Modulation Of Respiratory Pattern By Acetylcholine: Effects Mediated By Muscarinic M3-Like Receptor And Their Ionic Mechanism In Prebötzinger Complex Inspiratory Neurons. X. M. Shao and J. L. Feldman. Department of Neurobiology, UCLA, Los Angeles, CA 90095-1763.
APStracts 6:0544N, 1999.
Perturbations of cholinergic neurotransmission in the brainstem affect respiratory motor pattern both in vivo and in vitro; the underlying cellular mechanisms are unclear. Using a medullary slice preparation from neonatal rat which spontaneously generates respiratory rhythm, we patch-clamped inspiratory neurons in the preBötzinger complex (preBötC), the hypothesized site for respiratory rhythm generation, and simultaneously recorded respiratory-related motor output from the hypoglossal nerve (XIIn). Most (88%) of the inspiratory neurons tested responded to local application of acetylcholine (ACh), carbachol (CCh) or bath application of muscarine. Bath application of 50 (M muscarine increased the frequency, amplitude and duration of XIIn inspiratory bursts. At the cellular level, muscarine induced a tonic inward current, increased the duration and decreased the amplitude of the phasic inspiratory inward currents in preBötC inspiratory neurons recorded under voltage-clamp at -60 mV. Muscarine also induced seizure-like activity evident during expiratory periods in XIIn activity; these effects were blocked by atropine. In the presence of tetrodotoxin (TTX), local ejection of 2 mM CCh or ACh onto preBötC inspiratory neurons induced an inward current along with an increase in membrane conductance under voltage-clamp and induced a depolarization under current-clamp. This response was blocked by atropine in a concentration-dependent manner. Bath application of 1 (M pirenzepine, 10 (M gallamine or 10 (M himbacine had little effect on the CCh-induced current, whereas 10 (M 4-Diphenylacetoxy-N-methylpiperidine methiodide (4-DAMP) blocked the current. The current-voltage (I-V) relationship of the CCh-induced response was linear in the range of -110 to -20 mV and reversed at -11.4 mV. Similar responses were found in both pacemaker and non-pacemaker inspiratory neurons. The response to CCh was unaffected when patch electrodes contained a high concentration of EGTA (11 mM) or BAPTA(10 mM). The response to CCh was greatly reduced by substitution of 128mM Tris-Cl for NaCl in the bath solution; the I-V curve shifted to the left and the reversal potential shifted to -47 mV. Lowering extracellular Cl- concentration from 140 to 70 mM had no effect on the reversal potential. These results suggest that in preBötC inspiratory neurons ACh acts on M3-like receptors on the postsynaptic neurons to open a channel permeable to Na+ and K+ that is not Ca2+-dependent. This inward cation current plays a major role in depolarizing preBötC inspiratory neurons, including pacemakers, that may account for the ACh-induced increase in the frequency of respiratory motor output observed at the systems/behavioral level.
Received 29 July 1999; accepted in final form 26 October 1999.
APS Manuscript Number J628-9.
Article publication pending Journal of Neurophysiology.
ISSN 1080-4757 Copyright 1999 The American Physiological Society.
Published in APStracts on 21 December 1999