A subscription to JoVE is required to view this content. Sign in or start your free trial.
The present protocol describes the concepts and technical application of the tensometric myograph technique using a multi-chamber myograph system in the experimental ex vivo assessment of mouse aortic endothelial function.
Small volume chamber tensometric myography is a commonly used technique to evaluate the vascular contractility of small and large blood vessels in laboratory animals and small arteries isolated from human tissue. The technique allows researchers to maintain isolated blood vessels in a tightly controlled and standardized (near-physiological) setting, with the option of adjusting to various environmental factors, while challenging the isolated vessels with different pharmacological agents that can induce vasoconstriction or vasodilation. The myograph chamber also provides a platform to measure vascular reactivity in response to various hormones, inhibitors, and agonists that may impact the function of smooth muscle and endothelial layers separately or simultaneously. The blood vessel wall is a complex structure consisting of three different layers: the intima (endothelial layer), media (smooth muscle and elastin fibers), and adventitia (collagen and other connective tissue). To gain a clear understanding of the functional properties of each layer, it is critical to have access to an experimental platform and system that would allow for a combinational approach to study all three layers simultaneously. Such an approach demands access to a semi-physiological condition that would mimic the in vivo environment in an ex vivo setting. Small volume chamber tensometric myography has provided an ideal environment to evaluate the impact of environmental cues, experimental variables, or pharmacological agonists and antagonists on vascular properties. For many years, scientists have used the tensometric myograph technique to measure endothelial function and smooth muscle contractility in response to different agents. In this report, a small volume chamber tensometric myograph system is used to measure endothelial function in the isolated mouse aorta. This report focuses on how small volume chamber tensometric myography can be used to evaluate the functional integrity of the endothelium in small segments of a large artery such as the thoracic aorta.
For the last few decades, the small chamber myography system has been used to measure the reactivity of different layers of blood vessel walls in response to various pharmacological agents and neurotransmitters in an ex vivo, real-time setting. Vascular reactivity is a major component of a healthy functional blood vessel and is critical for the regulation of blood flow and perfusion in peripheral and cerebral vasculature1. Within the blood vessel wall, the interaction between endothelial and smooth muscle layers is a major determinant of vascular tone, which is also constantly impacted by structural changes in the connective tissue lay....
All surgical procedures and animal care were approved by the Institutional Animal Care and Use and Care Committee (IACUC) of Midwestern University (IACUC# AZ-3006, AZ-2936).
1. Buffer preparation
NOTE: Although the HEPES physiological salt solution (HEPES-PSS) buffer is stable at 4 °C for 7 days, it is recommended that all buffers are freshly made on the day of each experiment. All other reagents and agonists must be prepared freshly for each exp.......
The tensometric small chamber myography protocol explained here is the standard method for measuring vascular reactivity in small and large arteries and allows for simultaneous measurements of vascular reactivity in up to four blood vessel segments from the same experimental small laboratory animal. In this report, we specifically use the system to measure endothelial function in the isolated mouse aorta (Figure 1). In this protocol, isolated aortic segments are mounted onto a small organ ch.......
The field of vascular biology heavily relies on tools that help researchers to assess the functional and structural integrity of the blood vessel wall. It also demands special attention on the direct and indirect interactions between the three layers of blood vessels: the intima, media, and adventitia. Among those three layers, the intima is formed by a monolayer of endothelial cells and has a very important function in regulating vascular health and hemostasis.
It is well established that any.......
This work was supported by funding from the National Institutes of Health (R15HL145646) and Midwestern University College of Graduate Studies.
....Name | Company | Catalog Number | Comments |
Acetylcholine | SigmaAldrich | A6625-100G | |
CaCl2 | SigmaAldrich | C4901-1KG | |
Carbogen gas | Matheson | H103847 | |
Dissecting scissors | FST | 91460-11 | |
DMT 620 Multi chamber myograph system | DMT | DMT 620 | Multi chamber myograph system |
Dumont forceps | FST | 91150-20 | |
EDTA | SigmaAldrich | E5134-10G | |
Glucose | SigmaAldrich | G8270-1KG | |
HEPES | SigmaAldrich | H7006-1KG | |
KCl | SigmaAldrich | P9541-1KG | |
KH2PO4 | SigmaAldrich | P5655-1KG | |
LabChart | ADI instruments | Data acquisition software | |
Light source | Volpi | 14363 | |
L-Name | Fischer Scientific | 50-200-7725 | |
MgSO4 | SigmaAldrich | M2643-500G | |
Microscope | Leica | S6D | stereo zoom microscope |
NaCl | SigmaAldrich | S5886-5KG | |
NaHCO3 | SigmaAldrich | S5761-500G | |
Organ bath system | DMT | 720MO | |
Phenylephrine | SigmaAldrich | P6126-10G | |
Pump | Welch | 2546B-01 | |
Software | ADI instruments | LabChart 8.1.20 | |
Spring Scissors | FST | 15003-08 | |
Sylgard 184 Kit | Electron Microscopy Services | 24236-10 | silicone elastomer kit |
Tank Regulator | Fischer Scientific | 10575147 | |
Water bath system | Fischer Scientific | 15-462-10 |
This article has been published
Video Coming Soon
ABOUT JoVE
Copyright © 2024 MyJoVE Corporation. All rights reserved