기록하실 수 있습니다. 더 중요한 건,이 모델은 출혈, 염증, 부종, axotomy와의 연결을 죽음 7과 같은 외상성 뇌 손상 다음과 같은 변경 사항을 모방한 것이었 지요. 뿐...

<table border="1"><tbody><tr><td> 재료 이름 </td><td> 유형 </td><td> 회사 </td><td> 카탈로그 번호 </td></tr><tr><td> Foredom micromotor 키트 </td><td> 장비 </td><td> Foredom </td><td> K.1070 </td></tr><tr><td> 1.5 인치 22 게이지 주사 바늘 </td><td> 자료 </td><td> BD 회사 </td><td> 305156 </td></tr><tr><td> 1.5 인치 25 게이지 주사기 바늘 </td><td> 자료 </td><td> BD 회사 </td><td> 305127 </td></tr><tr><td> Cyanoacrylate 접착제 </td><td> 자료 </td><td> 테드 펠라 </td><td> 14,450 </td></tr></tbody></table>

preparing undercut model of posttraumatic epileptogenesis in rodents

부분적으로 격리 피질 ( &quot;가격을 내리다&quot;) 외상후 epileptogenesis의 동물 모델입니다. 수술은 sensorimotor 피질과 대뇌 피질의 특정 영역이 크게 인근 피질과 subcortical 지역 1-3에서 격리되도록 흰색 물질 (아래 부분을 잘라냄) 아래에. 통해 절단 포함 수술 다음과 같은 두 개 이상의 주 지연 시간 후, epileptiform의 방전은 설치류 1 뇌 슬라이스에 기록 수 있으며, 전기 또는 행동 압류는 그러한 고양이와 원숭이 4-6와 같은 다른 종 생체내에서 볼 수 있습니다. 이 잘 구축 동물 모델을 생성하는 효율적이며, 외상성 뇌 손상의 여러 가지 중요한 특성을 모방한 것이었 지요. 그러나, 그것은 기술적으로 무료로 손이 작은 쥐 두뇌의 대뇌 피질의 병변 정확한를 만들기 위해 시도 도전이다. 처음에 스탠포드 대학 1 박사 데이비드 왕자의 실험실에서 설립 절차에 따라, 우리가 마우스 및 쥐이 모델의 준비를위한 수술을 수행하는 향상된 기술을 제시한다. 우리는 간단한 수술 장치를 만들고 더 정확하고 일관된 결과를 생성하기 위해 깊이와 각도 절단의 더 나은 통제를 얻을 그것을 사용하는 방법을 보여줍니다. 장치 수 있도록 간단하고, 절차가 빨리 배웁니다. 이 동물 모델의 생성은 외상후 epileptogenesis의 메커니즘에 대한 연구에 대한 효율적인 시스템을 제공합니다.

Watch this Scientific Journal Video about Preparing Undercut Model of Posttraumatic Epileptogenesis in Rodents at JoVE.com

Preparing Undercut Model of Posttraumatic Epileptogenesis in Rodents

부분적으로 격리 피질 ( &quot;가격을 내리다&quot;) 외상후 epileptogenesis의 효율적인 동물 모델입니다. 여기 소설 수술 장치를 만들고이 모델을 생성하기 위해보다 정확하고 일관성있는 병변을 만들기 위해 그것을 사용하는 방법을 보여줍니다.

설치류에서 외상후 Epileptogenesis의 아래 부분을 잘라냄 모델 준비

Partially isolated cortex ("undercut") is an animal model of posttraumatic epileptogenesis. The surgical procedure involves cutting through the ...

preparing-undercut-model-of-posttraumatic-epileptogenesis-in-rodents

Research

JoVE Journal

Neuroscience

9.3K 조회수.  Indiana University School of Medicine. 부분적으로 격리 피질 ( "가격을 내리다") 외상후 epileptogenesis의 효율적인 동물 모델입니다. 여기 소설 수술 장치를 만들고이 모델을 생성하기 위해보다 정확하고 일관성있는 병변을 만들기 위해 그것을 사용하는 방법을 보여줍니다.

비디오: Preparing Undercut Model of Posttraumatic Epileptogenesis in Rodents

Habituation and Prepulse Inhibition of Acoustic Startle in Rodents

The acoustic startle response is a protective response, elicited by a sudden and intense acoustic stimulus. Facial and skeletal muscles are activated within a few milliseconds, leading to a whole body flinch in rodents1. Although startle responses are reflexive responses that can be reliably elicited, they are not stereotypic. They can be modulated by emotions such as fear (fear potentiated startle) and joy (joy attenuated startle), by non-associative learning processes such as habituation and sensitization, and by other sensory stimuli through sensory gating processes (prepulse inhibition), turning startle responses into an excellent tool for assessing emotions, learning, and sensory gating, for review see 2, 3. The primary pathway mediating startle responses is very short and well described, qualifying startle also as an excellent model for studying the underlying mechanisms for behavioural plasticity on a cellular/molecular level3.
We here describe a method for assessing short-term habituation, long-term habituation and prepulse inhibition of acoustic startle responses in rodents. Habituation describes the decrease of the startle response magnitude upon repeated presentation of the same stimulus. Habituation within a testing session is called short-term habituation (STH) and is reversible upon a period of several minutes without stimulation. Habituation between testing sessions is called long-term habituation (LTH)4. Habituation is stimulus specific5. Prepulse inhibition is the attenuation of a startle response by a preceding non-startling sensory stimulus6. The interval between prepulse and startle stimulus can vary from 6 to up to 2000 ms. The prepulse can be any modality, however, acoustic prepulses are the most commonly used.
Habituation is a form of non-associative learning. It can also be viewed as a form of sensory filtering, since it reduces the organisms' response to a non-threatening stimulus. Prepulse inhibition (PPI) was originally developed in human neuropsychiatric research as an operational measure for sensory gating7. PPI deficits may represent the interface of "psychosis and cognition" as they seem to predict cognitive impairment8-10. Both habituation and PPI are disrupted in patients suffering from schizophrenia11, and PPI disruptions have shown to be, at least in some cases, amenable to treatment with mostly atypical antipsychotics12, 13. However, other mental and neurodegenerative diseases are also accompanied by disruption in habituation and/or PPI, such as autism spectrum disorders (slower habituation), obsessive compulsive disorder, Tourette's syndrome, Huntington's disease, Parkinson's disease, and Alzheimer's Disease (PPI)11, 14, 15 Dopamine induced PPI deficits are a commonly used animal model for the screening of antipsychotic drugs16, but PPI deficits can also be induced by many other psychomimetic drugs, environmental modifications and surgical procedures.

Habituation and prepulse inhibition of startle are operational measures of sensory gating. Sensory gating is disrupted in schizophrenia, and some other mental disorders and neurodegenerative diseases. We here describe a standard protocol to assess short-term and long-term habituation as well as prepulse inhibition of acoustic startle responses in rats and mice.

설치류의 어쿠스틱 놀라게의 요법 이니 및 Prepulse 억제

The acoustic startle response is a protective response, elicited by a sudden and intense acoustic stimulus. Facial and skeletal muscles are activated ...

연구

신경과학

Large-scale Recording of Neurons by Movable Silicon Probes in Behaving Rodents

A major challenge in neuroscience is linking behavior to the collective activity of neural assemblies. Understanding of input-output relationships of neurons and circuits requires methods with the spatial selectivity and temporal resolution appropriate for mechanistic analysis of neural ensembles in the behaving animal, i.e. recording of representatively large samples of isolated single neurons. Ensemble monitoring of neuronal activity has progressed remarkably in the past decade in both small and large-brained animals, including human subjects1-11. Multiple-site recording with silicon-based devices are particularly effective because of their scalability, small volume and geometric design.
Here, we describe methods for recording multiple single neurons and local field potential in behaving rodents, using commercially available micro-machined silicon probes with custom-made accessory components. There are two basic options for interfacing silicon probes to preamplifiers: printed circuit boards and flexible cables. Probe supplying companies (<a href="http://www.neuronexustech.com/" >http://www.neuronexustech.com/</a>; <a href="http://www.sbmicrosystems.com/">http://www.sbmicrosystems.com/</a>; <a href="http://www.acreo.se/">http://www.acreo.se/</a>) usually provide the bonding service and deliver probes bonded to printed circuit boards or flexible cables. Here, we describe the implantation of a 4-shank, 32-site probe attached to flexible polyimide cable, and mounted on a movable microdrive. Each step of the probe preparation, microdrive construction and surgery is illustrated so that the end user can easily replicate the process.

We describe methods for large-scale recording of multiple single units and local field potential in behaving rodents with silicon probes. Drive fabrication, probe attachment to the drive and probe implantation processes are illustrated in sufficient details for easy replication.

설치류를 행동에 이동 가능한 실리콘 프로브에 의한 뉴런의 대규모 녹화

A major challenge in neuroscience is linking behavior to the collective activity of neural assemblies. Understanding of input-output relationships of ...

A Low Cost Setup for Behavioral Audiometry in Rodents

In auditory animal research it is crucial to have precise information about basic hearing parameters of the animal subjects that are involved in the experiments. Such parameters may be physiological response characteristics of the auditory pathway, e.g. via brainstem audiometry (BERA). But these methods allow only indirect and uncertain extrapolations about the auditory percept that corresponds to these physiological parameters. To assess the perceptual level of hearing, behavioral methods have to be used. A potential problem with the use of behavioral methods for the description of perception in animal models is the fact that most of these methods involve some kind of learning paradigm before the subjects can be behaviorally tested, e.g. animals may have to learn to press a lever in response to a sound. As these learning paradigms change perception itself 1,2 they consequently will influence any result about perception obtained with these methods and therefore have to be interpreted with caution. Exceptions are paradigms that make use of reflex responses, because here no learning paradigms have to be carried out prior to perceptual testing. One such reflex response is the acoustic startle response (ASR) that can highly reproducibly be elicited with unexpected loud sounds in na&iuml;ve animals. This ASR in turn can be influenced by preceding sounds depending on the perceptibility of this preceding stimulus: Sounds well above hearing threshold will completely inhibit the amplitude of the ASR; sounds close to threshold will only slightly inhibit the ASR. This phenomenon is called pre-pulse inhibition (PPI) 3,4, and the amount of PPI on the ASR gradually depends on the perceptibility of the pre-pulse. PPI of the ASR is therefore well suited to determine behavioral audiograms in na&iuml;ve, non-trained animals, to determine hearing impairments or even to detect possible subjective tinnitus percepts in these animals. In this paper we demonstrate the use of this method in a rodent model (cf. also ref. 5), the Mongolian gerbil (Meriones unguiculatus), which is a well know model species for startle response research within the normal human hearing range (e.g. 6).

A fast and inexpensive method for the behavioral determination of hearing parameters like hearing thresholds, hearing impairments or phantom perceptions (subjective tinnitus) is described. It uses pre-pulse inhibition of the acoustic startle response and can be easily implemented in a personal computer using a programmable AD/DA-converter and a piezo sensor.

쥐의 행동 Audiometry를위한 저비용 설정

In auditory animal research it is crucial to have precise information about basic hearing parameters of the animal subjects that are involved in the ...

Recording and Analysis of Circadian Rhythms in Running-wheel Activity in Rodents

When rodents have free access to a running wheel in their home cage, voluntary use of this wheel will depend on the time of day1-5. Nocturnal rodents, including rats, hamsters, and mice, are active during the night and relatively inactive during the day. Many other behavioral and physiological measures also exhibit daily rhythms, but in rodents, running-wheel activity serves as a particularly reliable and convenient measure of the output of the master circadian clock, the suprachiasmatic nucleus (SCN) of the hypothalamus. In general, through a process called entrainment, the daily pattern of running-wheel activity will naturally align with the environmental light-dark cycle (LD cycle; e.g. 12 hr-light:12 hr-dark). However circadian rhythms are endogenously generated patterns in behavior that exhibit a ~24 hr period, and persist in constant darkness. Thus, in the absence of an LD cycle, the recording and analysis of running-wheel activity can be used to determine the subjective time-of-day. Because these rhythms are directed by the circadian clock the subjective time-of-day is referred to as the circadian time (CT). In contrast, when an LD cycle is present, the time-of-day that is determined by the environmental LD cycle is called the zeitgeber time (ZT).
Although circadian rhythms in running-wheel activity are typically linked to the SCN clock6-8, circadian oscillators in many other regions of the brain and body9-14 could also be involved in the regulation of daily activity rhythms. For instance, daily rhythms in food-anticipatory activity do not require the SCN15,16 and instead, are correlated with changes in the activity of extra-SCN oscillators17-20. Thus, running-wheel activity recordings can provide important behavioral information not only about the output of the master SCN clock, but also on the activity of extra-SCN oscillators. Below we describe the equipment and methods used to record, analyze and display circadian locomotor activity rhythms in laboratory rodents.

Circadian rhythms in voluntary wheel-running activity in mammals are tightly coupled to the molecular oscillations of a master clock in the brain. As such, these daily rhythms in behavior can be used to study the influence of genetic, pharmacological, and environmental factors on the functioning of this circadian clock.

쥐에 활동 륜 실행에 녹음 및 Circadian 리듬 분석

When rodents have free access to a running wheel in their home cage, voluntary use of this wheel will depend on the time of day1-5. Nocturnal rodents, ...

Deep Brain Stimulation with Simultaneous fMRI in Rodents

In order to visualize the global and downstream neuronal responses to deep brain stimulation (DBS) at various targets, we have developed a protocol for using blood oxygen level dependent (BOLD) functional magnetic resonance imaging (fMRI) to image rodents with simultaneous DBS. DBS fMRI presents a number of technical challenges, including accuracy of electrode implantation, MR artifacts created by the electrode, choice of anesthesia and paralytic to minimize any neuronal effects while simultaneously eliminating animal motion, and maintenance of physiological parameters, deviation from which can confound the BOLD signal.&#160;Our laboratory has developed a set of procedures that are capable of overcoming most of these possible issues. For electrical stimulation, a homemade tungsten bipolar microelectrode is used, inserted stereotactically at the stimulation site in the anesthetized subject. In preparation for imaging, rodents are fixed on a plastic headpiece and transferred to the magnet bore. For sedation and paralysis during scanning, a cocktail of dexmedetomidine and pancuronium is continuously infused, along with a minimal dose of isoflurane; this preparation minimizes the BOLD ceiling effect of volatile anesthetics. In this example experiment, stimulation of the subthalamic nucleus (STN) produces BOLD responses which are observed primarily in ipsilateral cortical regions, centered in motor cortex. Simultaneous DBS and fMRI allows the unambiguous modulation of neural circuits dependent on stimulation location and stimulation parameters, and permits observation of neuronal modulations free of regional bias. This technique may be used to explore the downstream effects of modulating neural circuitry at nearly any brain region, with implications for both experimental and clinical DBS.

This protocol describes a standard method for simultaneous functional magnetic resonance imaging and deep brain stimulation in the rodent. The combined use of these experimental tools allows for the exploration of global downstream activity in response to electrical stimulation at virtually any brain target.

설치류에 동시 fMRI를 깊은 뇌 자극

In order to visualize the global and downstream neuronal responses to deep brain stimulation (DBS) at various targets, we have developed a protocol for ...

Acrylic Resin Molding Based Head Fixation Technique in Rodents

Head fixation is a technique of immobilizing animal's head by attaching a head-post on the skull for rigid clamping. Traditional head fixation requires surgical attachment of metallic frames on the skull. The attached frames are then clamped to a stationary platform resulting in immobilization of the head. However, metallic frames for head fixation have been technically difficult to design and implement in general laboratory environment. In this study, we provide a novel head fixation method. Using a custom-made head fixation bar, head mounter is constructed during implantation surgery. After the application of acrylic resin for affixing implants such as electrodes and cannula on the skull, additional resins applied on top of that to build a mold matching to the port of the fixation bar. The molded head mounter serves as a guide rails, investigators conveniently fixate the animal&#8217;s head by inserting the head mounter into the port of the fixation bar. This method could be easily applicable if implantation surgery using dental acrylics is necessary and might be useful for laboratories that cannot easily fabricate CNC machined metal head-posts.

Traditional head fixation techniques have used metallic frames attached on the skull as an interface for head fixation apparatus. This protocol demonstrates a frameless, acrylic resin molding based head fixation technique for behavioral tasks in rodents.

설치류에 아크릴 수지 성형 기반 헤드 고정 기법

Head fixation is a technique of immobilizing animal's head by attaching a head-post on the skull for rigid clamping. Traditional head fixation requires ...

Hydraulic Extrusion of the Spinal Cord and Isolation of Dorsal Root Ganglia in Rodents

Traditionally, the spinal cord is isolated by laminectomy, i.e. by breaking open the spinal vertebrae one at a time. This is both time consuming and may result in damage to the spinal cord caused by the dissection process. Here, we show how the spinal cord can be extruded using hydraulic pressure. Handling time is significantly reduced to only a few minutes, likely decreasing protein damage. The low risk of damage to the spinal cord tissue improves subsequent immunohistochemical analysis. By performing hydraulic spinal cord extrusion instead of traditional laminectomy, the rodents can further be used for DRG isolation, thereby lowering the number of animals and allowing analysis across tissues from the same rodent. We demonstrate a consistent method to identify and isolate the DRGs according to their localization relative to the costae. It is, however, important to adjust this method to the particular animal used, as the number of spinal cord segments, both thoracic and lumbar, may vary according to animal type and strain. In addition, we illustrate further processing examples of the isolated tissues.

Here, we present a protocol for hydraulic extrusion of the spinal cord as well as identification and isolation of specific dorsal root ganglia (DRGs) in the same rodent. Compared to standard spinal cord isolation methods, this method is significantly faster and reduces the risk of tissue damage.

설치류의 등쪽 뿌리 신경절의 척수 및 절연의 유압 압출

Traditionally, the spinal cord is isolated by laminectomy, i.e. by breaking open the spinal vertebrae one at a time. This is both time consuming and may ...

Transcranial Electrical Brain Stimulation in Alert Rodents

Transcranial electrical brain stimulation can modulate cortical excitability and plasticity in humans and rodents. The most common form of stimulation in humans is transcranial direct current stimulation (tDCS). Less frequently, transcranial alternating current stimulation (tACS) or transcranial random noise stimulation (tRNS), a specific form of tACS using an electrical current applied randomly within a pre-defined frequency range, is used. The increase of noninvasive electrical brain stimulation research in humans, both for experimental and clinical purposes, has yielded an increased need for basic, mechanistic, safety studies in animals. This article describes a model for transcranial electrical brain stimulation (tES) through the intact skull targeting the motor system in alert rodents. The protocol provides step-by-step instructions for the surgical set-up of a permanent epicranial electrode socket combined with an implanted counter electrode on the chest. By placing a stimulation electrode into the epicranial socket, different electrical stimulation types, comparable to tDCS, tACS, and tRNS in humans, can be delivered. Moreover, the practical steps for tES in alert rodents are introduced. The applied current density, stimulation duration, and stimulation type may be chosen depending on the experimental needs. The caveats, advantages, and disadvantages of this set-up are discussed, as well as safety and tolerability aspects.

This protocol describes a surgical set-up for a permanent epicranial electrode socket and an implanted chest electrode in rodents. By placing a second electrode into the socket, different types of transcranial electrical brain stimulation can be delivered to the motor system in alert animals through the intact skull.

경고 설치류에 Transcranial 전기 두뇌 자극

Transcranial electrical brain stimulation can modulate cortical excitability and plasticity in humans and rodents. The most common form of stimulation in ...

Cannula Implantation into the Cisterna Magna of Rodents

Cisterna magna cannulation (CMc) is a straightforward procedure that enables direct access to the cerebrospinal fluid (CSF) without operative damage to the skull or the brain parenchyma. In anesthetized rodents, the exposure of the dura mater by blunt dissection of the neck muscles allows the insertion of a cannula into the cisterna magna (CM). The cannula, composed either by a fine beveled needle or borosilicate capillary, is attached via a polyethylene (PE) tube to a syringe. Using a syringe pump, molecules can then be injected at controlled rates directly into the CM, which is continuous with the subarachnoid space. From the subarachnoid space, we can trace CSF fluxes by convective flow into the perivascular space around penetrating arterioles, where solute exchange with the interstitial fluid (ISF) occurs. CMc can be performed for acute injections immediately following the surgery, or for chronic implantation, with later injection in anesthetized or awake, freely moving rodents. Quantitation of tracer distribution in the brain parenchyma can be performed by epifluorescence, 2-photon microscopy, and magnetic resonance imaging (MRI), depending on the physico-chemical properties of the injected molecules. Thus, CMc in conjunction with various imaging techniques offers a powerful tool for assessment of the glymphatic system and CSF dynamics and function. Furthermore, CMc can be utilized as a conduit for fast, brain-wide delivery of signaling molecules and metabolic substrates that could not otherwise cross the blood brain barrier (BBB).

Here we describe a protocol to perform cisterna magna cannulation (CMc), a minimally invasive way to deliver tracers, substrates and signaling molecules into the cerebrospinal fluid (CSF). Combined with different imaging modalities, CMc enables glymphatic system and CSF dynamics assessment, as well as brain-wide delivery of various compounds.

설치류의 시스테 마그나에 정 맥 주입

Cisterna magna cannulation (CMc) is a straightforward procedure that enables direct access to the cerebrospinal fluid (CSF) without operative damage to ...

Intracranial Pharmacotherapy and Pain Assays in Rodents

Pain is a salient sensory experience with affective and cognitive dimensions. However, central mechanisms for pain remain poorly understood, hindering the development of effective therapeutics. Intracranial pharmacology presents an important tool for understanding the molecular and cellular mechanisms of pain in the brain, as well as for novel treatments. Here we present a protocol that integrates intracranial pharmacology with pain behavior testing. Specifically, we show how to infuse analgesic drugs into a select brain region, which may be responsible for pain modulation. Furthermore, to determine the effect of the candidate drug in the central nerve system, pain assays are performed after intracranial treatment. Our results demonstrate that intracranial administration of analgesic drugs in a targeted region can provide relief of pain in rodents. Thus, our protocol successfully demonstrates that intracranial pharmacology, combined with pain behavior testing, can be a powerful tool for the study of pain mechanisms in the brain.

Here we present a protocol to perform intracranial pharmacological experiments followed by pain behavior assays in rodents. This protocol allows researchers to deliver molecular and cellular targets in the brain, for pharmacologic agents in the treatment of pain.

Pain is a salient sensory experience with affective and cognitive dimensions. However, central mechanisms for pain remain poorly understood, hindering the ...