The Quest for the Inhibitory Neurotransmitter in Bovine Tracheal Smooth Muscle

The effect of inhibitory nerve stimulation on the mechanical, membrane potential and membrane conductance responses of isolated bovine tracheal smooth muscle has been studied. Membrane responses were measured in a sucrose-gap apparatus. In order to record inhibitory responses, it was necessary to increase tone in the preparation by applying a drug such as histamine. When tone was raised, repetitive field stimulation of intrinsic nerves caused depolarization and contraction, followed by relaxation and a suppression of histamine-induced slow waves. Hyperpolarization of the membrane was only seen following prolonged nerve stimulation, and there was no change in membrane conductance. The inhibitory effect of nerve stimulation was abolished by tetrodotoxin, but was not abolished by atropine, indomethacin, propranolol, naloxone or the purinergic blockers quinidine and theophylline. It was not satisfactorily mimicked by catecholamines, by gamma-amino-n-butyric acid (GABA) or by purines. Nerves with catecholamine fluorescence could not be found in the tracheal muscle layer. Neither adrenergic nor purinergic types of nerve terminal could be found in the tracheal muscle layer during ultrastructural examination of over one thousand nerve profiles. Vasoactive intestinal peptide (VIP) caused relaxation of the histamine-contracted tracheal muscle, suppressed the slow wave and caused slight hyperpolarization at higher concentrations, without affecting the membrane conductance. VIP was found in samples of tracheal muscle at a mean concentration of 1.95 ng/g. When the effluent solution flowing past isolated tracheal muscle strips was assayed for VIP, samples collected during inhibitory nerve stimulation had much higher concentrations of the peptide than samples collected before stimulation, after stimulation, or during stimulation in the presence of tetrodotoxin (10(-6) mol/l). The VIP content of the effluent during control periods was 73.8 pg/ml, and during stimulation was 167.5 pg/ml. It is suggested that VIP might be the non-adrenergic inhibitory neurotransmitter in bovine tracheal smooth muscle.