Colocalization of ATP and nicotinic ACh receptors in the identified vagal preganglionic neurone of rat

1. Effects of exogenous adenosine 5'-triphosphate (ATP) and acetylcholine (ACh) were investigated on acutely dissociated preganglionic neurones in the dorsal motor nucleus of vagus (DMV) of rate using whole-cell patch clamp recording methods. 2. The DMV neurones identified by retrograde transport of 1,1'-dioctadecyl-3,3,3'3'-tetra-methylindocarbocyanine perchlorate (DiI) fixed onto the cervical vagal nerve bundle were large in size (25-35 mu m diameter) and bipolar or tripolar in shape. 3. About 90% of DiI labelled DMV neurones responded to both ATP (10(-4) M) and ACh (10(-4) M) with inward currents at a holding potential (V-h) of -40 mV. 4. The ATP-induced current (I-ATP) and the ACh-induced current (I-ACh) reversed their polarities at membrane potentials between +5 and +15 mV, indicating that ATP and ACh increase the membrane permeability to cations. 5. The inhibitory potency of Reactive Blue on 5 x 10(-4) M I-ATP for half-inhibition (IC50), 4.4 x 10(-7) M) than suramin (IC50, 6.0 x 10(-6) M). In addition, alpha,beta-methylene ATP up to 10(-4) M could not induce any current. As intracellular application of guanosine 5'-O-(2-thiodiphosphate) (GDP beta S) did not block the I-ATP, the I-ATP was mediated not by guanosine triphosphate (GTP) binding protein, but rather by ligand-gated ionic channels, presumably via P-2X receptors. 6. Currents produced by ACh were due to activation of nicotinic receptors because they were mimicked by nicotine and carbachol, and blocked by hexamethonium. In addition, muscarine evoked no response. 7. Only 25% of nucleus tractus solitarii (NTS) neurones and no hypoglossal neurones responded to the exogenous application of ATP. 8. These results suggest that vagal preganglionic neurones colocalize functionally nicotinic and P-2X purinergic receptors.