Inhibitory mechanisms and binding site location for serotonin selective reuptake inhibitors on nicotinic acetylcholine receptors.

Functional and structural approaches were used to examine the inhibitory mechanisms and binding site location for fluoxetine and paroxetine, two serotonin selective reuptake inhibitors, on nicotinic acetylcholine receptors (AChRs) in different conformational states. The results establish that: (a) fluoxetine and paroxetine inhibit h alpha1beta1 gammadelta AChR-induced Ca(2+) influx with higher potencies than dizocilpine. The potency of fluoxetine is increased approximately 10-fold after longer pre-incubation periods, which is in agreement with the enhancement of [(3)H]cytisine binding to resting but activatable Torpedo AChRs elicited by these antidepressants, (b) fluoxetine and paroxetine inhibit the binding of the phencyclidine analog piperidyl-3,4-(3)H(N)]-(N-(1-(2 thienyl)cyclohexyl)-3,4-piperidine to the desensitized Torpedo AChR with higher affinities compared to the resting AChR, and (c) fluoxetine inhibits [(3)H]dizocilpine binding to the desensitized AChR, suggesting a mutually exclusive interaction.

Molecular mechanisms and binding site locations for noncompetitive antagonists of nicotinic acetylcholine receptors.

Nicotinic acetylcholine receptors are pentameric proteins that belong to the Cys-loop receptor superfamily. Their essential mechanism of functioning is to couple neurotransmitter binding, which occurs at the extracellular domain, to the opening of the membrane-spanning cation channel. The function of these receptors can be modulated by structurally different compounds called noncompetitive antagonists.

Molecular mechanisms and binding site location for the noncompetitive antagonist crystal violet on nicotinic acetylcholine receptors.

We investigated the molecular mechanisms and the binding site location for the fluorophor crystal violet (CrV), a noncompetitive antagonist of the nicotinic acetylcholine receptor (AChR). To this end, radiolabeled competition binding, fluorescence spectroscopy, Schild-type analysis, patch-clamp recordings, and molecular dynamics approaches were used. The results indicate that (i) CrV interacts with the desensitized Torpedo AChR with higher affinity than with the resting state at several temperatures (5-37 degrees C); (ii) CrV-induced inhibition of the phencyclidine (PCP) analogue [(3)H]thienylcyclohexylpiperidine binding to the desensitized or resting AChR is mediated by a steric mechanism; (iii) tetracaine inhibits CrV binding to the resting AChR, probably by a steric mechanism; (iv) barbiturates modulate CrV binding to the resting AChR by an allosteric mechanism; (v) CrV itself induces AChR desensitization; (vi) CrV decreases the peak of macroscopic currents by acting on the resting AChR but without affecting the desensitization rate from the open state; and (vii) two tertiary amino groups from CrV may bind to the alpha1-Glu(262) residues (located at position 20') in the resting state.

Anesthetics as chemical tools to study the structure and function of nicotinic acetylcholine receptors.

The nicotinic acetylcholine receptor (AChR) is the archetype of the Cys-loop ligand-gated ion channel receptor superfamily. Noncompetitive antagonists inhibit the AChR without interacting directly with agonist sites. Among noncompetitive antagonists, general and local anesthetics have been used for decades to study the structure and function of muscle- as well as neuronal-type AChRs.