THE REMOVAL OF ACETYLCHOLINE BY DIFFUSION AT NICOTINIC SYNAPSES IN THE RAT OTIC GANGLION

1. We have examined the clearance of synaptically released acetylcholi ne in the otic ganglion when acetylcholinesterase was blocked with ese rine. 2. Intracellular recordings were made from otic ganglion neurone s, in vitro. The decay of the excitatory postsynaptic potential (EPSP) , in response to stimulation of afferent fibres, was greatly prolonged in the presence of eserine. Low frequency (0.05-4 Hz) repetitive syna ptic stimulation led to a slow depolarization of the postsynaptic cell that persisted throughout the period of stimulation. This slow depola rization was blocked by the nicotinic antagonists mecamylamine (100 mu M) or (+) tubocurarine (100 mu M), but was unaffected by atropine (1 mu M), indicating that the response was due to the activation of nicot inic receptors. 3. Following 2 Hz synaptic stimulation (30 s), the rat e of rise of the slow depolarization had a time constant of 3.1 +/- 0. 4 s and a peak amplitude of 12 +/- 1 mV. Upon cessation of stimulation , the depolarization decayed to resting levels with a time constant of 18.3 +/- 1.5 s (n = 23). At increasing stimulation frequencies the ra te of rise of the depolarization increased. Lowering the probability o f release, by adding cadmium to the perfusing solution or by lowering extracellular calcium, slowed the rise time of the response. 4. Both t he onset and decay kinetics of the slow depolarization had a low tempe rature sensitivity indicating that they reflect diffusional processes. 5. Repetitive stimulation (2 Hz) of the afferent nerve supplying the ganglion, in the presence of eserine, also caused a slow depolarizatio n in cells in which we could not demonstrate a synaptic input. This in dicates that synaptically released acetylcholine can spill over onto n earby neurones. 6. We conclude that at parasympathetic synapses, under physiological conditions, transmitter action is terminated by the enz ymatic degradation of acetylcholine. When acetylcholinesterase is bloc ked, acetylcholine accumulates because its removal by diffusion is slo w.