The three-dimensional detection of microvasculatory bed in the brain of white rat Rattus norvegicus by a Ca2+-ATPase method

A procedure is described which allows for the selective and non-injectional staining of the three-dimensional microvasculatory bed (MVB) in thick sections (60–140 μm) of formalin-fixed brain tissue of white rats Rattus norvegicus. This histochemical method detects ATPase activity and takes place between pH 10.5 and 11.2. Calcium ion is used to capture inorganic phosphate, calcium phosphate is converted to lead phosphate, and subsequently converted to black or dark brown lead sulfide. All vessels are revealed due to a precipitate on the endothelium and smooth muscle cells of arterioles. In some vessels, red blood cells also stain. The background is transparent with no staining of neurons, nerve fibers, glial cells, or nuclei. This allows for clear identification of arterioles, venules, and capillaries, which is difficult using other methods. New observations are described including the presence of webs connecting branching parts of arterioles and constrictions along vessels. This procedure should be useful in investigations of the MVB in rat brain.