scholarly journals Novel structural determinants of single channel conductance and ion selectivity in 5-hydroxytryptamine type 3 and nicotinic acetylcholine receptors

2010 ◽  
Vol 588 (4) ◽  
pp. 587-596 ◽  
Author(s):  
John A. Peters ◽  
Michelle A. Cooper ◽  
Jane E. Carland ◽  
Matthew R. Livesey ◽  
Tim G. Hales ◽  
...  
Keyword(s):  
Nicotinic Acetylcholine Receptors ◽  
Acetylcholine Receptors ◽  
Single Channel ◽  
Ion Selectivity ◽  
Channel Conductance ◽  
Single Channel Conductance ◽  
Structural Determinants ◽  
Nicotinic Acetylcholine ◽  
Type 3

Novel structural determinants of single-channel conductance in nicotinic acetylcholine and 5-hydroxytryptamine type-3 receptors

2006 ◽  
Vol 34 (5) ◽  
pp. 882-886 ◽  
Author(s):  
J.A. Peters ◽  
J.E. Carland ◽  
M.A. Cooper ◽  
M.R. Livesey ◽  
T.Z. Deeb ◽  
...  
Keyword(s):  
Transmembrane Domain ◽  
Single Channel ◽  
Ion Selectivity ◽  
Atomic Scale ◽  
Scale Model ◽  
Amino Acid Residues ◽  
Channel Conductance ◽  
Single Channel Conductance ◽  
Structural Determinants ◽  
Type 3

Nicotinic ACh (acetylcholine) and 5-HT3 (5-hydroxytryptamine type-3) receptors are cation-selective ion channels of the Cys-loop transmitter-gated ion channel superfamily. Numerous lines of evidence indicate that the channel lining domain of such receptors is formed by the α-helical M2 domain (second transmembrane domain) contributed by each of five subunits present within the receptor complex. Specific amino acid residues within the M2 domain have accordingly been demonstrated to influence both single-channel conductance (γ) and ion selectivity. However, it is now clear from work performed on the homomeric 5-HT3A receptor, heteromeric 5-HT3A/5-HT3B receptor and 5-HT3A/5-HT3B receptor subunit chimaeric constructs that an additional major determinant of γ resides within a cytoplasmic domain of the receptor termed the MA-stretch (membrane-associated stretch). The MA-stretch, within the M3–M4 loop, is not traditionally thought to be implicated in ion permeation and selection. Here, we describe how such observations extend to a representative neuronal nicotinic ACh receptor composed of α4 and β2 subunits and, by inference, probably other members of the Cys-loop family. In addition, we will attempt to interpret our results within the context of a recently developed atomic scale model of the nicotinic ACh receptor of Torpedo marmorata (marbled electric ray).

The 5-hydroxytryptamine type 3 (5-HT3) receptor reveals a novel determinant of single-channel conductance

2004 ◽  
Vol 32 (3) ◽  
pp. 547-552 ◽  
Author(s):  
J.A. Peters ◽  
S.P. Kelley ◽  
J.I. Dunlop ◽  
E.F. Kirkness ◽  
T.G. Hales ◽  
...  
Keyword(s):  
Ion Channels ◽  
Acetylcholine Receptor ◽  
Nicotinic Acetylcholine Receptor ◽  
Single Channel ◽  
Channel Conductance ◽  
Single Channel Conductance ◽  
Nicotinic Acetylcholine ◽  
Ultrastructural Studies ◽  
Arginine Residues ◽  
Type 3

5-HT3 (5-hydroxytryptamine type 3) receptors are cation-selective ion channels of the Cys-loop transmitter-gated ion channel superfamily. Two 5-HT3 receptor subunits, 5-HT3A and 5-HT3B, have been characterized in detail, although additional putative 5-HT3 subunit genes (HTR3C, HTR3D and HTR3E) have recently been reported. 5-HT3 receptors function as homopentameric assemblies of the 5-HT3 subunit, or heteropentamers of 5-HT3A and 5-HT3B subunits of unknown stoichiometry. The single-channel conductances of human recombinant homomeric and heteromeric 5-HT3 receptors are markedly different, being <1 and approx. 16 pS respectively. Paradoxically, from the results of studies performed on the closely related nicotinic acetylcholine receptor, the channel-lining M2 domain of the 5-HT3A subunit is predicted to enhance cation conduction, whereas that of the 5-HT3B subunit would not. The present study describes a novel determinant of single-channel conductance, outwith the M2 domain, which accounts for this anomaly. Utilizing a panel of chimaeric 5-HT3A and 5-HT3B subunits, a profound determinant of single-channel conductance was traced to a putative amphipathic helix (the ‘HA stretch’) within the large cytoplasmic loop of the receptor. Replacement of three arginine residues (R432, R436 and R440) unique to the HA stretch of the 5-HT3A subunit with the aligned residues (Q395, D399 and A403) of the 5-HT3B subunit increased the single-channel conductance 28-fold. Significantly, from ultrastructural studies of the Torpedo nicotinic acetylcholine receptor, the key residues may be components of narrow openings within the inner vestibule of the channel, located in the cytoplasm, which contribute to the permeation pathway. Our findings indicate an important and hitherto unappreciated function for the HA stretch in the Cys-loop family of transmitter-gated ion channels.

scholarly journals Common Determinants of Single Channel Conductance within the Large Cytoplasmic Loop of 5-Hydroxytryptamine Type 3 and α4β2Nicotinic Acetylcholine Receptors

2006 ◽  
Vol 281 (12) ◽  
pp. 8062-8071 ◽  
Author(s):  
Tim G. Hales ◽  
James I. Dunlop ◽  
Tarek Z. Deeb ◽  
Jane E. Carland ◽  
Stephen P. Kelley ◽  
...  
Keyword(s):  
Acetylcholine Receptors ◽  
Single Channel ◽  
Channel Conductance ◽  
Single Channel Conductance ◽  
Cytoplasmic Loop ◽  
Type 3

Differential Sensitivities of Mammalian Neuronal and Muscle Nicotinic Acetylcholine Receptors to General Anesthetics 

1997 ◽  
Vol 86 (4) ◽  
pp. 866-874 ◽  
Author(s):  
J. M. Violet ◽  
D. L. Downie ◽  
R. C. Nakisa ◽  
W. R. Lieb ◽  
N. P. Franks
Keyword(s):  
Nicotinic Acetylcholine Receptors ◽  
Acetylcholine Receptors ◽  
Xenopus Oocytes ◽  
General Anesthetics ◽  
Response Curves ◽  
Nicotinic Acetylcholine ◽  
Considerable Uncertainty ◽  
Messenger Ribonucleic Acid ◽  
The Central Nervous System ◽  
Type 3

Background Nicotinic acetylcholine receptors (nAChRs) are members of a superfamily of fast neurotransmitter-gated receptor channels that includes the gamma-aminobutyric acidA (GABAA), glycine and serotonin type 3 (5-HT3) receptors. Most previous work on the interactions of general anesthetics with nAChRs has involved the muscle-type receptor. The authors investigate the effects of general anesthetics on defined mammalian neuronal and muscle nAChRs expressed in Xenopus oocytes. Methods Complementary deoxyribonucleic acid (cDNA) or messenger ribonucleic acid (mRNA) encoding for various neuronal or muscle nAChR subunits was injected into Xenopus oocytes, and the resulting ACh-activated currents were studied using the two-electrode voltage-clamp technique. The effects of halothane, isoflurane, sevoflurane, and propofol on the peak acetylcholine-induced currents were investigated, and concentration-response curves were constructed. Results The neuronal nAChRs were found to be much more sensitive to general anesthetics than were the muscle nAChRs, with IC50 concentrations being 10- to 35-fold less for the neuronal receptors. For the inhalational general anesthetics, the IC50 concentrations were considerably less than the free aqueous concentrations that cause general anesthesia in mammals. In addition, qualitative (dependence on acetylcholine concentration) and quantitative (steepness of concentration-response curves) differences in the anesthetic interactions between the neuronal and muscle nAChRs suggest that different mechanisms of inhibition may be involved. Conclusions Although there is considerable uncertainty about the physiologic roles that neuronal nAChRs play in the central nervous system, their extraordinary sensitivity to general anesthetics, particularly the inhalational agents, suggests they may mediate some of the effects of general anesthetics at surgical, or even subanesthetic, concentrations.

scholarly journals Cytosolic [Ca2+] regulation of InsP3-evoked puffs

2012 ◽  
Vol 449 (1) ◽  
pp. 167-173 ◽  
Author(s):  
Michiko Yamasaki-Mann ◽  
Angelo Demuro ◽  
Ian Parker
Keyword(s):  
Plasma Membrane ◽  
Nicotinic Acetylcholine Receptors ◽  
Acetylcholine Receptors ◽  
Xenopus Oocytes ◽  
Single Channel ◽  
Building Blocks ◽  
Electrochemical Gradient ◽  
Intact Cells ◽  
Nicotinic Acetylcholine ◽  
Insp3 Receptors

InsP3-mediated puffs are fundamental building blocks of cellular Ca2+ signalling, and arise through the concerted opening of clustered InsP3Rs (InsP3 receptors) co-ordinated via Ca2+-induced Ca2+ release. Although the Ca2+ dependency of InsP3Rs has been extensively studied at the single channel level, little is known as to how changes in basal cytosolic [Ca2+] would alter the dynamics of InsP3-evoked Ca2+ signals in intact cells. To explore this question, we expressed Ca2+-permeable channels (nicotinic acetylcholine receptors) in the plasma membrane of voltage-clamped Xenopus oocytes to regulate cytosolic [Ca2+] by changing the electrochemical gradient for extracellular Ca2+ entry, and imaged Ca2+ liberation evoked by photolysis of caged InsP3. Elevation of basal cytosolic [Ca2+] strongly increased the amplitude and shortened the latency of global Ca2+ waves. In oocytes loaded with EGTA to localize Ca2+ signals, the number of sites at which puffs were observed and the frequency and latency of puffs were strongly dependent on cytosolic [Ca2+], whereas puff amplitudes were only weakly affected. The results of the present study indicate that basal cytosolic [Ca2+] strongly affects the triggering of puffs, but has less of an effect on puffs once they have been initiated.

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