Name: assessing murine resistance artery function using pressure myography
Uploaded: 2013-06-07T12:00:00
Description: Watch this Scientific Journal Video about Assessing Murine Resistance Artery Function Using Pressure Myography at JoVE.com

Authors would like to acknowledge Drs. Paul Huang and Dmitriy Atochin for use of DMT pressure myograph and Drs. Binglan Yu and Chong Lei for providing mice fed with high fat diet or standard diet.
Sources of Funding
This work was supported by Scientist Development Grant 10SDG2610313 from the American Heart Association (to E. S. Buys), and an Eleanor and Miles Shore 50th Anniversary Fellowship program for Scholars in Medicine from Harvard Medical School (to E. S. Buys).

1. Preparation of Solution
<ol>
	<li>500X EDTA stock: weigh 500 mg EDTA-Na2&bull;2H2O and dissolve in 50 ml deionized water. Store at room temperature.</li>
	<li>KCl depolarizing solution.
	<ol>
		<li>Prepare K10X stock solution: 3.69 g NaCl, 18.64 g KCl, 0.36 g MgSO4 anhydrous, 0.41 g KH2PO4, 0.46 g CaCl2&bull;2H2O. Dissolve in 250 ml deionized water. Store at room temperature.</li>
		<li>For 100 ml final KCl depolarizing solution 1X: D...

<ol>
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Mice are the experimental model of choice for many investigators, in part because of the possibility to introduce genetic modifications, thereby generating mouse models for human pathophysiology. The vasoactive status of small resistance but not of larger conduit vessels largely defines the regulation of blood flow throughout the vascular system 11. The size of resistance arteries in small animals, such as mice, prevents the use of a wire myograph to study microvascular function. Pressure myographs not only ov...

Pressure myograph systems are used to measure the physiological function and properties of small arteries, veins and other vessels. An intact small segment of an artery or vein is mounted onto two small glass cannulas and pressurized to a suitable luminal pressure, allowing the vessel to maintain most of its in vivo characteristics (Figures 1 and 2). The near physiological condition in a pressure myograph reflects the in vivo behavior of the vascular bed, allowing the investigation of intrinsic properties (e.g. myogenic tone) of isolated vessels. Some of the advantages of pressure myography over wire myography, where muscle contraction is assessed by direct mechanical coupling to a force transducer 1, include (i) that micro resistance arteries, which define the overall resistance developed in vascular system, can be studied, whereas wire myograph is limited to larger conduit arteries, (ii) that the risk to damage the endothelium is reduced since no wires need to be passed through the vessel lumen, (iii) that the natural shape of the vessel is better maintained, and (iv) that vessel dimension can be studied at a wide range of pressures or shear stress 2.
Studying micro vessels can be more informative than studying larger conduit arteries to help understand the pathophysiology and molecular mechanisms that contribute to altered vascular tone in cardiovascular diseases such as hypertension. For example, impairment of endothelial function associated with feeding mice a high fat diet for 8 weeks could be demonstrated in 2nd-order mesenteric arteries 3 but not in aortic rings (Figure 3). Another advantage of pressure myography is that intrinsic myogenic constriction of the pressurized vessel is present, and that the role and function of the endothelium in this phenomenon can be studied. Here, we describe the use of a pressure myograph to study vascular reactivity of mouse mesenteric 3rd order resistance arteries in a setting of impaired nitric oxide (NO)-cGMP signaling.

<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

Pressure myograph systems are exquisitely useful in the functional assessment of small arteries, pressurized to a suitable transmural pressure. The near physiological condition achieved in pressure myography permits in-depth characterization of intrinsic responses to pharmacological and physiological stimuli, which can be extrapolated to the in vivo behavior of the vascular bed. Pressure myograph has several advantages over conventional wire myographs. For example, smaller resistance vessels can be studied at tightly controlled and physiologically relevant intraluminal pressures. Here, we study the ability of 3rd order mesenteric arteries (3-4 mm long), preconstricted with phenylephrine, to vaso-relax in response to acetylcholine. Mesenteric arteries are mounted on two cannulas connected to a pressurized and sealed system that is maintained at constant pressure of 60 mmHg. The lumen and outer diameter of the vessel are continuously recorded using a video camera, allowing real time quantification of the vasoconstriction and vasorelaxation in response to phenylephrine and acetylcholine, respectively.
	To demonstrate the applicability of pressure myography to study the etiology of cardiovascular disease, we assessed endothelium-dependent vascular function in a murine model of systemic hypertension. Mice deficient in the &alpha;1 subunit of soluble guanylate cyclase (sGC&alpha;1-/-) are hypertensive when on a 129S6 (S6) background (sGC&alpha;1-/-S6) but not when on a C57BL/6 (B6) background (sGC&alpha;1-/-B6). Using pressure myography, we demonstrate that sGC&alpha;1-deficiency results in impaired endothelium-dependent vasorelaxation. The vascular dysfunction is more pronounced in sGC&alpha;1-/-S6 than in sGC&alpha;1-/-B6 mice, likely contributing to the higher blood pressure in sGC&alpha;1-/-S6 than in sGC&alpha;1-/-B6 mice. 
	Pressure myography is a relatively simple, but sensitive and mechanistically useful technique that can be used to assess the effect of various stimuli on vascular contraction and relaxation, thereby augmenting our insight into the mechanisms underlying cardiovascular disease.

NO is centrally involved in maintenance of blood pressure homeostasis both in humans 6 and in animal models 7. The ability of NO to control vascular smooth muscle relaxation is mediated by soluble guanylate cyclase (sGC), a heme-containing heterodimeric enzyme that generates cGMP 8. Recently, a blood pressure-modifying genetic variant was identified in a locus that contains the sGC&alpha;1 and sGC&beta;1 genes, illustrating the relevance of sGC in regulating blood pr...

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

In pressure myography, an intact small segment of a vessel is mounted onto two small cannulas and pressurized to a suitable luminal pressure. Here, we describe the method to measure vasorelaxation response of the mouse 3rd order mesenteric arteries in c57 and sGC&alpha;1-/- mice using pressure myography.

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

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