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Single-cell contraction assay for human ciliary muscle cells. Effect of carbachol
Purpose:
The authors developed an assay to observe the contraction of a single human ciliary muscle cell.
Methods:
Cultured human ciliary muscle cells were partially detached from the culture dish by incubation with a nonenzymatic dissociation buffer and treated with carbachol or pilocarpine. Contraction was quantified by measuring the cross-sectional surface areas of the cells.
Results:
Carbachol decreased the cell surface area in a time-dependent manner. Contraction was observed within 1 min after the addition of carbachol and completed in less than 15 min. The effect of carbachol was dose dependent. For example, at 10 min after treatment with 10 mumol/l carbachol, the relative surface areas of cells decreased to 47% +/- 4% (mean +/- standard error of the mean, n = 7, with surface area at 0 min defined as 100%). The relative surface areas were 74% +/- 4% (n = 7) after 1 mumol/l and 100% +/- 9% (n = 7) after 0.1 mumol/l carbachol treatment. This contractile effect was antagonized by pretreatment with atropine, a specific muscarinic antagonist.
Conclusions:
A simple method was established to study the functional changes of human ciliary muscle cells.