α‐Bungarotoxin Binds to Low‐Affinity Nicotine Binding Sites in Rat Brain

Abstract

Reported differences in the pharmacology and distribution of [3H]nicotine and [125I]alpha-bungarotoxin binding sites in mammalian brain suggest that these ligands label separate receptor sites. Affinity purification of an alpha-bungarotoxin binding protein from rat brain failed to copurify the high-affinity nicotine binding site, which remained in the nonbound soluble fraction after the affinity chromatography step. This confirms the independence of these putative receptor sites. Nevertheless, the binding of [125I]alpha-bungarotoxin to P2 membranes was inhibited by (-)-nicotine (Ki = 9 X 10(-6) M), and this sensitivity was preserved after affinity purification. It is proposed that alpha-bungarotoxin binds to a population of low-affinity nicotine binding sites. Comparison of the enantiomers of nicotine in competition studies at both radioligand binding sites revealed an 80-fold preference for the (-) form at the high-affinity [3H]nicotine binding site, whereas the site labelled by [125I]alpha-bungarotoxin displayed little stereoselectivity. In this respect, the brain alpha-bungarotoxin binding site resembles the nicotinic acetylcholine receptor from Torpedo electric organ.

Agonist pharmacology of the neuronal α7 nicotinic receptor expressed in Xenopus oocytes

“…In addition, (+)-anatoxin-a is shown to be the most potent agonist, as it is at other nAChR [17], while (+)-nicotine is two-fold weaker than the naturally occurring enantiomer. Such slight stereo~l~tivity in favour of (-)-nicotine is a property of aBgt-sensitive sites in Torpedo and brain [18], and is in marked contrast to aBgt-insensitive nAChR which show a loo-fold preference for (-)-nicotine [19]. Interestingly, DMPP was a relatively potent agonist at a7 nAChR but displayed only 20% of the efficacy of (-)-nicotine.…”

Comparison of the regional expression of nicotinic acetylcholine receptor ?7 mRNA and [125I]-?-bungarotoxin binding in human postmortem brain

“…Nucleic acid coding sequences are known for at least eight neuronal ␣ subtypes (␣2-␣9; Deneris et al, 1989;Boulter et al, 1990;Couturier et al, 1990;Fornasari et al, 1990;Schoepfer et al, 1990;Séguéla et al, 1993;Chini et al, 1994;Doucette-Stamm et al, 1994;Elgoyhen et al, 1994;Peng et al, 1994) and two ␤ subtypes (␤2 and ␤4; Deneris et al, 1988;Lindstrom, 1990, Tarroni et al, 1992;Monteggia et al, 1995). The nicotinic receptors are functionally subdivided into two principal classes based on their affinity for nicotine or ␣-bungarotoxin (␣BTX; Marks and Collins, 1982;Marks et al, 1986;Wonnacott, 1986;Amar et al, 1993). Receptors with a high affinity for nicotine are thought to be primarily composed of ␣2-␣6 and ␤2 and ␤4 subtypes Deneris et al, 1989;Wada et al, 1989;Happe et al, 1994), with the ␣4 and ␤2 receptor subtypes demonstrating the most widespread expression in mammalian brain Court and Clementi, 1995).…”

Visualization, characterization and modulation of calcium signaling during the development of slow muscle cells in intact zebrafish embryos

“…The tip of the tail was excised as described previously, after which embryos were treated with either 3 M -bungarotoxin-Alexa Fluor 488 (Molecular Probes) or 500 M nifedipine (Sigma-Aldrich Corp.) in 30% Danieau's solution at ~16.5 hpf. The former is an inhibitor of nicotinic acetylcholine receptors, nAChRs (Wonnacott, 1986;Pugh and Berg, 1994), while the latter is an organic blocker of the L-type voltage sensitive Ca 2+ channel, which targets the dihydropyridine binding site on the -subunit of the L-type Ca 2+ channel (Zamponi, 1997;Fischer and Schäfer, 2002). The Alexa Fluor 488 conjugate of -bungarotoxin was used so that the distribution of the toxin could be determined within the musculature of the embryonic trunk.…”