No abstract

With the recent availability of ( -)-[3H]nicotine of high specific activity, binding studies were performed on rat brain membranes in the presence of a variety of nicotine analogues and cholinergic drugs. Both a higher affinity (& = 2 x lo-" M) and a lower affinity (2 x 10e9 M) site were observed; the stereoselectivity of both sites being similar. A good correlation was observed between Z&-values and psychotropic potency of a series of N'alkyl substituted nicotine analogues. Nicotine receptor Rat brain Nicotine analogue Cholinergic drugNicotine psychopharmacology

A comparison of the binding of (-)- and (+)-[3H]nicotine to rat brain membranes revealed that only the (-)-enantiomer showed high affinity binding; while the (+)-enantiomer was at least 1/10 as effective as the (-)-enantiomer when in competition with (-)-[3H]nicotine as the ligand. Positive cooperativity, which is observed with (-)-[3H]nicotine in the presence of low concentrations of (+)-nicotine, may account for the seeming paradox.

No abstract

With the use of affinity chromatography, a [3H]-nicotine binding site was purified almost 1,000-fold from a Triton X-100-solubilized extract of rat brain neural membranes. The affinity column was prepared by conjugation of (R,S)-6-(2-hydroxyethyl)nicotine to epoxy-activated Sepharose. Further purification of the material from the affinity column was resolved by using another column of the same affinity gel, resulting in the isolation of a major protein (about 95% purity) that had a Mr of 56,000, as determined by NaDodSO4/polyacrylamide gel electrophoresis, with very minor components ranging in Mr from 47,000 to 83,000. With the use of various nicotine analogues, it was shown that the purified material exhibited nearly identical binding characteristics to rat brain membrane preparations, including stereoselectivity for the nicotine enantiomers. The Kd of the purified site, 3.5 X 10-9 M, was similar to that observed with membrane and Triton X-100-soluble preparations, whereas the binding capacity was >25 pmol/mg of protein, as compared to 0.07 pmol/mg of protein in the starting material. The results are discussed in relation to the purified nicotinic cholinergic receptor from electroplax. It was concluded that the nicotine site in rat brain was different from the cholinergic receptor of electroplax or calf skeletal muscle.A significant achievement in modern pharmacology and neurochemistry has been the isolation and characterization of the peripheral nicotinic cholinergic receptor (AcChoR) from the electric organ of Torpedo (fish) (1-3), electric eel (4, 5), and housefly brain (6). Various types of nicotinic cholinergic ligands, including a-bungarotoxin (BuTX) and bisquaternary amines, have been successfully employed in such studies, utilizing detergents such as Triton X-100 and desoxycholate to solubilize the receptor. The availability of the purified receptor has proven crucial to the understanding of receptor function and in characterizing the molecular nature of the reactive sites (7)(8)(9)(10)(11)(12).On the other hand, progress has been considerably slower in regard to the isolation and characterization of brain nicotinic receptors. In the past few years, numerous studies have focused on nicotine's neuropharmacologic and behavioral properties (13)(14)(15)(16)(17)(18)(19)(20)(21)(22)(23). Recently, there have been demonstrations of high-affinity nicotine binding sites in mammalian brain that exhibit a high degree of specificity for chemical structures related to the nicotine molecule and considerably lower affinity for nicotinic cholinergic antagonists and agonists (16,(24)(25)(26)(27)(28). Evidence has also been presented that the brain site's receptors may be noncholinergic (16,25,29).Considerable difficulties are encountered in the isolation of brain nicotinic receptors. Although they appear to be widely distributed throughout the brain (24), the nicotinic receptors are located on neuronal membranes rather than on glands or effector organs as in the peripheral nervous system (10). Although BuTX has...