BUMETANIDE-SENSITIVE NACL UPTAKE IN RABBIT TRACHEAL EPITHELIAL-CELLS IS STIMULATED BY NEUROHORMONES AND HYPERTONICITY

Loop diuretic-sensitive NaCl(K) cotransport plays a fundamental role in absorption and secretion of electrolytes in epithelial tissues. Cotransport activity was measured as uptake of Na-22, Cl-36, or Rb-86 at 27-degrees-C in isolated rabbit tracheal epithelial cells. Uptake of radiotracer was linear from 1 to 2 min after initiation of radiotracer transport. Bumetanide at 10-mu-M final concentration did not affect tracer uptake. The endogeneous catecholamine l-epinephrine and alpha(2)-adrenergic agent clonidine increased sodium and chloride uptake at least 5.5-fold. Bumetanide blocked sodium uptake by 85% and chloride uptake by 72%. Rb-86 uptake was not affected by l-epinephrine, clonidine, or bumetanide. The alpha(2)-adrenergic antagonist yohimbine blocked the effects of l-epinephrine and clonidine on Na-22 and Cl-36 uptake. In Ca2+-depleted transport medium, baseline levels of sodium and chloride uptake increased 3.8- and 2.4-fold, respectively, in a bumetanide-independent manner. Nevertheless, l-epinephrine and clonidine induced a net stimulation of sodium and chloride uptake similar to that found in Ca2+-replete medium. This response was reduced by bumetanide and yohimbine. The Ca2+-elevating agent ionomycin increased bumetanide-sensitive sodium and chloride uptake 7.2-and 6.2-fold, respectively. Replacement of chloride with gluconate or sodium with N-methyl-D-glucamine in the extracellular medium inhibited l-epinephrine and clonidine-stimulated bumetanide-sensitive sodium and chloride uptake, respectively. Osmotic shrinkage in hyperosmotic (500 mM NaCl with all other electrolytes at normal concentration) transport medium markedly increased bumetanide-inhibitable sodium and chloride uptake. Analysis of the bumetanide-sensitive component of stimulated transport demonstrated a Cl-to-Na uptake ratio that varied from 1.0 with hyperosmotic perturbation to 1.6 with hormonal stimulation. We conclude that loop diuretic-sensitive cotransport in rabbit tracheocytes is best modeled as K-independent NaCl cotransport. The cotransporter is inactive until stimulated by alpha(2)-adrenergic agents, increased intracellular Ca2+, or osmotic perturbation.