Receptor‐coupled shortening of α‐toxin‐permeabilized single smooth muscle cells from the guinea‐pig stomach

Abstract

1. Isolated single smooth muscle cells from the fundus of the guinea-pig stomach were permeabilized by use of Staphylococcus aureus alpha-toxin. Receptor-coupled shortening of individual cells was monitored under phase contrast microscopy. 2. Most of the isolated cells responded to 0.6 microM Ca2+, but not to 0.3 microM Ca2+, with a resulting maximal shortening to approximately 65% of the resting cell length. The contractile activity of these permeabilized cells lasted for several hours and repeated shortening was readily achieved after washing out. 3. Addition of acetylcholine (ACh) at a maximal concentration (10 microM) resulted in a marked decrease in the concentration of Ca2+ required to trigger a threshold response from 0.6 microM to 0.2 microM, and 1 mM guanosine 5'-diphosphate (GDP) blocked this decrease. Moreover, treatment with 100 microM guanosine 5'-triphosphate (GTP) mimicked the action of ACh. 4. Addition of 100 microM inositol 1,4,5-trisphosphate (InsP3) with 0.2 microM Ca2+ did not cause cell shortening, whereas 10 microM ACh with 0.2 microM Ca2+ did, suggesting that InsP3-induced Ca2+ release is not involved in ACh-operated cell shortening. 5. The present study demonstrates an alpha-toxin-permeabilized single smooth muscle cell preparation which retains its receptor function and also provides an insight into mechanisms leading to augmentation of Ca2+ sensitivity by stimulation of muscarinic receptors or GTP-binding proteins.