Abstract
The binding characteristics of [ 3H]cytisine, a putative CNS nicotinic receptor ligand, were examined in 4 regions of the human brain. [ 3H]Cytisine was found to bind non-cooperatively with high affinity to a single site in tissue homogenates and to exhibit low non-specific association. The binding characteristics of this ligand were evaluated in thalamus at 4°C and 24°C. The association constants were found to be 0.234 and 0.308 min −1nM −1, while the dissociation constants were 0.007 and 0.098 min −1, respectively. Saturation analysis of thalamus revealed the equilibrium K d to be 147 pM (4°C) and 245 pM (24°C), values in good agreement with those determined kinetically. The Hill coefficient varied slightly between brain regions; however, the mean values in all regions were examined were close to 1.0 at 0.95 ± 0.03 (4°C) and 0.91 ± 0.04 (24°C). [ 3H]Cytisine binding could be displaced using both nicotinic agonists and antagonists. Cytisine was the most potent displacer of [ 3H]cytisine binding with an K i of 250 pM. Nicotine and acetylcholine were also potent displacers with K i values of 1.8 and 8.1 nM, respectively. The nicotinic antagonists α-bungarotoxin and mecamylamine were ineffective competitors for the [ 3H]cytisine binding site while dihydro-β-erythroidine had an K i value of 109 nM. Thalamus showed the highest density of cytisine binding sites of all the regions examined (48 fmol/mg protein) while the hippocampus, cingulate gyrus and the cortex showed B max values of 18.9, 19.3 and 8.8 fmol/mg protein, respectively. The results of this characterization of [ 3H]cytisine binding are in excellent agreement with previously published reports on the nicotinic cholinergic receptor population in the human brain. We conclude that [ 3H]cytisine exhibits both high affinity for the receptor and a very low non-specific binding component which makes this a useful ligand for the quantitation of nicotinic receptors in tissues with low receptor density.
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