L-3,3',5-triiodothyronine and pregnenolone sulfate inhibit Torpedo nicotinic acetylcholine receptors.

Steven X Moffett,Grace Brannigan,Joseph V Martin,Eric A Klein,Tina Hinton

doi:10.1371/journal.pone.0223272

Steven X Moffett, Grace Brannigan + Show 3 more

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https://doi.org/10.1371/journal.pone.0223272

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Journal: PloS one	Publication Date: Oct 4, 2019
Citations: 4	License type: CC BY 4.0

Affiliation: Rutgers, The State University of New Jersey

Abstract

The nicotinic acetylcholine receptor (nAChR) is an excitatory pentameric ligand-gated ion channel (pLGIC), homologous to the inhibitory γ-aminobutyric acid (GABA) type A receptor targeted by pharmaceuticals and endogenous sedatives. Activation of the GABAA receptor by the neurosteroid allopregnanolone can be inhibited competitively by thyroid hormone (L-3,3’,5-triiodothyronine, or T3), but modulation of nAChR by T3 or neurosteroids has not been investigated. Here we show that allopregnanolone inhibits the nAChR from Torpedo californica at micromolar concentrations, as do T3 and the anionic neurosteroid pregnenolone sulfate (PS). We test for the role of protein and ligand charge in mediated receptor inhibition by varying pH in a narrow range around physiological pH. We find that both T3 and PS become less potent with increasing pH, with remarkably similar trends in IC50 when T3 is neutral at pH < 7.3. After deprotonation of T3 (but no additional deprotonation of PS) at pH 7.3, T3 loses potency more slowly with increasing pH than PS. We interpret this result as indicating the negative charge is not required for inhibition but does increase activity. Finally, we show that both T3 and PS affect nAChR channel desensitization, which may implicate a binding site homologous to one that was recently indicated for accelerated desensitization of the GABAA receptor by PS.

Highlights

The nicotinic acetylcholine receptor is an excitatory receptor protein localized in the central nervous system [1], the peripheral nervous system and neuromuscular junction
We tested the hypothesis that a population of anionic T3 transduces inhibition of nicotinic acetylcholine receptor (nAChR) at physiological pH. by extracting functional nAChRs from Torpedo californica and used twoelectrode voltage clamp (TEVC) to demonstrate functional effects of pregnanolone sulfate (PS) and T3 on nAChRs. We found that both PS and T3 inhibit nAChR activity due to acetylcholine, representing a case in which the same effect on channel function is demonstrated in both nAChRs and GABAA receptors
The apparent maximal effect of T3 reduced the nAChR control response by 80 ± 6%, with an IC50 of 5.4 ± 1 μM T3. This is very similar to the IC50 of T3 for the GABAA receptor (8 ± 2 μM) [29]

Summary

Introduction

The nicotinic acetylcholine receptor (nAChR) is an excitatory receptor protein localized in the central nervous system [1], the peripheral nervous system and neuromuscular junction (reviewed in [2]). Pathologies of the receptor, including epilepsy [3] and myasthenia gravis (in muscle-type nAChRs) [4, 5], demonstrate its crucial function in fast synaptic transmission. In muscle-type nAChRs, it is a cation-conducting member of the pentameric ligand-gated ion channel (pLGIC), or “Cys-loop” receptor superfamily [6,7,8,9,10]. Of the five homologous subunits that comprise the nAChR structure and central pore, two are identical (α, γ, α, β, δ). When acetylcholine molecules bind to the α-γ and α-δ subunit interfaces in the receptor’s extracellular.

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