MUSCARINIC RECEPTOR SUBTYPES IN FELINE TRACHEAL SUBMUCOSAL GLAND SECRETION

To determine what muscarinic receptor subtype regulates [Ca2+]i mediating airway submucosal gland secretion, we examined the effects of atropine (Atr), pirenzepine (PZ), 11{[2-(diethylamino)methyl-1-piperi-dinyl]acetyl}-5,11-dihydro-6H-pyrido (2,3-b)(1,4)-benzo-diazepin-6-one (AF-DX116) and 4-diphenylacetoxy-N-methyl-piperidine methiodide (4-DAMP) on methacholine (MCh)-evoked [Ca2+]i rise in acinar cells, and compared this with mucus glycoprotein (MGP) and electrolyte secretion evoked by MCh from submucosal glands isolated from feline trachea. [Ca2+]i was measured with the Ca2+-sensitive fluorescent dye, fura 2. We determined MGP secretion by measuring TCA-precipitable H-3-labeled glycoconjugates and electrolyte secretion by the change in the rate constant of Na-22-efflux from isolated glands. Half-maximal inhibitory concentrations (IC50) of PZ, AF-DX116, 4-DAMP, and Atr against MCh-evoked [Ca2+]i rise were 10(-7) M, 6 x 10(-6) M, 8 x 10(-9) M, and 6 x 10(-9) M, respectively. IC50 of PZ, AF-DX116, 4-DAMP, and Atr against MCh-evoked MGP secretion were 10(-6) M, 2 x 10(-5) M, 8 x 10(-9) M, and 6 x 10(-9) M, respectively. MCh (10(-5) M)-evoked Na-22 efflux was significantly inhibited by 10(-7) M 4-DAMP and 10(-7) M Atr (P < 0.01, each) but not by 10(-7) M PZ. Receptor binding assays with [H-3]quinuclidinyl benzilate showed that the K(i) values for PZ, AF-D x 116, 4-DAMP and Atr were 2.2 x 10(-8) M, 6.6 x 10(-7) M, 6.2 x 10(-10) M, and 2.9 x 10(-10) M, respectively. These findings suggest that cholinergic stimulation induces MGP and electrolyte secretions by the activation of muscarinic M3 receptors, involving [Ca2+]i as the second messenger in the secretory cells of airway submucosal glands.