Assessment of nicotinic acetylcholine receptor-mediated release of [ 3H]-norepinephrine from rat brain slices using a new 96-well format assay

• Published in: Neuropharmacology Vol. 39; no. 13; pp. 2663 - 2672
• Main Authors: Anderson, David J., Puttfarcken, Pamela S., Jacobs, Iris, Faltynek, Connie
• Format: Journal Article
• Language: English
• Published: Oxford Elsevier Ltd 01.10.2000; Elsevier
• Subjects: 1,1-Dimethyl-4-phenylpiperazine; Animals; Biological and medical sciences; Brain Chemistry - physiology; Central nervous system; Central neurotransmission. Neuromudulation. Pathways and receptors; Dimethylphenylpiperazinium Iodide - pharmacology; Fundamental and applied biological sciences. Psychology; Hippocampus - drug effects; Hippocampus - metabolism; In Vitro Techniques; Kinetics; Male; Neurotransmitter release; Nicotine; Nicotine - pharmacology; Nicotinic acetylcholine receptors; Nicotinic Agonists - pharmacology; Nicotinic Antagonists - pharmacology; Noradrenergic; Norepinephrine; Norepinephrine - metabolism; Prefrontal Cortex - drug effects; Prefrontal Cortex - metabolism; Rats; Rats, Sprague-Dawley; Receptors, Nicotinic - drug effects; Receptors, Nicotinic - metabolism; Spinal Cord - drug effects; Spinal Cord - metabolism; Thalamus - drug effects; Thalamus - metabolism; Vertebrates: nervous system and sense organs; Neurotransmitter release; Norepinephrine; 1,1-Dimethyl-4-phenylpiperazine; Noradrenergic; Nicotinic acetylcholine receptors; Nicotine; Agonist; Rat; Central nervous system; Molecular interaction; Frontal cortex; Well testing; Dose activity relation; Alkaloid; Cholinergic receptor; Investigation method; Thalamus; Antagonist; Nicotinic receptor; Dopamine; Rodentia; Catecholamine; In vitro; Vertebrata; Mammalia; Animal; Neurotransmitter; Hippocampus; Brain (vertebrata)
• ISSN: 0028-3908; 1873-7064
• DOI: 10.1016/S0028-3908(00)00143-X

The study of the modulatory effects of nicotinic acetylcholine receptor (nAChR) agonists on neurotransmitter release from tissue slices has been hampered by laborious and limiting superfusion techniques. A new methodology was developed utilizing 96-well filter plates. This new method produced comparable results to previously published data, yet expanded throughput to permit more complete pharmacological characterization. Rat brain slices, preloaded with [ 3H]-norepinephrine ([ 3H]-NE), were distributed onto 96-well filter plates. Following a 5 min preincubation, the slices were incubated for 5 min with nicotinic agonists or antagonists. (−)-Nicotine (NIC) and 1,1-dimethyl-4-phenylpiperazine (DMPP) evoked release of [ 3H]-NE from a number of brain regions and spinal cord, with the highest response seen in the hippocampus. Concentration–response curves revealed a rank order of potency of (±)-epibatidine>>anatoxin-a>A-85380>DMPP=NIC=(−)-cytisine in the hippocampus, thalamus, and frontal cortex. EC 50 values were approximately 0.005, 0.2, 1, 5, 5 and 5 μM, respectively. Concentration–inhibition curves of nicotine evoked [ 3H]-NE release from hippocampal and thalamic slices resulted in a rank order of potency of mecamylamine>hexamethonium>d-tubocurare>DHβE. Schild analysis revealed apparent noncompetitive antagonism of [ 3H]-NE release from hippocampus by mecamylamine, hexamethonium, and DHβE. In contrast, DHβE antagonism of [ 3H]-dopamine release from striatal slices using a similar methodology was competitive.