Binding of κ-Conotoxin PVIIA to Shaker K+ Channels Reveals Different K+ and Rb+ Occupancies within the Ion Channel Pore

The x-ray structure of the KcsA channel at different [K+] and [Rb+] provided insight into how K+ channels might achieve high selectivity and high K+ transit rates and showed marked differences between the occupancies of the two ions within the ion channel pore. In this study, the binding of κ-conotoxin PVIIA (κ-PVIIA) to Shaker K+ channel in the presence of K+ and Rb+ was investigated. It is demonstrated that the complex results obtained were largely rationalized by differences in selectivity filter occupancy of this 6TM channels as predicted from the structural work on KcsA. κ-PVIIA inhibition of the Shaker K+ channel differs in the closed and open state. When K+ is the only permeant ion, increasing extracellular [K+] decreases κ-PVIIA affinity for closed channels by decreasing the “on” binding rate, but has no effect on the block of open channels, which is influenced only by the intracellular [K+]. In contrast, extracellular [Rb+] affects both closed- and open-channel binding. As extracellular [Rb+] increases, (a) binding to the closed channel is slightly destabilized and acquires faster kinetics, and (b) open channel block is also destabilized and the lowest block seems to occur when the pore is likely filled only by Rb+. These results suggest that the nature of the permeant ions determines both the occupancy and the location of the pore site from which they interact with κ-PVIIA binding. Thus, our results suggest that the permeant ion(s) within a channel pore can determine its functional and pharmacological properties.

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Ion Channel Pore Sizing in Patch-Clamp Experiments

Springer Protocols Handbooks - Patch Clamp Techniques ◽

10.1007/978-4-431-53993-3_26 ◽

2012 ◽

pp. 389-402 ◽

Cited By ~ 1

Author(s):

Ravshan Z. Sabirov ◽

Yasunobu Okada

Keyword(s):

Patch Clamp ◽

Ion Channel ◽

Channel Pore ◽

Ion Channel Pore

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Selective Proton-Mediated Transport by Electrogenic K+-Binding Macrocycles

Chemistry ◽

10.3390/chemistry2010003 ◽

2020 ◽

Vol 2 (1) ◽

pp. 11-21

Author(s):

Yu-Hao Li ◽

Shao-Ping Zheng ◽

Dawei Wang ◽

Mihail Barboiu

Keyword(s):

Ion Channels ◽

Therapeutic Agents ◽

K Channel ◽

Transport Activity ◽

Selective Activation ◽

K Channels ◽

High K ◽

Carbonyl Cyanide ◽

Self Assembled ◽

K Transport

Synthetic K+-binding macrocycles have potential as therapeutic agents for diseases associated with KcsA K+ channel dysfunction. We recently discovered that artificial self-assembled n-alkyl-benzoureido-15-crown-5-ether form selective ion-channels for K+ cations, which are highly preferred to Na+ cations. Here, we describe an impressive selective activation of the K+ transport via electrogenic macrocycles, stimulated by the addition of the carbonyl cyanide-4-(trifluoromethoxy) phenylhydrazone (FCCP) proton carrier. The transport performances show that both the position of branching or the size of appended alkyl arms favor high transport activity and selectivity SK+/Na+ up to 48.8, one of the best values reported up to now. Our study demonstrates that high K+/Na+ selectivity obtained with natural KcsA K+ channels is achievable using simpler artificial macrocycles displaying constitutional functions.

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Architecture of the Alzheimer?s A�P Ion Channel Pore

The Journal of Membrane Biology ◽

10.1007/s00232-003-0638-7 ◽

2004 ◽

Vol 197 (1) ◽

pp. 33-48 ◽

Cited By ~ 70

Author(s):

N. Arispe

Keyword(s):

Ion Channel ◽

Channel Pore ◽

Ion Channel Pore

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Piezo1 ion channel pore properties are dictated by C-terminal region

Nature Communications ◽

10.1038/ncomms8223 ◽

2015 ◽

Vol 6 (1) ◽

Cited By ~ 102

Author(s):

Bertrand Coste ◽

Swetha E. Murthy ◽

Jayanti Mathur ◽

Manuela Schmidt ◽

Yasmine Mechioukhi ◽

...

Keyword(s):

Ion Channel ◽

Terminal Region ◽

Channel Pore ◽

Ion Channel Pore ◽

Pore Properties

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Docosahexaenoic Acid-Induced Vasorelaxation in Hypertensive Rats: Mechanisms of Action

Biological Research For Nursing ◽

10.1177/109980040000200202 ◽

2000 ◽

Vol 2 (2) ◽

pp. 85-95 ◽

Cited By ~ 41

Author(s):

Mary B. Engler ◽

Marguerite M. Engler

Keyword(s):

Nitric Oxide ◽

Inhibitory Effect ◽

Significant Inhibition ◽

Isometric Tension ◽

K Channel ◽

Omega 3 ◽

Hypertensive Rats ◽

Blood Pressure Lowering Effect ◽

K Channels ◽

High K

The authors investigated the vasorelaxant properties of the omega-3 fatty acid, docosahexaenoic (DHA, 22:6n-3), and the possible involvement of endothelium-derived nitric oxide, prostanoids, opening of K+ channels, and/or modulation of calcium-mediated events. Isolated aorta from male spontaneously hypertensive rats (SHR) (age 16-17 weeks) were used to measure isometric tension. DHA-induced (1-100 mol/l) relaxation was examined following contraction to norepinephrine (NE) (10– 6 mol/l) or high-K+ (80 mmol/l) solution in the presence and absence of various inhibitors and calcium-containing solution. DHA acid induced a significant vasorelaxant effect in both NE and high-K+-induced contracted SHR aortic rings, although DHA relaxations were greater in high-K+-induced contracted rings. In the absence of extracellular calcium, DHA (5-30 mol/l) inhibited the initial phasic and sustained components of NE-induced contraction under different conditions. Inhibition of nitric oxide synthesis by N•-nitro-L-arginine methyl ester hydrochloride (100 mol/l) had no effect on DHA relaxations; however, indomethacin or nifedipine caused significant inhibition at• 30 mol/l DHA. The K+ channel blocker, glibenclamide, but not tetraethyl-ammonium, also had an inhibitory effect on DHA-induced relaxation. These results indicate that DHA’s vasorelaxant actions in SHR aorta are independent of endothelium-derived nitric oxide; however, at DHA concentrations• 30 mol/l, vasodilatory prostanoids that activate AT Psensitive K+ channels (KATP) may be involved. At lower concentrations, DHA-induced relaxation appears to be attributed to modulation of intracellular Ca2+release and L-type Ca2+channels in vascular smooth muscle cells. The vasorelaxant properties of DHA may contribute, in part, to the blood pressure–lowering effect of dietary fish oil in this hypertensive model.

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A Conserved Arginine Residue in the Pore Region of an Inward Rectifier K Channel (IRK1) as an External Barrier for Cationic Blockers

The Journal of General Physiology ◽

10.1085/jgp.110.6.665 ◽

1997 ◽

Vol 110 (6) ◽

pp. 665-677 ◽

Cited By ~ 26

Author(s):

Ravshan Z. Sabirov ◽

Tomoko Tominaga ◽

Akiko Miwa ◽

Yasunobu Okada ◽

Shigetoshi Oiki

Keyword(s):

Open Channel ◽

Single Channel ◽

K Channel ◽

Channel Blockers ◽

Channel Activity ◽

Wild Type ◽

K Channels ◽

Blocking Rate ◽

External Barrier ◽

Kir2.1 Channel

The number, sign, and distribution of charged residues in the pore-forming H5 domain for inward-rectifying K channels (IRK1) are different from the otherwise homologous H5 domains of other voltage-gated K channels. We have mutated Arg148, which is perfectly conserved in all inward rectifiers, to His in the H5 of IRK1 (Kir2.1). Channel activity was lost by the mutation, but coexpression of the mutant (R148H) along with the wild-type (WT) mRNA revealed populations of channels with reduced single-channel conductances. Long-lasting and flickery sublevels were detected exclusively for the coexpressed channels. These findings indicated that the mutant subunit formed hetero-oligomers with the WT subunit. The permeability ratio was altered by the mutation, while the selectivity sequence (K+ > Rb+ > NH4+ >> Na+) was preserved. The coexpression made the IRK1 channel more sensitive to extracellular block by Mg2+ and Ca2+, and turned this blockade from a voltage-independent to a -dependent process. The sensitivity of the mutant channels to Mg2+ was enhanced at higher pH and by an increased ratio of mutant:WT mRNA, suggesting that the charge on the Arg site controlled the sensitivity. The blocking rate of open channel blockers, such as Cs+ and Ba2+, was facilitated by coexpression without significant change in the steady state block. Evaluation of the electrical distance to the binding site for Mg2+ or Ca2+ and that to the barrier peak for block by Cs+ or Ba2+ suggest that Arg148 is located between the external blocking site for Mg2+ or Ca2+ and the deeper blocking site for Cs+ or Ba2+ in the IRK1 channel. It is concluded that Arg148 serves as a barrier to cationic blockers, keeping Mg2+ and Ca2+ out from the electric field of the membrane.

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Role of 20-HETE in mediating the effect of dietary K intake on the apical K channels in the mTAL

AJP Renal Physiology ◽

10.1152/ajprenal.2001.280.2.f223 ◽

2001 ◽

Vol 280 (2) ◽

pp. F223-F230 ◽

Cited By ~ 22

Author(s):

Ruimin Gu ◽

Yuan Wei ◽

Houli Jiang ◽

Michael Balazy ◽

Wenhui Wang

Keyword(s):

Gas Chromatography Mass Spectrometry ◽

K Channel ◽

Thick Ascending Limb ◽

Patch Clamp Technique ◽

Open Probability ◽

Deficient Diet ◽

K Channels ◽

Medullary Thick Ascending Limb ◽

High K

We have used the patch-clamp technique to study the effect of dietary K intake on the apical K channels in the medullary thick ascending limb (mTAL) of rat kidneys. The channel activity, defined by the number of channels in a patch and the open probability ( NP o), of the 30- and 70-pS K channels, was 0.18 and 0.11, respectively, in the mTAL from rats on a K-deficient diet. In contrast, NP o of the 30- and 70-pS K channels increased to 0.60 and 0.80, respectively, in the tubules from animals on a high-K diet. The concentration of 20-hydroxyeicosatetraenoic acid (20-HETE) measured with gas chromatography-mass spectrometry was 0.8 pg/μg protein in the mTAL from rats on a high-K diet and increased significantly to 4.6 pg/μg protein in the tubules from rats on a K-deficient diet. Addition of N-methylsulfonyl-12,12-dibromododec-11-enamide (DDMS) or 17-octadecynoic acid (17-ODYA), agents that inhibit the formation of 20-HETE, had no significant effect on the activity of the 30-pS K channels. However, DDMS/17-ODYA significantly increased the activity of the apical 70-pS K channel from 0.11 to 0.91 in the mTAL from rats on a K-deficient diet. In contrast, inhibition of the cytochrome P-450 metabolism of arachidonic acid increased NP o from 0.64 to 0.81 in the tubules from animals on a high-K diet. Furthermore, the sensitivity of the 70-pS K channel to 20-HETE was the same between rats on a high-K diet and on a K-deficient diet. Finally, the pretreatment of the tubules with DDMS increased NP o of the 70-pS K channels in the mTAL from rats on a K-deficient diet to 0.76. We conclude that an increase in 20-HETE production is involved in reducing the activity of the apical 70-pS K channels in the mTAL from rats on a K-deficient diet.

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Endothelium-dependent responses in human isolated thyroid arteries from donors

Journal of Endocrinology ◽

10.1677/joe.0.1810379 ◽

2004 ◽

Vol 181 (3) ◽

pp. 379-384 ◽

Cited By ~ 16

Author(s):

B Torondel ◽

JM Vila ◽

G Segarra ◽

P Lluch ◽

P Medina ◽

...

Keyword(s):

Nitric Oxide ◽

Dominant Role ◽

Human Thyroid ◽

K Channel ◽

Organ Donors ◽

Inhibitory Effects ◽

K Channels ◽

High K ◽

Inwardly Rectifying ◽

Endothelium Dependent Relaxation

The functional properties of the endothelium of human thyroid arteries remain unexplored. We investigated the intervention of nitric oxide (NO), prostacyclin (PGI(2)) and endothelium-derived hyperpolarizing factor (EDHF) in the responses to acetylcholine and noradrenaline in isolated thyroid arteries obtained from multi-organ donors. Artery rings were suspended in organ baths for isometric recording of tension. The contribution of NO, PGI(2) and EDHF to endothelium-dependent relaxation was determined by the inhibitory effects of N(G)-monomethyl-L-arginine (L-NMMA), indomethacin, and K(+) channel inhibitors respectively. Acetylcholine induced concentration-dependent relaxation; this effect was not modified by indomethacin and was only partly reduced by L-NMMA, but was abolished in endothelium-denuded rings. The relaxation resistant to indomethacin and L-NMMA was abolished by using either apamin combined with charybdotoxin, ouabain plus barium, or a high-K(+) solution. Noradrenaline induced concentration-dependent contractions which were of greater magnitude in arteries denuded of endothelium or in the presence of L-NMMA.In conclusion, the results indicate that in human thyroid arteries the endothelium significantly modulates responses to acetylcholine and noradrenaline through the release of NO and EDHF. EDHF plays a dominant role in acetylcholine-induced relaxation through activation of Ca(2+)-activated K(+) channels, inwardly rectifying K(+) channels and Na(+)-K(+)-ATPase.

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