The distributions of connexin 43 (Cx43) and connexin 40 (Cx40) in smooth muscle and endothelium of resistance vessels were examined using indirect immunofluorescence techniques coupled with confocal microscopy. Cx43 and Cx40 were found in smooth muscle and endothelium. Similar staining patterns were found in microvessel samples from brain and cremaster of the rat and from arterioles of the hamster cheek pouch. Double-labeling studies showed a high degree of colocalization of Cx40 with Cx43, suggesting the presence of multiple connexins within a single junctional plaque. Quantitative comparisons were made of the fluorescent patterns in the endothelium and smooth muscle of rat brain arterioles. Cx43 and Cx40 plaque diameters were 0.9 +/- 0.1 and 0.8 +/- 0.1 (SE) microns, respectively, in the endothelial layer and 0.5 +/- 0.1 and 0.5 +/- 0.1 microns, respectively, in the smooth muscle. There was no difference between mean plaque diameters of Cx43 and Cx40 in endothelium or smooth muscle. However, plaques were significantly larger in endothelium than in smooth muscle (P < 0.05). These findings demonstrate the potential for cell-cell communication in both cell types of the wall of arterioles from three different tissues. The data also suggest a greater level of coupling within the endothelium.