Summary: Acetylcholine is an important regulator of local cerebral blood flow. There is, however, limited information available on the possible sites of action of this neurotransmitter on brain intraparenchymal microvessels. In this study, a com bination of molecular and functional approaches was used to identify which of the five muscarinic acetylcholine receptors (mAChR) are present in human brain microvessels and their intimately associated astroglial cells. Microvessel and capillary fractions isolate<;i from human cerebral cortex were found by reverse transcriptase-polymerase chain reaction to express m2, m3, and, occasionally, ml and m5 receptor subtypes. To local ize these receptors to a specific cellular compartment of the vessel wall, cultures of human brain microvascular endothelial and smooth muscle cells were used, together with cultured human brain astrocytes. Endothelial cells invariably expressed m2 and m5 receptors, and occasionally the ml receptor; smooth muscle cells exhibited messages for all except the m4 mAChR subtypes, whereas messages for all five muscarinic receptors Acetylcholine (ACh) is an important regulator of CBF in man and in many other species (for review see Sato and Sato, 1995;Dauphin and MacKenzie, 1995). Intra cortical and basalocortical cholinergic nerve fibers have been shown to intimately associate with microarterioles and capillaries in the rat (Vaucher and Hamel, 1995)