Cholinergic Modulation of Dural Mast Cells

To study the action of cholinergic drugs on degranulation of dural mast cells, hemiskulls of Wistar rats of both sexes were used. One control hemiskull was filled for 20 min with the basic saline solution (BSS), containing (in millimolars) 152 NaCl, 5 KCl, 10 HEPES, 10 glucose, 2.6 CaCl₂, 2.1 MgCl₂ (pH adjusted to 7.4 with NaOH), whereas others were filled with BSS containing 50 µM carbachol or 100 µM nicotine. Then, the hemiskulls were left in 4% PFA solution for at least 4 h. After that meninges were dissected carefully and placed on a microscope slide, where they were stained with 0.1% toluidine blue (26 (link)). Images of meninges were acquired with Olympus AX70 microscope (20× objective). Homogeneously stained and well-shaped mast cells were classified as non-degranulated, whereas pale poorly stained mast cells as well as mast cells with distorted borders were classified as degranulated (22 (link), 26 (link)). To study an effect of carbachol and nicotine on mast cells degranulation, 10 randomly chosen, same in different animals, perivascular areas of meninges enriched by mast cells were analyzed (27 (link)). Then, total number and number of degranulated mast cells were counted in a blind manner.