Monolayer co-culture of rat heart cells and bovine adrenal chromaffin paraneurons |
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Authors: | J M Trifaró R Tang M L Novas |
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Institution: | (1) Secretory Process Research Program, Department of Pharmacology, Faculty of Medicine, University of Ottawa, 451 Smyth Rd., K1H 8M5 Ottawa, Ontario, Canada |
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Abstract: | Summary This paper describes a method for the preparation of co-cultures of rat heart cells and bovine adrenal chromaffin paraneurons.
The most suitable condition for heart cell isolation was when a combination of paraneurons. The most suitable condition for
heart cell isolation was when a combination of trypsin-DNAse I in Locke's solution was used for digestion. The best co-culture
conditions were obtained when 106 heart cells were plated on 7- to 8-d-old adrenal chromaffin paraneuron cultures containing 0.5×106 cells per 35-mm diameter culture dishes. Measurements of DNA (heart cells and chromaffin paraneurons), monitoring of beating
frequency (heart cells), and catecholamine (chromaffin paraneurons) levels and release indicated that both cell types remain
viable and functional, for several weeks. Heart cells started their characteristic contractile activity 24 h earlier when
plated either on viable or lysed chromaffin paraneurons, an effect apparently due to faster surface adhesion of heart cells.
The beating frequency of heart cells increased after treatment of co-cultures with either noradrenaline or nicotine, with
the latter agent acting indirectly through, the release of chromaffin paraneuron catecholamines. Propranolol produced a dose-related
inhibition of the responses to either noradrenaline or nicotine, thus suggesting that the increase in myocyte's beating activity
was mediated through β-receptors. Anti-myosin and anti-dopamine-β-hydroxylase immunostaining was used for cell type identification
and for the demonstration of body-to-body and process-to-process contacts between adrenal chromaffin paraneurons and heart
cells. This co-culture system will serve as a starting point of further studies directed to understand a) the influence of
a cell type on the development and on the phenotypic characteristics of a second cell type and b) the interaction of cells
derived from different organs and species.
This study was supported by grant PG-20 from Medical Research Council of Canada and a grant from the Ontario Heart and Stroke
Foundation. M. L. N. is a postdoctoral fellow of the MRC. |
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Keywords: | chromaffin paraneuron co-culture dopamine β -hydroxylase immunohistochemistry myocytes myosin |
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