Effect of cholinergic stimulation on the release of macromolecules by canine trachea in vitro.

Abstract

SUMMARy _ The incorporation of D-[6-3H] glucosamine into the macromolecules synthesized by canine tracheo­ bronchial tissue provided a method for evaluating the influence of various physiologic and pharma­ cologic parameters on respiratory glycoprotein secretion. The cervical portion of canine trachea and the bifurcation were incubated in vitro for 18 hours with tissue culture medium 199 containing [3H] glucosamine. The tissues were rinsed and allowed to incubate further for 2-hour periods in unlabeled tissue culture medium. To study the effects of various agents on the release of total 3H-labeled macromolecules, compounds were added during the second and/or third bath periods after an initial basal control period. Addition of acetylcholine chloride to the bath fluid significantly increased the release of total [3H] macromolecules, presumably by stimulation of cholinergically innervated submucosal mucous glands, an effect that could be pharmacologically antagonized by atropine sulfate. Histamine, norepinephrine, and bromhexine were without any significant effect on the release of [3H] macro­ molecules at the concentrations studied. Concentrated aliquots of the dialyzed bath solutions were chromatographed on columns of I per cent agarose by continuous elution with Tris-phosphate buffer in 6 M urea. The increased release of [3H] macromolecules induced by cholinergic stimulation appeared to be accounted for by increases in the void volume peak of tritium eluted from agarose columns at a mobility identi­ cal with large molecular weight mucin type glycoproteins of canine tracheal pouch secretions collected in vivo.