Nicotinic receptor binding sites in rat primary neuronal cells in culture: characterization and their regulation by chronic nicotine.
We have characterized high affinity neuronal nicotinic acetylcholine receptors labeled by [3H]cytisine in primary neuronal cell cultures from fetal rat brains. After 15 days in culture, the highest density of [3H]cytisine binding sites (Bmax approximately 57 fmol/mg protein) was found in cells from the brainstem, which includes the following subcortical brain areas: the septum, thalamus, hypothalamus, midbrain, pons and medulla. A lower density of sites was found in cells from the cerebral cortex, hippocampus, and caudate nucleus. [3H]Cytisine binds to receptors in primary cells from the brainstem and cerebral cortex with a Kd of approximately 0. 5 nM, and the binding is inhibited by the agonists nicotine, acetylcholine, and epibatidine with IC50 values of 1 to 20 nM, and by carbachol and the antagonist dihydro-beta-erythroidine with IC50 values of 0.5 to 1.5 microM. Chronic treatment of neuronal cultures with nicotine for 7 days differentially affected the number of nicotinic receptors in cells from different brain areas; it significantly increased the number of nicotinic binding sites in cells from the cerebral cortex, hippocampus, and caudate, but not in cells from the brainstem. The nicotine-induced increase of receptors in cerebral cortical cultures was not blocked by either mecamylamine or dihydro-beta-erythroidine. These results indicate that primary cultures of rat neuronal cells provide a good model system in which to study and compare the properties and regulation of native neuronal nicotinic acetylcholine receptors.
Dávila-García
M I
MI
Department of Pharmacology, Georgetown University School of Medicine, 3900 Reservoir Road, Washington, DC 20007, USA.
Houghtling
R A
RA
Qasba
S S
SS
Kellar
K J
KJ
D000109:Acetylcholine / Q000502:physiology; D000470:Alkaloids / Q000494:pharmacology; D000818:Animals; D001392:Azocines; D001667:Binding, Competitive / Q000187:drug effects / Q000502:physiology; D001923:Brain Chemistry; D001933:Brain Stem / Q000737:chemistry / Q000166:cytology; D002478:Cells, Cultured; D002540:Cerebral Cortex / Q000737:chemistry / Q000166:cytology; D004079:Dihydro-beta-Erythroidine / Q000494:pharmacology; D005260:Female; D008464:Mecamylamine / Q000494:pharmacology; D009474:Neurons / Q000737:chemistry* / Q000166:cytology / Q000502:physiology; D009538:Nicotine / Q000494:pharmacology*; D018722:Nicotinic Agonists / Q000494:pharmacology*; D018733:Nicotinic Antagonists / Q000494:pharmacology; D011247:Pregnancy; D011807:Quinolizines; D011869:Radioligand Assay; D051381:Rats; D017207:Rats, Sprague-Dawley; D011978:Receptors, Nicotinic / Q000737:chemistry / Q000378:metabolism*; D014316:Tritium; D000093843:Quinolizidine Alkaloids
D000109:Acetylcholine; D000470:Alkaloids; D001392:Azocines; D004079:Dihydro-beta-Erythroidine; D008464:Mecamylamine; D009538:Nicotine; D018722:Nicotinic Agonists; D018733:Nicotinic Antagonists; D011807:Quinolizines; D011978:Receptors, Nicotinic; D014316:Tritium; C004712:cytisine; D000093843:Quinolizidine Alkaloids