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Preparation of primary cultured mesenteric artery smooth muscle cells for fluorescent imaging and physiological studies

Nature Protocols volume 1, pages 2681–2687 (2006)Cite this article

Abstract

In this protocol, we describe a method for isolation and culture of smooth muscle cells derived from the adult rat (or mouse) superior mesenteric artery. Arterial myocytes are obtained by enzymatic dissociation and established in primary culture. The cultured cells retain expression of smooth muscle-specific α-actin and physiological responses to agonists. Cultured arterial myocytes (prepared from wild-type or transgenic animals) provide a useful model for studying the regulation of a wide range of vascular smooth muscle responses at the cellular and subcellular levels. Plasmids, RNA interference and antisense oligodeoxynucleotides can be readily introduced into the cells to alter protein expression. Fluorescent dyes can also be introduced to visualize a variety of activities, some of which may be specific to vascular smooth muscle cells. This protocol requires about 3 h on each of 2 consecutive days to complete.

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Figure 1: Cultured arterial myocytes retain Ca2+ responses to agonists.
Figure 2: Purity of rat ASMCs in primary culture.
Figure 3: High-magnification wide-field image of a small peripheral portion of intact (nonpermeabilized) arterial myocyte loaded with Furaptra (F346) at 36 °C (see ref. 15) (left panel).

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Acknowledgements

This work was supported by NHLBI grants HL-P01-078870 Project 1 (to M.P.B.) and Project 2 (to V.A.G.), and HL-R01-045215 (to M.P.B.).

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Authors and Affiliations

  1. Department of Physiology, University of Maryland School of Medicine, 685 West Baltimore St., HSF1, Room 571, Baltimore, 21201, Maryland, USA

    Vera A Golovina

  2. Department of Physiology, University of Maryland School of Medicine, 655 West Baltimore St., Room 5-007A, FCBRL, Baltimore, 21201, Maryland, USA

    Mordecai P Blaustein

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  1. Vera A Golovina
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Correspondence to Vera A Golovina.

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Golovina, V., Blaustein, M. Preparation of primary cultured mesenteric artery smooth muscle cells for fluorescent imaging and physiological studies. Nat Protoc 1, 2681–2687 (2006). https://doi.org/10.1038/nprot.2006.425

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