The role of interstitial cells of Cajal (ICC) is difficult to determine because these cells are not easily identified by light microscopy, and there are no compounds available to specifically lesion ICC. Ultrastructural studies have shown an abundance of mitochondria in ICC. Therefore, we have used rhodamine 123, a fluorescent dye that is specifically accumulated by mitochondria, to identify ICC in canine proximal colon. This technique provided good discrimination between ICC and smooth muscle cells, but enteric neurons were labeled with rhodamine 123. This compound has cytotoxic properties in some cells. Therefore, we treated intact muscle strips with rhodamine 123 while recording intracellular electrical activity from circular muscle cells. Uptake of rhodamine 123 by ICC was associated with an alteration in electrical rhythmicity. These data suggest that rhodamine 123 may be a useful tool for visualizing and perhaps chemically lesioning ICC.