저자는 Drs를 인정하고 싶습니다. DMT 압력 myograph 및 박사 부부의 용 폴 황과 드미트리 Atochin. 높은 지방 다이어트 또는 표준 식단에 공급 쥐를 제공 Binglan 유와 종 레이. 자금 출처 이 작품은 하버드 의과 대학 (ES로 매입)에서 의학 학자 과학자 개발 미국 심장 협회 (ES로 매입)에서 부여 10SDG2610313, 그리고 엘레 및 마일 쇼어 50 주년 기념 친목 프로그램에 의해 지원되었다.

1. 용액의 조제 <ol><li> 500X EDTA (에틸렌 다이아 민 테트라 초산) 재고 : 500 MG EDTA-나 2 • 2H 2 O의 무게를 50 ML 탈 이온수에 용해. 상온에서 보관하십시오. </li><li> KCl을이 솔루션을 탈분극. <ol><li> 3.69 g의 NaCl, 18.64 g KCl을, 0.36 g 황산 마그네슘 무수, 0.41 g KH 2 PO 4, 0.46 g 염화칼슘 2 • 2H 2 O : K10X 재고 솔루션을 준비합니다 250 ML 탈 이온수에 용해. 상온에서 보관하십시오. </li><li> 100 ML 최종 KCl을이 솔루션 1X를 탈분극의 경우 : 0.21 NaHCO3를, 90 ML 탈 이온수에 0.18 g의 포도당을 녹여. K10X 원액 10 ㎖를 추가합니다. 500X EDTA 용액 95 μl를 추가합니다. 잘 섞는다. 4 ° C.에 저장 일주일 이내에 사용하십시오. </li></ol></li><li> 실험의 날 (1 리터) 신선한 HEPES-PSS 솔루션을 준비합니다. <ol><li> 6.96 g의 NaCl, 0.35 g KCl을, 0.288 g 황산 마그네슘 • 7H 2 O, 0.1605 g KH 2 PO 4, 2.38 g의 HEPES. 100 ㎖에 용해탈 이온수. 플라스크 A.로 표시 </li><li> 0.312 g 염화칼슘 2 • 2H 2 O 100 ML 탈 이온수에 용해. 플라스크 B.로 표시 </li><li> 1.25 g NaHCO3를, 0.991 g 포도당. 98 ML 탈 이온수에 용해. 플라스크 C.로 표시 </li></ol></li><li> 플라스크 C.이 큰 플라스크에 700 ML 탈 이온수을 가지고있는 500X EDTA 2 ㎖를 추가합니다. 연속 소용돌이로 교반 큰 플라스크에 플라스크 A, B 및 C의 내용을 붓는다. 수산화 나트륨과 7.4 산도를 조정합니다. </li></ol> 2. 약물 사용 <ol><li> 페닐 (10 -2 M) 10 ㎖ 탈 이온수 20.37 밀리그램을 녹인다. -80에서 나누어지는 1 ML 및 저장 ° C. 직렬 실험의 날에 10 -3, 10 -4, 10 -5, 10 -6 몰 / L를 얻기 위해 희석. </li><li> 아세틸 콜린 (10 -2 M) 10 ㎖ 탈 이온수 18.17 밀리그램을 녹인다. -80에서 나누어지는 1 ML 및 저장 ° C. 직렬 10 -3, 10 -4, 10 -5, 10 -6을 얻기 위해 희석몰 / 실험의 날에 L. </li></ol> 3. 마우스 남성 WT와 sGCα 1 - / - S6와 B6 배경에 마우스는이 연구를 통해 조사 하였다. sGCα 1 - / - 기술 이전에 4,5로 생성되었다. 주택과 실험 동물 (쥐)와 관련된 절차는 매사 추세 츠 종합 병원, 보스톤, MA의 연구 동물 관리에 분과위원회에 의해 승인되었습니다. 4. 장간막 동맥의 혈관 반응성의 측정 <ol><li> 와 쥐를 안락사 펜 토바 비탈 (200 ㎎ / kg, IP). 복강을 열고 장간막 조직을 해부. 분리 및 결합과 지방 조직의 무료 3 차 주문 장간막 동맥을 해부하고 얼음처럼 차가운에 HEPES-PSS 95 % O 2 / 5 %, 15 분 CO 2와 사전 평형 솔루션입니다. 동맥 배치 아니 장간막에서 분기 2 ~ 3 mm 길이의 고리를 잘라동맥. 혈관에는 혈액이 없을 때 마이크로 배 수준에서 동맥과 정맥을 구별하는 것은 문제가있다. 그것을 밖으로 해부하기 전에 여전히 동물에 그대로있는 동안 따라서, 우리는 용기를 식별하는 것이 좋습니다. </li><li> 입구와 출구 밸브를 통해 10 ML의 주사기의 도움으로 HEPES-PSS 용액을 압력 myograph 챔버 (DMT 모델 110P &amp; 11P 버전 1.31, 그림 2)의 유리 캐 뉼러를 입력합니다. 과도한 압력이 캐 뉼러에 연결된 깨지기 쉬운 센서를 손상시킬 수 있으므로 정맥 충만 부드럽게하고 신중하게 수행해야합니다. 작성 후 캐 뉼러 단단히 입구와 출구 밸브를 모두 닫습니다. </li><li> 오른쪽 정맥 위에 용기의 한쪽 끝을 탑재하고주의 깊게 나일론 봉합사의 한 미세 가닥을 묶어. 주사기의 도움으로, 세척 및 흡입 밸브를 통해 HEPES 용액을 용기에 채우십시오. 가까운 용기에 왼쪽 정맥에 가져와 그것에 용기의 다른 쪽 끝을 마운트합니다. 나일론 봉합사로 묶어. 채우기챔버까지 HEPES 용액을 10 ㎖로와. 조심스럽게 주사기의 도움으로 유입 밸브를 통해 HEPES 용액을 눌러 누출을 확인합니다. </li><li> myograph에와 비디오 카메라에 따라 챔버를 설치합니다. 챔버에서 산소와 열 (37 ° C)를 시작합니다. 밸브가 닫혀있을 때, 첫 번째 HEPES 솔루션 저수지 튜브 P1과 급수 밸브를 연결합니다. 관에는 거품이 없어 질 때까지 튜브를 통해 모든 거품과 솔루션 패스를 할 수 있습니다. 밸브를 엽니 다. </li><li> 관 P2 압력 레귤레이터 오는 챔버의 왼쪽 밸브를 연결합니다. 모든 거품을 다시 확인합니다. 전체 시스템의 모든 거품이나 누설이 없어야합니다. </li><li> 묘 인터페이스 패널이나 소프트웨어의 압력 메뉴로 이동 흐름을 돌리는 동안 펌프를 켭니다. </li><li> 프로그램을 열고 COLLECT을 클릭합니다. 용기는 이제 화면 (그림 3)에서 볼 수 있어야합니다. 배는 monitori을 허용 현미경에 장착 된 카메라로 시각화NG와 혈관 내강, 혈관의 직경 및 두께 분석. </li><li> 점차적으로 묘 인터페이스 패널이나 프로그램 P1 압력을 선택하여 P1 밸브를 통해 interluminal 압력을 높이는 다음과 같이 압력 값을 입력, 5 mmHg로 → 10 → 20 → 40 → 60 mmHg로. 장력을 조정하거나 수직 또는 세로 micropositioner (그림 2)의 도움으로 따라 변형, 배는 언젠가 비틀거나 압력이 증가하면서 나선상하는 경향이있다. 60 mmHg로에서 선박 평형을 37 ° C 45 분 이상. 평형 동안 예열 HEPES 한 번 목욕 솔루션을 변경합니다. </li><li> KCl을 탈분극 용액 10 mL를 적용, 완전히 평활근 세포를 탈분극 최대 수축을 달성하기 위해 목욕에, 37 ° C에서 prewarmed. 안정 수축 후 HEPES 용액 3 회 매 10 분에 목욕을 씻어. </li><li> 안정과 재생산에 의해 내피 세포의 혈관 및 무결성의 가능성을 확인페닐의 추가 (10 -5 M) 및 아세틸 콜린 ible 반응 (10 -5 M). HEPES 용액 10 분 간격으로 3 번 씻어. </li><li> (10 -5 M) 페닐 추가 선박을 preconstrict하고, 안정적인 수축을 얻을되었을 때, 아세틸 콜린의 증가 복용 (10-9, 3 × 10의 연속 추가로 누적 농도 혈관 확장 반응 곡선 (CRC)을 수행 - 9, 10 8, 3 × 10 -8, 10 7, 3 × 10 -7, 10-6, 3 × 10 -6, 10 -5 M). 마지막 투여 후, HEPES 용액으로 3 회 새로운 CRC를 시작하기 전에 매 10 분 헹구십시오. </li><li> 2 밀리미터 EGTA을 포함하는 2 + 무료 PSS에게 CA를 적용하여 수동 내강의 직경을 결정합니다. 모든 매개 변수를 계산하는 데 사용되는 각 용량 반응 (그림 4)에 대한 기록 트레이싱 측정 루멘 직경에서. </li><li> 퍼크로 모두 아세틸 콜린 이완 반응을 표현페닐의 preconstriction 후 내강 직경 entage 변화는 다음 방정식을 사용하여, 페닐 수축 후 칼슘이없는 직경 직경의 차이와 비교 : </li><li> 백분율 팽창 = 100 % X [(D X - D I) / (D 캘리포니아 자유로운 - D I)], D는 측정 된 루멘 지름 및 X, I, 그리고 캘리포니아 자유로운 작용제의 각 복용량 동맥 직경을 나타냅니다 (X), 초기 직경 다음은 (I) 수축 페닐 및 CA 2 + 프리 버퍼 (CA-무료). </li></ol> 5. 통계 분석 모든 연속적인 측정은 평균 ± SEM을로 표현됩니다. 장간막 동맥 혈관 반응이 반복 - 조치 양방향 ANOVA로 분석된다. 모든 경우에, P는 &lt;0.05를 통계적으로 유의 간주됩니다.

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저자는 공개 아무것도 없어.

마우스는 부분 때문에함으로써 인간의 병태 생리에 대한 마우스 모델을 생성하는 유전자 변형을 소개 할 수있는 가능성, 많은 연구자에 대한 선택의 실험적인 모델입니다. 혈관 활성 작은 저항의 상태가 아니라 더 큰 도관 혈관은 주로 혈관 시스템 (11)을 통해 혈액의 흐름의 조절을 정의합니다. 같은 쥐 같은 작은 동물에 저항 동맥의 크기는 미세 혈관 기능을 연구하는 와이어 myograph의 사...

압력 myograph 시스템은 작은 동맥, 정맥 등 혈관의 생리적 기능과 특성을 측정하는 데 사용됩니다. 동맥이나 정맥의 손상 작은 세그먼트는 두 개의 작은 유리 캐 뉼러에 장착하고 용기 생체 특성 (그림 1과 2)에있는 그것의 대부분을 유지할 수 있도록 적절한 내강의 압력으로 가압된다. 압력 myograph에 가까운 생리적 조건은 고립 된 혈관의 고유 특성 (예를 들어, 근원 성 톤)의 조사를 허용, 혈관 침대의 생체 내 행동을 반영합니다. 근육 수축은 힘 센서 1에 직접 기계적 커플 링에 의해 평가되는 와이어 myography, 이상 압력 myography의 장점 중 일부는 반면, 혈관 시스템 개발의 전체 저항을 정의하는 마이크로 저항 동맥, 공부 할 수있는 (I) 포함 와이어 myograph 더 큰 도관 동맥, (ⅱ) t로 제한됩니다더 와이어 (ⅲ) 선박의 자연 형태가 잘 유지되는, 혈관 내강을 통과 할 필요가 없기 때문에 내피 세포를 손상하는 모자의 위험이 감소되고, (IV)는 용기 치수는 폭 넓은 범위에서 공부하실 수 있습니다 압력이나 전단 응력 2. 마이크로 혈관을 공부하는 것은 고혈압과 같은 심혈관 질환의 변형 된 혈관 톤에 기여하는 병태 생리 및 분자 메커니즘을 이해하는 데 도움이 큰 도관 동맥을 공부하는 것보다 더 많은 정보가 될 수 있습니다. 예를 들어, 8 주 동안 쥐에게 고지방식이를 공급과 관련된 내피 세포 기능의 손상이 있지만 대동맥 고리에 2 차 주문 장간막 동맥 3 (그림 3)에서 설명 될 수있다. 압력 myography의 또 다른 장점은 가압 용기의 본질적인 근원 성 수축이 있는지, 그리고이 현상의 내피 세포의 역할과 기능을 연구 할 수있다. 여기, 우리는 DESCR장애인 산화 질소 (NO) cGMP의 신호의 설정에서 마우스 장간막 3 차 주문 저항 동맥의 혈관 반응성을 연구하는 압력 myograph의 사용을 IBE.

<table border="1">
		<tbody>
		 <tr>
			<td>Name</td>
			<td>Company</td>
			<td>Cataloge No.</td>
		 </tr>
		 <tr>
			<td>NaCl</td>
			<td>Fisher Scientific</td>
			<td>BP358</td>
		 </tr>
		 <tr>
			<td>CaCl2 (2H2O)</td>
			<td>Fisher Scientific</td>
			<td>C79-500</td>
		 </tr>
		 <tr>
			<td>KCl</td>
			<td>Sigma</td>
			<td>P9333</td>
		 </tr>
		 <tr>
			<td>MgSO4 </td>
			<td>Fisher Scientific</td>
			<td>M65-500</td>
		 </tr>
		 <tr>
			<td>KH2PO4 </td>
			<td>Sigma</td>
			<td>P3786</td>
		 </tr>
		 <tr>
			<td>NaHCO3 </td>
			<td>Fisher Scientific</td>
			<td>BP328</td>
		 </tr>
		 <tr>
			<td>NaOH</td>
			<td>Fisher Scientific</td>
			<td>S318</td>
		 </tr>
		 <tr>
			<td>D-Glucose</td>
			<td>Sigma</td>
			<td>G8270</td>
		 </tr>
		 <tr>
			<td>EDTA</td>
			<td>Fisher Scientific</td>
			<td>BP121</td>
		 </tr>
		 <tr>
			<td>HEPES</td>
			<td>Sigma</td>
			<td>H3375</td>
		 </tr>
		 <tr>
			<td>Phenylephrine</td>
			<td>Acros Organics</td>
			<td>AC20724</td>
		 </tr>
		 <tr>
			<td>Acetylcholine</td>
			<td>Sigma</td>
			<td>A6625</td>
		 </tr>
		 <tr>
			<td>Pressure Myograph System</td>
			<td>DMT</td>
			<td></td>
		 </tr>
		</tbody>
 </table>

assessing murine resistance artery function using pressure myography

압력 myograph 시스템은 작은 동맥의 기능 평가에 절묘하게 유용에 적합한 전층 압력으로 가압. 압력 myography에서 달성 근처 생리적 조건은 혈관 침대의 생체 내 행동을 추정 할 수있는 약물 및 생리적 자극에 고유 반응의 심도 특성을 허용합니다. 압력 myograph은 기존의 와이어 myographs에 비해 여러 가지 장점이 있습니다. 예를 들어, 작은 저항 혈관 엄격하게 제어 및 생리학 관련 강내 압력에서 공부하실 수 있습니다. 여기, 우리는 아세틸 콜린에 대한 응답으로 혈관 이완에 페닐와 preconstricted 3 차 주문 장간막 동맥 (긴 3-4밀리미터)의 기능을 공부합니다. 장간막 동맥은 60 mmHg로 일정한 압력으로 유지되는 압력 및 봉인 시스템에 연결된 두 개의 캐 뉼러에 장착되어 있습니다. 혈관의 루멘 및 외경 continuou 있습니다교활한는 각각 페닐 아세틸 콜린 님의 질문에 답변 혈관 수축과 혈관 이완의 실시간 정량을 허용 비디오 카메라를 사용하여 촬영. 심혈관 질환의 원인을 연구하는 압력 myography의 적용을 설명하기 위해, 우리는 전신 고혈압의 쥐 모델에서 내피 세포 의존적 혈관 기능을 평가 하였다. 수용성 구아닐산의 cyclase의 (- / - sGCα 1)의 α 1 소단위의 결핍 생쥐 때 129S6 (S6) 배경 (sGCα 1 -/-S6)에 고혈압하지만하지 않는 경우 C57BL / 6 (B6) 배경 (sGCα에 1 -/-B6). 압력 myography를 사용하여, 우리는 보여 저해하는 내피 세포 의존적 혈관 이완에 sGCα 1 결핍이 발생합니다. 혈관 장애는 sGCα 1보다 sGCα 1 -/-S6에서 더 뚜렷하다 -/-B6 마우스, 가능성 contributisGCα 1 -/-B6 마우스에 비해 sGCα 1 -/-S6의 높은 혈압 NG. 압력 myography 그로 인하여 심혈관 질환을 기본 메커니즘에 대한 우리의 통찰력을 보강, 혈관 수축과 이완에 다양한 자극의 효과를 평가하는 데 사용할 수있는 비교적 간단하지만, 민감하고 기작 유용한 기술이다.

NO는 중앙 인간 6 및 동물 모델 7 모두에서 혈압 항상성 유지에 관여하지 않습니다. 혈관 평활근의 이완을 제어 할 수있는 능력이 용해 구아닐산 cyclase의 (SGC), cGMP의 8 생성 헴 함유 heterodimeric 효소에 의해 매개된다. 최근에는 혈압 - 수정 유전자 변형 인간 9 혈압 조절에 SGC의 관련성을 보여주는 sGCα 1 sGCβ 1 유전자를 포함하는 현장에서 발견되었다. ?...

Watch this Scientific Journal Video about Assessing Murine Resistance Artery Function Using Pressure Myography at JoVE.com

Assessing Murine Resistance Artery Function Using Pressure Myography

압력 myography에서 혈관의 손상 작은 세그먼트는 두 개의 작은 캐 뉼러에 장착하고 적절한 내강 압력으로 가압. 여기, 우리는 마우스 3의 혈관 이완 반응을 측정하는 방법을 설명<sup&gt; 회</supC57와 sGCα에&gt; 주문 장간막 동맥<sub&gt; 1</sub<sup&gt; - / -</sup&gt; 압력 myography를 사용하여 마우스.

압력 Myography를 사용 쥐과 저항 동맥의 기능을 평가

Pressure myograph systems are  exquisitely useful in the functional assessment of small arteries, pressurized  to a suitable transmural pressure. The  ...

assessing-murine-resistance-artery-function-using-pressure-myography

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JoVE Journal

Biology

21.9K 조회수.  Massachusetts General Hospital, Harvard Medical School. 압력 myography에서 혈관의 손상 작은 세그먼트는 두 개의 작은 캐 뉼러에 장착하고 적절한 내강 압력으로 가압. 여기, 우리는 마우스 3의 혈관 이완 반응을 측정하는 방법을 설명 회 주문 장간막 동맥 1

비디오: Assessing Murine Resistance Artery Function Using Pressure Myography

Measuring Blood Pressure in Mice using Volume Pressure Recording, a Tail-cuff Method

The CODA 8-Channel High Throughput Non-Invasive Blood Pressure system measures the blood pressure in up to 8 mice or rats simultaneously. The CODA tail-cuff system uses Volume Pressure Recording (VPR) to measure the blood pressure by determining the tail blood volume. A specially designed differential pressure transducer and an occlusion tail-cuff measure the total blood volume in the tail without the need to obtain the individual pulse signal. Special attention is afforded to the length of the occlusion cuff in order to derive the most accurate blood pressure readings. VPR can easily obtain readings on dark-skinned rodents, such as C57BL6 mice and is MRI compatible. The CODA system provides you with measurements of six (6) different blood pressure parameters; systolic and diastolic blood pressure, heart rate, mean blood pressure, tail blood flow, and tail blood volume. Measurements can be made on either awake or anesthetized mice or rats. The CODA system includes a controller, laptop computer, software, cuffs, animal holders, infrared warming pads, and an infrared thermometer. There are seven different holder sizes for mice as small as 8 grams to rats as large as 900 grams.

The CODA 8-Channel High Throughput Non-Invasive Blood Pressure system measures the blood pressure in up to 8 mice or rats simultaneously. This tail-cuff system uses Volume Pressure Recording (VPR) to measure the blood pressure by determining the tail blood volume.

볼륨 압력 녹음, 꼬리 즉석 방법을 사용하여 마우스의 측정 혈압

The CODA 8-Channel High Throughput Non-Invasive Blood Pressure system measures the blood pressure in up to 8 mice or rats simultaneously. The CODA ...

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생물학

Using the optokinetic response to study visual function of zebrafish

Optokinetic response (OKR) is a behavior that an animal vibrates its eyes to follow a rotating grating around it. It has been widely used to assess the visual functions of larval zebrafish1-5. Nevertheless, the standard protocol for larval fish is not yet readily applicable in adult zabrafish. Here, we introduce how to measure the OKR of adult zebrafish with our simple custom-built apparatus using a new protocol which is established in our lab. Both our apparatus and step-by-step procedure of OKR in adult zebrafish are illustrated in this video. In addition, the measurements of the larval OKR, as well as the optomotor response (OMR) test of adult zebrafish, are also demonstrated in this video. This OKR assay of adult zebrafish in our experiment may last for up to 4 hours. Such OKR test applied in adult fish will benefit to visual function investigation more efficiently when the adult fish vision system is manipulated.
Su-Qi Zou and Wu Yin contributed equally to this paper.

Optokinetic response has been widely used to assess the visual functions of larval zebrafish. Nevertheless, the standard protocol for larval fish is not yet readily applicable in adults1-5. Here, we introduce how to measure the OKR of adult zebrafish using a new protocol which is established in our lab.

zebrafish의 시각 기능을 연구하기 위해 optokinetic 응답을 사용하여

Optokinetic response (OKR) is a behavior that an animal vibrates its eyes to follow a rotating grating around it. It has been widely used to assess the ...

Bioassays for Monitoring Insecticide Resistance

Pest resistance to pesticides is an increasing problem because pesticides are an integral part of high-yielding production agriculture. When few products are labeled for an individual pest within a particular crop system, chemical control options are limited. Therefore, the same product(s) are used repeatedly and continual selection pressure is placed on the target pest. There are both financial and environmental costs associated with the development of resistant populations. The cost of pesticide resistance has been estimated at approximately $ 1.5 billion annually in the United States. This paper will describe protocols, currently used to monitor arthropod (specifically insects) populations for the development of resistance. The adult vial test is used to measure the toxicity to contact insecticides and a modification of this test is used for plant-systemic insecticides. In these bioassays, insects are exposed to technical grade insecticide and responses (mortality) recorded at a specific post-exposure interval. The mortality data are subjected to Log Dose probit analysis to generate estimates of a lethal concentration that provides mortality to 50% (LC50) of the target populations and a series of confidence limits (CL's) as estimates of data variability. When these data are collected for a range of insecticide-susceptible populations, the LC50 can be used as baseline data for future monitoring purposes. After populations have been exposed to products, the results can be compared to a previously determined LC50 using the same methodology.

This manuscript demonstrates and discusses techniques used to survey pesticide susceptibility and detect resistance to contact and systemic pesticides in arthropod pests.

모니터링 살충제 저항 Bioassays

Pest resistance to pesticides is an increasing problem because pesticides are an integral part of high-yielding production agriculture. When few products ...

Exploring Arterial Smooth Muscle Kv7 Potassium Channel Function using Patch Clamp Electrophysiology and Pressure Myography

Contraction or relaxation of smooth muscle cells within the walls of resistance arteries determines the artery diameter and thereby controls flow of blood through the vessel and contributes to systemic blood pressure. The contraction process is regulated primarily by cytosolic calcium concentration ([Ca2+]cyt), which is in turn controlled by a variety of ion transporters and channels. Ion channels are common intermediates in signal transduction pathways activated by vasoactive hormones to effect vasoconstriction or vasodilation. And ion channels are often targeted by therapeutic agents either intentionally (e.g. calcium channel blockers used to induce vasodilation and lower blood pressure) or unintentionally (e.g. to induce unwanted cardiovascular side effects).
Kv7 (KCNQ) voltage-activated potassium channels have recently been implicated as important physiological and therapeutic targets for regulation of smooth muscle contraction. To elucidate the specific roles of Kv7 channels in both physiological signal transduction and in the actions of therapeutic agents, we need to study how their activity is modulated at the cellular level as well as evaluate their contribution in the context of the intact artery.
The rat mesenteric arteries provide a useful model system. The arteries can be easily dissected, cleaned of connective tissue, and used to prepare isolated arterial myocytes for patch clamp electrophysiology, or cannulated and pressurized for measurements of vasoconstrictor/vasodilator responses under relatively physiological conditions. Here we describe the methods used for both types of measurements and provide some examples of how the experimental design can be integrated to provide a clearer understanding of the roles of these ion channels in the regulation of vascular tone.

Measurements of Kv7 (KCNQ) potassium channel activity in isolated arterial myocytes (using patch clamp electrophysiological techniques) in parallel with measurements of constrictor/dilator responses (using pressure myography) can reveal important information about the roles of Kv7 channels in vascular smooth muscle physiology and pharmacology.

패치 클램프 전기 생리학 및 압력 Myography를 사용하여 간선 평활근 Kv7 칼륨 채널 기능을 탐색

Contraction  or relaxation of smooth muscle cells within the walls of resistance arteries  determines the artery diameter and thereby controls flow of ...

RNA Secondary Structure Prediction Using High-throughput SHAPE

Understanding the function of RNA involved in biological processes requires a thorough knowledge of RNA structure. Toward this end, the methodology dubbed "high-throughput selective 2' hydroxyl acylation analyzed by primer extension", or SHAPE, allows prediction of RNA secondary structure with single nucleotide resolution. This approach utilizes chemical probing agents that preferentially acylate single stranded or flexible regions of RNA in aqueous solution. Sites of chemical modification are detected by reverse transcription of the modified RNA, and the products of this reaction are fractionated by automated capillary electrophoresis (CE). Since reverse transcriptase pauses at those RNA nucleotides modified by the SHAPE reagents, the resulting cDNA library indirectly maps those ribonucleotides that are single stranded in the context of the folded RNA. Using ShapeFinder software, the electropherograms produced by automated CE are processed and converted into nucleotide reactivity tables that are themselves converted into pseudo-energy constraints used in the RNAStructure (v5.3) prediction algorithm. The two-dimensional RNA structures obtained by combining SHAPE probing with in silico RNA secondary structure prediction have been found to be far more accurate than structures obtained using either method alone.

High-throughput selective 2' hydroxyl acylation analyzed by primer extension (SHAPE) utilizes a novel chemical probing technology, reverse transcription, capillary electrophoresis and secondary structure prediction software to determine the structures of RNAs from several hundred to several thousand nucleotides at single nucleotide resolution.

높은 처리량 SHAPE를 사용하여 RNA 이차 구조 예측

Understanding the function of RNA involved in biological processes requires a thorough knowledge of RNA structure. Toward this end, the methodology dubbed ...

Assessing Cerebral Autoregulation via Oscillatory Lower Body Negative Pressure and Projection Pursuit Regression

The process by which cerebral perfusion is maintained constant over a wide range of systemic pressures is known as &#8220;cerebral autoregulation.&#8221; Effective dampening of flow against pressure changes occurs over periods as short as ~15 sec and becomes progressively greater over longer time periods. Thus, slower changes in blood pressure are effectively blunted and faster changes or fluctuations pass through to cerebral blood flow relatively unaffected. The primary difficulty in characterizing the frequency dependence of cerebral autoregulation is the lack of prominent spontaneous fluctuations in arterial pressure around the frequencies of interest (less than ~0.07 Hz or ~15 sec). Oscillatory lower body negative pressure (OLBNP) can be employed to generate oscillations in central venous return that result in arterial pressure fluctuations at the frequency of OLBNP. Moreover, Projection Pursuit Regression (PPR) provides a nonparametric method to characterize nonlinear relations inherent in the system without a priori assumptions and reveals the characteristic non-linearity of cerebral autoregulation. OLBNP generates larger fluctuations in arterial pressure as the frequency of negative pressure oscillations become slower; however, fluctuations in cerebral blood flow become progressively lesser. Hence, the PPR shows an increasingly more prominent autoregulatory region at OLBNP frequencies of 0.05 Hz and below (20 sec cycles). The goal of this approach it to allow laboratory-based determination of the characteristic nonlinear relationship between pressure and cerebral flow and could provide unique insight to integrated cerebrovascular control as well as to physiological alterations underlying impaired cerebral autoregulation (e.g., after traumatic brain injury, stroke, etc.).

Cerebral perfusion is maintained across a range of pressures via cerebral autoregulation. However, characterizing autoregulation requires prominent pressure fluctuations at regulated frequencies. The described protocol will show how oscillatory lower body negative pressure can generate pressure fluctuations to provide data for projection pursuit regression for quantification of the autoregulatory curve.

진 동성 하체 부정적인 압력 및 프로젝션 추구 회귀 분석을 통해 뇌 혈류의자가 평가

The process by which cerebral perfusion is maintained constant over a wide range of systemic pressures is known as &#8220;cerebral autoregulation.&#8221; ...

Measuring Respiratory Function in Mice Using Unrestrained Whole-body Plethysmography

Respiratory dysfunction is one of the leading causes of morbidity and mortality in the world and the rates of mortality continue to rise. Quantitative assessment of lung function in rodent models is an important tool in the development of future therapies. Commonly used techniques for assessing respiratory function including invasive plethysmography and forced oscillation. While these techniques provide valuable information, data collection can be fraught with artefacts and experimental variability due to the need for anesthesia and/or invasive instrumentation of the animal. In contrast, unrestrained whole-body plethysmography (UWBP) offers a precise, non-invasive, quantitative way by which to analyze respiratory parameters. This technique avoids the use of anesthesia and restraints, which is common to traditional plethysmography techniques. This video will demonstrate the UWBP procedure including the equipment set up, calibration and lung function recording. It will explain how to analyze the collected data, as well as identify experimental outliers and artefacts that results from animal movement. The respiratory parameters obtained using this technique include tidal volume, minute volume, inspiratory duty cycle, inspiratory flow rate and the ratio of inspiration time to expiration time. UWBP does not rely on specialized skills and is inexpensive to perform. A key feature of UWBP, and most appealing to potential users, is the ability to perform repeated measures of lung function on the same animal.

The assessment of respiratory physiology has traditionally relied upon techniques, which require restraint or sedation of the animal. Unrestrained whole-body plethysmography, however, provides precise, non-invasive, quantitative analysis of respiratory physiology in animal models. In addition, the technique allows repeated respiratory assessment of mice allowing for longitudinal studies.

무제한 전신 혈량 측정법을 사용하여 마우스의 호흡 기능을 측정

Respiratory dysfunction is one of the leading causes of morbidity and mortality in the world and the rates of mortality continue to rise. Quantitative ...

Evaluation of Zebrafish Kidney Function Using a Fluorescent Clearance Assay

The zebrafish embryo offers a tractable model to study organogenesis and model human genetic disease. Despite its relative simplicity, the zebrafish kidney develops and functions in almost the same way as humans. A major difference in the construction of the human kidney is the presence of millions of nephrons compared to the zebrafish that has only two. However, simplifying such a complex system into basic functional units has aided our understanding of how the kidney develops and operates. In zebrafish, the midline located glomerulus is responsible for the initial blood filtration into two pronephric tubules that diverge to run bilaterally down the embryonic axis before fusing to each other at the cloaca. The pronephric tubules are heavily populated by motile cilia that facilitate the movement of filtrate along the segmented tubule, allowing the exchange of various solutes before finally exiting via the cloaca2-4. Many genes responsible for CKD, including those related to ciliogenesis, have been studied in zebrafish5. However, a major draw back has been the difficulty in evaluating zebrafish kidney function after genetic manipulation. Traditional assays to measure kidney dysfunction in humans have proved non translational to zebrafish, mainly due to their aquatic environment and small size. For example, it is not physically possible to extract blood from embryonic staged fish for analysis of urea and creatinine content, as they are too small. In addition, zebrafish do not produce enough urine for testing on a simple proteinuria &#8216;dipstick&#8217;, which is often performed during initial patient examinations. We describe a fluorescent assay that utilizes the optical transparency of the zebrafish to quantitatively monitor the clearance of a fluorescent dye, over time, from the vasculature and out through the kidney, to give a read out of renal function1,6-9.

The zebrafish is a popular tool to model chronic kidney disease (CKD). However, their small size makes it impossible to evaluate renal function using traditional methods. We describe a fluorescent dye kidney clearance assay1 that allows quantitative analysis of zebrafish kidney function in CKD.

형광 정리 분석을 사용하여 Zebrafish의 신장 기능의 평가

The zebrafish embryo offers a tractable model to study organogenesis and model human genetic disease. Despite its relative simplicity, the zebrafish ...

Cardiac Pressure-Volume Loop Analysis Using Conductance Catheters in Mice

Cardiac pressure-volume loop analysis is the &#8220;gold-standard&#8221; in the assessment of load-dependent and load-independent measures of ventricular systolic and diastolic function. Measures of ventricular contractility and compliance are obtained through examination of cardiac response to changes in afterload and preload. These techniques were originally developed nearly three decades ago to measure cardiac function in large mammals and humans. The application of these analyses to small mammals, such as mice, has been accomplished through the optimization of microsurgical techniques and creation of conductance catheters. Conductance catheters allow for estimation of the blood pool by exploiting the relationship between electrical conductance and volume. When properly performed, these techniques allow for testing of cardiac function in genetic mutant mouse models or in drug treatment studies. The accuracy and precision of these studies are dependent on careful attention to the calibration of instruments, systematic conduct of hemodynamic measurements and data analyses. We will review the methods of conducting pressure-volume loop experiments using a conductance catheter in mice.

Cardiac pressure-volume loop analysis is the most comprehensive way to measure cardiac function in the intact heart. We describe a technique to perform and analyze cardiac pressure volume loops, using conductance catheters.

마우스에서 컨덕턴스 카테터를 사용하여 심장 압력 볼륨 루프 분석

Cardiac pressure-volume loop analysis is the &#8220;gold-standard&#8221; in the assessment of load-dependent and load-independent measures of ventricular ...

Intracavernosal Pressure Recording to Evaluate Erectile Function in Rodents

Erectile dysfunction (ED) is defined as the inability to attain or keep an erection of the penis, and this has become a prevalent male sexual disorder. Rodents are employed by many studies to research the physiology/pathology of erectile function. Erectile function in rodents can be evaluated by measuring the intracavernosal pressure (ICP). In practice, ICP can be monitored following electrical stimulation of the cavernous nerves (CNs). The arterial pressure of the carotid artery (the mean arterial pressure) is used as the reference for ICP. Using ICP recording protocols, many key parameters of erectile function can be measured from the ICP response curve. The ICP measurement provides more information than the apomorphine-induced penile erection test, and is cheaper than telemetric monitoring of the corpus spongiosum penis, making this method the most popular one to evaluate erectile function. However, compared to the easily-performed APO-induced erectile function test, successful ICP recordings require attention to detail, practice, and adherence to the operation method. In this work, an introduction to ICP recording in rats is provided to complement the procedure efficiently.

Intracavernosal pressure recording (ICP) is an important method to evaluate the erectile function of experimental animals. Here, a detailed protocol is demonstrated for the recording procedure of ICP by catheterizing the crura penis and then electrically stimulating the cavernous nerves in rats.

Intracavernosal 압력 기록 설치류에 발기 기능을 평가 하

Erectile dysfunction (ED) is defined as the inability to attain or keep an erection of the penis, and this has become a prevalent male sexual disorder. ...