Name: the ex vivo isolated skeletal microvessel preparation for investigation of vascular reactivity
Uploaded: 2012-04-28T14:00:00
Description: Watch this Scientific Journal Video about The ex vivo Isolated Skeletal Microvessel Preparation for Investigation of Vascular Reactivity at JoVE.com

Эта работа выполнена при финансовой поддержке Американской Ассоциации Сердца (ОВОС 0740129N) и NIH T32 HL90610.

1. До эксперимента <ol><li> До начала эксперимента день, стеклянные капилляры соответствующих размеров для станции тянут в микропипетки (горизонтальное или вертикальное съемник может использоваться). Диаметр кончика легко регулируется в зависимости от судна, изолированный, хотя мы обыч?...

<ol>
	<li>Goodwill, A. G., Frisbee, S. J., Stapleton, P. A., James, M. E., Frisbee, J. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Impact+of+Chronic+Anticholesterol+Therapy+on+Development+of+Microvascular+Rarefaction+in+the+Metabolic+Syndrome.">Impact of Chronic Anticholesterol Therapy on Development of Microvascular Rarefaction in the Metabolic Syndrome.</a> Microcirculation. , 1-18 (2009).</li><li>Goodwill, A. G., James, M. E., Frisbee, J. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Increased+vascular+thromboxane+generation+impairs+dilation+of+skeletal+muscle+arterioles+of+obese+Zucker+rats+with+reduced+oxygen+tension.">Increased vascular thromboxane generation impairs dilation of skeletal muscle arterioles of obese Zucker rats with reduced oxygen tension.</a> Am. J. Physiol. Heart Circ. 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A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Increased+myogenic+responsiveness+of+skeletal+muscle+arterioles+with+juvenile+growth.">Increased myogenic responsiveness of skeletal muscle arterioles with juvenile growth.</a> Am. J. Physiol. Heart Circ. Physiol. 294, 2344-2351 (2008).</li><li>Dacey, R. G., Duling, B. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+study+of+rat+intracerebral+arterioles:+methods,+morphology,+and+reactivity.">A study of rat intracerebral arterioles: methods, morphology, and reactivity.</a> Am. J. Physiol. Heart Circ. Physiol. 243, H598-H606 (1982).</li><li>Fredricks, K. T., Liu, Y., Lombard, J. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Response+of+extraparenchymal+resistance+arteries+of+rat+skeletal+muscle+to+reduce+PO2.">Response of extraparenchymal resistance arteries of rat skeletal muscle to reduce PO2.</a> Am. J. Physiol. 267, H706-H715 (1994).</li><li>Durand, M. J., Raffai, G., Weinberg, B. D., Lombard, J. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Angiotensin-(1-7)+and+low-dose+angiotensin+II+infusion+reverse+salt-induced+endothelial+dysfunction+via+different+mechanisms+in+rat+middle+cerebral+arteries.">Angiotensin-(1-7) and low-dose angiotensin II infusion reverse salt-induced endothelial dysfunction via different mechanisms in rat middle cerebral arteries.</a> Am. J. Physiol. Heart Circ. Physiol. 299, H1024-H1033 (2010).</li><li>LeBlanc, A. J., Cumpston, J. L., Chen, B. T., Frazer, D., Castranova, V., Nurkiewicz, T. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Nanoparticle+inhalation+impairs+endothelium-dependent+vasodilation+in+subepicardial+arterioles.">Nanoparticle inhalation impairs endothelium-dependent vasodilation in subepicardial arterioles.</a> J. Toxicol. Environ. Health A. 72, 1576-1584 (2009).</li><li>Jernigan, N. L., LaMarca, B., Speed, J., Galmiche, L., Granger, J. P., Drummond, H. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Dietary+salt+enhances+benzamil-sensitive+component+of+myogenic+constriction+in+mesenteric+arteries.">Dietary salt enhances benzamil-sensitive component of myogenic constriction in mesenteric arteries.</a> Am. J. Physiol. Heart Circ. Physiol. 294, H409-H420 (2008).</li><li>Stapleton, P. A., Goodwill, A. G., James, M. E., Frisbee, J. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Altered+mechanisms+of+endothelium-dependent+dilation+in+skeletal+muscle+arterioles+with+genetic+hypercholesterolemia.">Altered mechanisms of endothelium-dependent dilation in skeletal muscle arterioles with genetic hypercholesterolemia.</a> Am. J. Physiol. Regul. Integr. Comp. Physiol. 293, R1110-R1119 (2007).</li><li>Goodwill, A. G., Stapleton, P. A., James, M. E., d'Audiffret, A. C., Frisbee, J. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Increased+arachidonic+acid-induced+thromboxane+generation+impairs+skeletal+muscle+arteriolar+dilation+with+genetic+dyslipidemia.">Increased arachidonic acid-induced thromboxane generation impairs skeletal muscle arteriolar dilation with genetic dyslipidemia.</a> Microcirculation. 15, 621-631 (2008).</li><li>Baumbach, G. L., Hadju, M. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mechanics+and+composition+of+cerebral+arterioles+in+renal+and+spontaneously+hypertensive+rats.">Mechanics and composition of cerebral arterioles in renal and spontaneously hypertensive rats.</a> Hypertension. 21, 816-826 (1993).</li><li>Uchida, E., Bohr, D. F., Hoobler, S. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+method+for+studying+isolated+resistance+vessel+from+rabbit+mesentery+and+brain+and+their+responses+to+drugs.">A method for studying isolated resistance vessel from rabbit mesentery and brain and their responses to drugs.</a> Circ. Res. 4, 525-536 (1967).</li><li>Davis, M. J., Kuo, L., Chilian, W. M., Muller, J. M. I. s. o. l. a. t. e. d., Barker, J. H., Anderson, G. L., Menger, M. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Chapter+23.">Chapter 23.</a> Isolated, perfused microvessels. In: Clinically Applied Microcirculation Research. 32, 435-456 (1995).</li><li>Lombard, J. H., Liu, Y., Fredricks, K. T., Bizub, D. M., Roman, R. J., Rusch, N. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Electrical+and+mechanical+responses+of+rat+middle+cerebral+arterieal+to+reduced+PO2+and+prostacyclin.">Electrical and mechanical responses of rat middle cerebral arterieal to reduced PO2 and prostacyclin.</a> Am. J. Physiol. 276, H509-H516 (1994).</li></ol>

Протокол представлен описывает изоляции, удаление и двойной катетеризации из скелетных мышц сосудистой системы, хотя это общий метод может быть легко применены к большинству тканей. В текущем рукописи, термин &quot;артериол&quot; был использован авторами для описания сопротивления судна ?...

<table border="1"><tbody><tr><td> Реактивы и оборудование </td><td> Компания </td><td> Комментарии / Catalogue # </td></tr><tr><td> Судно палаты </td><td> Обычай </td><td> Дэйв Эйк (MCW) </td></tr><tr><td> Подогрев оборотной воды ванны </td><td> PolyScience и Haake </td><td> Хааке DC 10 </td></tr><tr><td> Пипец </td><td> Фредерик Haer и Ко </td><td> Капиллярные трубки диаметром 2,0 мм х 1,0 мм ID (27-33-1) </td></tr><tr><td> Контроля давления </td><td> Инструменты Всемирной Precision </td><td></td></tr><tr><td> Водяной рубашкой водохранилище </td><td> Обычай </td><td></td></tr><tr><td> Внешний источник света </td><td> Инструменты Всемирной Precision </td><td> Novaflex </td></tr><tr><td> Внесите Puller </td><td> Микроданных инструменты </td><td> PMP102 микропипеткой Puller </td></tr><tr><td> Полный дополненияМент хирургических инструментов </td><td> Изобразительных средств наук </td><td> Дюмон </td></tr><tr><td> Ultra Fine Пинцет </td><td> Изобразительных средств наук </td><td> Inox # 5 </td></tr><tr><td> Шелковой нити шовного </td><td> Ethilon </td><td> # 10-0 или 9-0 </td></tr><tr><td> Стерео микроскоп </td><td> Олимп </td><td> Olympus SZ-11 </td></tr><tr><td> Аналоговое видео Штангенциркули </td><td> Boeckeler </td><td> С помощью контроллера (Via-100) </td></tr><tr><td> Высокое разрешение аналоговой камеры </td><td> Panasonic </td><td> GP-MF 602 </td></tr><tr><td> Кислородный баллон </td><td> Региональный </td><td> 21% баланса азота и 5% CO 2, баланс азота </td></tr><tr><td> Трубы </td><td> Tygon </td><td></td></tr><tr><td> Дренажный насос </td><td> Коул Parmer машиностроительный завод </td><td></td></tr><tr><td> Измененный PSS Крысы </td><td> См. рецепт ниже </td><td></TD&gt; </tr><tr><td> Ван Breemen в релаксант PSS </td><td> См. рецепт ниже </td><td></td></tr></tbody></table> Таблица 1. Перечень основных компонентов изолированной установке станции микрососудов представлены на рисунках. <table border="1"><tbody><tr><td> Измененный Рецепт Крыса PSS </td><td> Для того, чтобы два литра PSS </td><td> 20X соль бирже (2L) </td><td> 20X буферных запасов (2 л) </td></tr><tr><td> NaCl </td><td></td><td> 278,0 г </td><td></td></tr><tr><td> KCl </td><td></td><td> 14,0 г </td><td></td></tr><tr><td> MgSO 4-7H 2 O </td><td></td><td> 11,5 г </td><td></td></tr><tr><td> CaCl 2-H 2 O </td><td></td><td> 9,4 г </td><td></td></tr><tr> &lt;TD&gt; NaHCO 3 </td><td></td><td></td><td> 80,8 г </td></tr><tr><td> ЭДТА </td><td></td><td></td><td> 0,4 г </td></tr><tr><td> NaH 2 PO 4 </td><td> 0,28 г </td><td></td><td></td></tr><tr><td> Глюкоза </td><td> 1,98 г </td><td></td><td></td></tr><tr><td> 20x соль со </td><td> 100 мл </td><td></td><td></td></tr><tr><td> 20x буферных запасов </td><td> 100 мл </td><td></td><td></td></tr><tr><td> Дистиллированная вода </td><td> 1800 мл </td><td></td><td></td></tr></tbody></table> Таблица 2. Рецепт стандартный физиологический раствор поваренной соли (PSS), используемые в изолированных протоколов микрососудов. Комментарии к рецепта: Сделайте 2 л соль бирже и 2 л буферных запасов. Они схранить в холодильнике, когда не используется, но избавиться от них хорошо, и часто перед приготовлением PSS. Дополнительные ингредиенты добавляются в момент подготовки окончательного PSS. <table border="1"><tbody><tr><td> Ван Breemen в релаксант PSS </td><td> Для того, чтобы 2 литра PSS </td><td> 20X соль бирже (1L) </td><td> 20X буферных запасов (1L) </td></tr><tr><td> NaCl </td><td></td><td> 107,4 г </td><td></td></tr><tr><td> KCl </td><td></td><td> 7,0 г </td><td></td></tr><tr><td> MgSO 4-7H 2 O </td><td></td><td> 5,76 г </td><td></td></tr><tr><td> MgCl 2 6H 2 O </td><td></td><td> 81,32 г </td><td></td></tr><tr><td> NaHCO 3 </td><td></td><td></td><td> 40,4 г </td></tr><tr><td> EDT</td><td></td><td></td><td> 0,2 г </td></tr><tr><td> EGTA </td><td></td><td></td><td> 15,22 </td></tr><tr><td> NaH 2 PO 4 </td><td> 0,28 г </td><td></td><td></td></tr><tr><td> Глюкоза </td><td> 1,98 г </td><td></td><td></td></tr><tr><td> 20x соль со </td><td> 100 мл </td><td></td><td></td></tr><tr><td> 20x буферных запасов </td><td> 100 мл </td><td></td><td></td></tr><tr><td> Дистиллированная вода </td><td> 1800 мл </td><td></td><td></td></tr></tbody></table> Таблица 3. Рецепт для расслабляет физиологические Ван Breemen в солевом растворе (PSS), используемые в изолированных протоколов микрососудов в условиях нулевой активный сигнал. Комментарии к рецепта: Сделайте 1 л соль сом 1 л буферного запаса. Их можно хранить в холодильнике, когда не используется, но избавиться от них хорошо, и часто перед приготовлением PSS. Дополнительные ингредиенты добавляются в момент подготовки окончательного релаксант PSS.

the ex vivo isolated skeletal microvessel preparation for investigation of vascular reactivity

The isolated microvessel preparation is an ex vivo preparation that allows for examination of the different contributions of factors that control vessel diameter, and thus, perfusion resistance1-5. This is a classic experimental preparation that was, in large measure, initially described by Uchida et al.15 several decades ago. This initial description provided the basis for the techniques that was extensively modified and enhanced, primarily in the laboratory of Dr. Brian Duling at the University of Virginia6-8, and we present a current approach in the following pages. This preparation will specifically refer to the gracilis arteriole in a rat as the microvessel of choice, but the basic preparation can readily be applied to vessels isolated from nearly any other tissue or organ across species9-13. Mechanical (i.e., dimensional) changes in the isolated microvessels can easily be evaluated in response to a broad array of physiological (e.g., hypoxia, intravascular pressure, or shear) or pharmacological challenges, and can provide insight into mechanistic elements comprising integrated responses in an intact, although ex vivo, tissue. The significance of this method is that it allows for facile manipulation of the influences on the integrated regulation of microvessel diameter, while also allowing for the control of many of the contributions from other sources, including intravascular pressure (myogenic), autonomic innervation, hemodynamic (e.g., shear stress), endothelial dependent or independent stimuli, hormonal, and parenchymal influences, to provide a partial list. Under appropriate experimental conditions and with appropriate goals, this can serve as an advantage over in vivo or in situ tissue/organ preparations, which do not readily allow for the facile control of broader systemic variables. 
The major limitation of this preparation is essentially the consequence of its strengths. By definition, the behavior of these vessels is being studied under conditions where many of the most significant contributors to the regulation of vascular resistance have been removed, including neural, humoral, metabolic, etc. As such, the investigator is cautioned to avoid over-interpretation and extrapolation of the data that are collected utilizing this preparation. The other significant area of concern with regard to this preparation is that it can be very easy to damage cellular components such as the endothelial lining or the vascular smooth muscle, such that variable source of error can be introduced. It is strongly recommended that the individual investigator utilize appropriate measurements to ensure the quality of the preparation, both at the initiation of the experiment and periodically throughout the course of a protocol.

Watch this Scientific Journal Video about The ex vivo Isolated Skeletal Microvessel Preparation for Investigation of Vascular Reactivity at JoVE.com

The ex vivo Isolated Skeletal Microvessel Preparation for Investigation of Vascular Reactivity

<em&gt; Бывший естественных условиях</em&gt; Подготовка описана в изоляции из крупнейших гасШз мышц сопротивление артериол для допроса и сосудистых реакций на вазоактивные стимулы и оценка основных структурных свойств с помощью пассивных механики стены.

Бывший естественных условиях Изолированный скелета Подготовка микрососудов для исследования сосудистой реактивности

The isolated  microvessel preparation is an ex vivo preparation that allows for examination of the different contributions of  factors that control vessel ...

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The ex vivo Isolated Skeletal Microvessel Preparation for Investigation of Vascular Reactivity

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BACKGROUND: Regulatory factors and detailed physiology of in vivo microcirculation have remained not fully clarified after many different modalities of ...

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Ex Vivo Assessment of Contractility, Fatigability and Alternans in Isolated Skeletal Muscles

Ex Vivo оценке сократимости, утомляемость и аНегпапз в изолированных скелетных мышц

 
Described here is a method to measure contractility of isolated skeletal muscles. Parameters such as muscle force, muscle power, contractile kinetics, ...

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Developmental studies in the mouse are hampered  by the inaccessibility of the embryo during gestation. Thus, protocols to  isolate and culture individual ...

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Количественная Single микрососудов проницаемость в изолированном крови-перфузии Rat Подготовка легких

The isolated blood-perfused lung preparation is widely used to visualize and define signaling in single microvessels. By coupling this preparation with ...

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Сократимость Измерения на изолированных папиллярных мышц по расследованию сердечной инотропию у мышей

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