Functional behavior of the beta-adrenergic receptor-adenylyl cyclase system in rabbit airway epithelium.

Abstract

Stimulation of adenylyl cyclase mediates the effects of beta-adrenergic agonists and prostaglandin E2 (PGE2) on tracheobronchial epithelial cell function by increasing intracellular cyclic adenosine monophosphate (cAMP). In turn, increases in cAMP affect airway function by modulating ciliary beating, chloride and water transport, mucus secretion, and release of bronchoactive substances. This study examined the function and regulation of the beta-adrenergic receptor-adenylyl cyclase system (beta AR-AC) in tracheal epithelial cells isolated from the rabbit, a frequently used animal model of airway reactivity, inflammation, and electrolyte transport. beta AR number, assessed by ligand binding using the non-subtype-specific beta-antagonist [125I]iodopindolol, averaged approximately 10,700 beta AR/cell (400 fmol/mg membrane protein). Greater than 85% of the receptors were of the beta 2 subtype as determined by competitive antagonist displacement of iodopindolol by selective beta 1- (betaxolol) and beta 2- (ICI 118,551) antagonists. cAMP synthesis was stimulated with isoproterenol, PGE2, and forskolin in a time- and concentration-dependent fashion. Preincubation of epithelial cells for 30 min with either isoproterenol (10 microM) or the peptide inflammatory mediator, bradykinin (100 microM), markedly depressed subsequent isoproterenol-stimulated cAMP synthesis. Isoproterenol-induced beta AR-AC desensitization appeared to be homologous since cAMP responses to PGE2 and forskolin, a direct activator of adenylyl cyclase, were not reduced. The effect of bradykinin on isoproterenol-stimulated cAMP response was mimicked by preincubation with either dioctanoyl glyceride or phorbol myristate acetate, activators of protein kinase C.(ABSTRACT TRUNCATED AT 250 WORDS)

Activation of endogenous deltaF508 cystic fibrosis transmembrane conductance regulator by phosphodiesterase inhibition.

“…PDE inhibitor-␤ agonist synergism and PDE inhibitor class specificity requirements for the activation of chloride transport in CF-T43 cells. 36 Cl efflux rate is plotted as a function of time, and the addition of stimulatory compounds is indicated by the solid bar. Error bars represent SEM.…”

“…In these experiments, chloride efflux appeared to be of larger magnitude when albuterol was used as an agonist, possibly indicating the involvement of ␤ 2 receptors in this process. Alternatively, this effect may be due to receptor desensitization known to occur with isoproterenol (36).…”

“…After the final efflux interval, cells were lysed with 1% Triton X-100 in HBR for 20 min at 37 Њ C. Scintillation fluid was added and vials were counted in a liquid scintillation counter (LS 5801; Beckman Instruments, Inc., Fullerton, CA). Efflux was calculated as the percentage of 36 Cl lost from the cells per 30-s interval from 36 Cl remaining in the cells and plotted as a function of time. Effects of drugs on efflux were calculated as the area under the curve corresponding to the presence of drugs and dividing by the average area for control.…”

Signature current of SO2-induced bronchitis in rabbit.

“…As an alternative explanation, the expression of I BE may have resulted from de novo generation of several kinds of membrane receptors including adenosine-and ␤-adrenergic receptors in bronchitis model epithelium. However, this possibility is remote because the normal rabbit epithelium isolated from trachea already possesses abundant ␤-adrenergic receptors and generates cAMP in response to ISO, epinephrine, or prostaglandin E 2 (20,21). Therefore, we assumed that I BE was evoked through a newly generated effector sensitive to a common cellular mechanism shared by ATP, ISO, and ADO.…”

“…On the other hand, this type of epithelium has been shown to have no Isc response to ␤ -agonists or agents that raise cellular cyclic adenosine 3 Ј ,5 Ј -monophosphate (cAMP) (13,14,19), although it possesses ␤ -adrenergic receptors as well as an ability to increase the cellular level of cAMP in the presence of epinephrine, isoproterenol, or forskolin (20,21). The electric refractoriness of the freshly isolated rabbit trachea to isoproterenol and other cAMP-mediated agents was also confirmed in our previous study, and extracellular ATP was found to elicit a transient increase in Isc that was carried by Cl Ϫ , probably via P 2u -receptor (22).…”