The use of a monoclonal antibody (HHF-35) against muscle-cell-specific actin has led to new insights into the early development and differentiation of the tunica media of blood vessels. The localization of this substance was studied by light and electron microscopy, in 22 rat embryos ranging between 10 and 15 days post coitum. Actin expression was already seen in the mesoderm around the dorsal aortae at 10 days post coitum, i.e. 1.5 day before circular mesenchymal condensations were detectable by light microscopy. These condensations are usually regarded as the first indication of arterial tunica media formation. The actin expression starts in the dorso-medial mesoderm surrounding the dorsal aortae at the level of the developing pharyngeal arch system, followed at 11.5 days by positive cells in the lateral mesoderm. In 12-day-old embryos most of the mesenchymal cells around the dorsal aortae contain actin, except for those aspects of the dorsal aortae which participate in outgrowing and connecting vessels to specific organs, such as the pharyngeal arch arteries, dorsal intersegmental arteries and aorto-pulmonary plexus, which themselves are still negative. At this stage the vitelline and umbilical arteries, which belong to the early developing ventral segmental arteries, are already surrounded by actin-containing mesenchymal cells. From 13 days onwards the tributary arteries, such as the subclavian and vertebral arteries, start to present the first differentiation of a tunica media in a proximo-distal development. In general the actin-negative areas are involved in vascular remodelling, implying the formation of new vessels, as as well as the disappearance of those previously developed.(ABSTRACT TRUNCATED AT 250 WORDS)