Myosin II regulates actin protrusive activity and is necessary to establish proper SC polarity. Subconfluent SC cultures stained with phalloidin and antibodies to N-cadherin, MLC, and vinculin. (A and B) In control cultures, SC display abundant cortical actin and stress fibers (dotted line). MLC staining is also found at stress fibers (insets 1 and 3) but not at focal adhesions (arrowheads, inset 3). (B) Two randomly polarized (front-to-back) SC showing concentration of N-cadherin staining in actin-rich protrusions at the leading edge (arrowheads) and also at the tip of the cell tail (asterisks, inset 2). Accumulation of MLC is observed at the retracting tail (inset 2), where it colocalizes with vinculin (inset 4). (C and D) SC treated with blebbistatin (25 μM for 60 min) display striking changes in the organization of the actin cytoskeleton, including loss of stress fibers and the formation of multiple actin spikes throughout the entire cell body, where N-cadherin is highly concentrated (arrowheads). Many long and very thin cell processes (asterisks) are also observed in treated cultures. (E–H) SC-DRG cocultures maintained for 3 d in myelin-promoting media with or without blebbistatin and stained for N-cadherin, neurofilament, and Hoechst nuclear staining. In control conditions (E and F), SCs that are starting to elongate (asterisks) show a “track-like” pattern of N-cadherin staining along the axons they contact (broken line). In blebbistatin-treated cultures (G and H), multipolar SCs (asterisks) extend many N-cadherin–positive processes toward various axons. Staining in these processes at the SC–axon interface (dotted line) does not show the track-like pattern found in controls. Arrowheads indicate processes with patchy N-cadherin staining that do not align with any axon in the field. Bars, 10 μm.