Bioassay development: The implications of cardiac myocyte motility in vitro

Abstract

Cardiac myocytes cultured over microfabricated extracellular recording devices can be used to assay bioactive compounds. However, electrophysiological signals recorded from these devices vary in amplitude with time. Theoretically, changes in signal amplitude arise from myocytes being moved over recording sites by cocultured fibroblasts. To test this, neonatal rat cardiac myocytes were cultured at high densities and low densities on fibronectin-coated glass. After 36.5 h, myocytes were identified by their rhythmic contractions and then time-lapse-recorded for 3.5 h. Length, width, and angle of orientation was then determined every 30 min for five cells in low density and five cells in high-density culture. Low-density cells had mean lengths of 65.3 microm and widths of 35.1 microm, whereas cells in high-density culture had greater mean lengths of 74.2 microm and lower mean widths of 24.3 microm. Length, width, and angle of orientation of cells in low- and high-density culture changed by 4.1%, 11.8%, and 2.7 degrees, and 6.4%, 10%, and 4.6 degrees, respectively, every half hour. We found no evidence of myocyte-fibroblast interactions influencing cell position or shape in low density, but in high density, we found evidence that fibroblast-myocyte interactions could transiently influence cell shape. We conclude that fibroblast-independent changes in cell shape are largely responsible for the changes in signal amplitude recorded from cardiac myocytes cultured on microfabricated extracellular recording devices. However, there is some evidence that myocyte-fibroblast interactions may augment this process in high-density culture. The implications of these findings for bioassay development are discussed.

Recording of cell action potentials with AlGaN∕GaN field-effect transistors

“…The preparation for the cardiac myocyte cells of embryonic Wistar rats ͑Charles River GmbH, Sulzbach, Germany, E 19͒ was adapted from previously published protocols. 22 The AlGaN / GaN chips were cleaned with 70% ͑v/v͒ ethanol ͑p.a.͒ and coated for 30 min with 20 l of 12.5 g/ml fibronectin ͑Sigma-Aldrich͒ in Hanks' balanced salt solution ͑HBSS͒ at 37°C. The surfaces were rinsed with Phosphatebuffered saline ͑PBS͒ buffer solution ͑137 mM NaCl, 2.7 mM KCl, 4.3 mM Na 2 HPO 4 , 1.4 mM NaH 2 PO 4 , pH 7.3͒.…”

Relationship between Adsorbed Fibronectin and Cell Adhesion on a Honeycomb-patterned Film

“…CMYs were isolated by enzyme treatment of minced heart tissues of 19-day rat embryos (Sprague Dawley rats, Japan SLC, Inc) [38]. CMYs were seeded onto the pre-fabricated cell-culture substrates (i.e., a flat film and a self-supported honeycomb film) at a density of 1.0×10 5 Results and Discussion Figure 1 shows SEM images of the structure of the PCL honeycomb film.…”

Honeycomb Films of Biodegradable Polymers for Tissue Engineering

“…HEPs were cultured with a Williams'E medium containing dexamethasone (I jiM, Sigma), ascorbic acid (0.28 mM, Sigma), insulin (0.57 mg/L, Sigma), epidermal growth factor (0.02 mg/L, Sigma), gentamicin (48 mg/L, Schellingplau), and aprotinin (5000 KIU/L, Sigma). Cardiac myocytes (CMYs) were isolated by enzyme treatment of minced, heart tissues of 19-day rat embryos (Sprague Dawley rats; Japan SLC, lnc) [8]. CMYs were cultured with a Hepes-buffered Hams FI0 medium containing 0.5 % insulin-transferrin-selenium-X (Gibco) and 3 % fetal calf serum (Gibco).…”