Regulation of force in skinned, single cells of ferret aortic smooth muscle.
An isolation technique was developed for single cells from the ferret aorta, which resulted in the isolation of long (87 +/- 27 microns; 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 microN (n = 17). After cell skinning with saponin, the threshold for force development was 0.05 microM [Ca2+], and force reached a maximum of 4.4 +/- 1.6 microN (n = 36) at 0.5 microM [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 microM calmodulin was added to the solutions, the calcium sensitivity of force was significantly increased (P less than 0.05) without changing the maximal force (P greater than 0.05). In a solution of pCa 7, the skinned cells developed 2.5 +/- 0.5 microN (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 (P less than 0.05) the maximally Ca2(+)-activated force without a change in the calcium sensitivity of force (P greater than 0.05). These data strongly suggest that in vascular smooth muscle, protein kinase C may be involved in a physiological, regulatory system for force.
Brozovich
F V
FV
Department of Medicine, Charles A. Dana Research Institute, Harvard-Thorndike Laboratory, Harvard Medical School, Beth Israel Hospital, Boston, MA 02215.
Walsh
M P
MP
Morgan
K G
KG
D000818:Animals; D001011:Aorta / Q000166:cytology* / Q000502:physiology / Q000503:physiopathology; D001696:Biomechanical Phenomena; D002118:Calcium / Q000494:pharmacology / Q000502:physiology; D002469:Cell Separation / Q000379:methods; D005289:Ferrets / Q000502:physiology*; D009119:Muscle Contraction / Q000187:drug effects; D009131:Muscle, Smooth, Vascular / Q000166:cytology* / Q000502:physiology / Q000503:physiopathology; D010656:Phenylephrine / Q000494:pharmacology; D010703:Phorbol Esters / Q000494:pharmacology; D011493:Protein Kinase C / Q000037:antagonists & inhibitors
D010703:Phorbol Esters; D010656:Phenylephrine; D011493:Protein Kinase C; D002118:Calcium