JCS02655 Supplementary Material

Files in this Data Supplement:

• Supplemental Figure 1 -

  Fig. S1. Effect of cGMP on motility of rat osteoclasts. (A) Cellular outlines of sequential images at 1 minute intervals of rat osteoclasts, with the initial position in red. A cluster of ten similar outlines near the initial position are outlines before addition of the agonist 8-Br-cGMP. (B) Change in the geometric cell center in sequential images of rat cells before (above red line) and after addition of 250 mM 8-Br-cGMP (below red line) showing the overall cell movement.

• Movie 1 -

  Movie 1. Effect of cGMP and NO on motility of human osteoclasts. Three sequences of 21 phase-contrast frames are shown. Each frame is separated by two minutes. Each frame is 290x225 mm. (1) Motion in osteoclasts over 40 minutes of cells with the cGMP blocking agent Rp-cGMPS, 50 mM, added 10 minutes prior to the first frame. Only minor motion is seen. There are small amounts of membrane ruffling and motion of nuclei and vacuoles in the cells. (2) In contrast, 2 hours after addition of the hydrolysis-resistant cGMP activator 8-pCPT-cGMP (100 mM), there is rapid motility. On average, the attached surface of the cells has decreased, and many cells appear rounded, although no floating cells are seen. The cells move in increments of ~10 mm every 5-10 minutes, with some movements back and forth and some circular. Net motion over 30 minutes (Fig. 1B of paper) or 1 hour is clear but typically less than total movement because the cells change direction aimlessly. (3) Motility after addition of the rapidly hydrolyzed NO donor sodium nitroprusside (100 mM). The agent is added after Frame 1. Note that the cell spread area declines over 5 minutes (three frames) with the cells then undergoing net, but random, motion similar to that after 8-pCPT-cGMP.