Regulation of force in skinned, single cells of ferret aortic smooth muscle

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Abstract
An isolation technique was developed for single cells from the ferret aorta, which resulted in the isolation of long (87±27 μm; x±SD, n=62), relaxed, pharmacologically active smooth muscle cells. These cells were attached to microtools, one of which was connected to a force transducer. Force in maximally phenylephrine-stimulated contractions of the intact cells averaged 2.3 ±1.4 μN (n=17). After cell skinning with saponin, the threshold for force development was 0.05 μM [Ca2+], and force reached a maximum of 4.4±1.6 μN (n = 36) at 0.5 μM [Ca2+]. Plots of relative steady-state force vs pCa (−log10[Ca2+]) were fit to the Hill equation, which yielded a pCa at half-maximal force of 6.87 ± 0.30 and a Hill coefficient of 2.3±1.4 (n = 29). When 2.5 μM calmodulin was added to the solutions, the calcium sensitivity of force was significantly increased (P0.05). In a solution of pCa 7, the skinned cells developed 2.5±0.5 μN (n = 5) of force when stimulated with a phorbol ester. The addition of a specific inhibitor (17 kDa) of protein kinase C to the calcium buffers depressed (P0.05). These data strongly suggest that in vascular smooth muscle, protein kinase C may be involved in a physiological, regulatory system for force.