Single-channel properties of a G-protein-coupled inward rectifier potassium channel in brain neurons

1996 ◽  
Vol 75 (1) ◽  
pp. 318-328 ◽  
Author(s):  
J. J. Grigg ◽  
T. Kozasa ◽  
Y. Nakajima ◽  
S. Nakajima
Keyword(s):  
G Protein ◽  
Single Channel ◽  
Inward Rectifier ◽  
Equilibrium Potential ◽  
K Channel ◽  
Open Probability ◽  
K Concentration ◽  
Inwardly Rectifying ◽  
G Protein Coupled ◽  
Cytoplasmic Side

1. In cultured rat locus coeruleus neurons, somatostatin or met-enkephalin induces an inwardly rectifying K+ conductance. This inward rectifier was analyzed at the single-channel level. 2. Using the inside-out patch-clamp, guanosine 5'-triphosphate (GTP) application to the cytoplasmic side in the presence of somatostatin or met-enkephalin in the pipette produced a large increase in channel activity, which disappeared on switching from GTP to guanosine 5'-diphosphate. 3. The unitary conductance was approximately 30 pS at -95 mV with an extracellular K+ concentration of 156 mM and an intracellular K+ concentration of 124 mM at 23 degrees C. The channel showed burst behavior, and the closed time histogram was fit by two exponentials, with the fast time constant being 0.4 ms. The burst time histogram was also fit by two exponentials, with time constants of 0.24 and 2.0 ms (at 10 nM somatostatin). When the somatostatin concentration was changed from 500 to 1 nM, the kinetic behavior of the channel did not change, except that the open probability of the patch was decreased. 4. The current-voltage relation of the unitary channel current showed inward rectification. The reversal potential coincided with the K+ equilibrium potential, and it shifted according to a change in the K+ equilibrium potential. 5. In the presence of external somatostatin, the application of guanosine 5'-O-(3-thiotriphosphate) to the cytoplasmic side induced an irreversible activation of this channel. 6. These results indicate that this K+ channel is the microscopic counterpart of the somatostatin- or met-enkephalin-induced inwardly rectifying K+ current in whole cell recording, and that the channel is activated by a G protein without a diffusible second messenger. Thus this channel is identified as a neuronal G-protein-coupled inward rectifier K+ channel. 7. Analysis of the burst behavior, based on a close-close-open kinetic model, revealed that there are at least four states in the K+ channel, a short gap, a longer closing, a short opening, and a long opening, and that the neuronal inward rectifier is activated at faster rates than the atrial inward rectifier.

scholarly journals G protein–coupled receptors differentially regulate glycosylation and activity of the inwardly rectifying potassium channel Kir7.1

2018 ◽  
Vol 293 (46) ◽  
pp. 17739-17753 ◽  
Author(s):  
Sheridan J. Carrington ◽  
Ciria C. Hernandez ◽  
Daniel R. Swale ◽  
Oluwatosin A. Aluko ◽  
Jerod S. Denton ◽  
...  
Keyword(s):  
Potassium Channel ◽  
G Protein ◽  
Single Channel ◽  
Pigment Epithelium ◽  
Retinal Pigment ◽  
G Protein Coupled Receptors ◽  
Open Probability ◽  
Inwardly Rectifying Potassium Channel ◽  
Inwardly Rectifying ◽  
G Protein Coupled

Kir7.1 is an inwardly rectifying potassium channel with important roles in the regulation of the membrane potential in retinal pigment epithelium, uterine smooth muscle, and hypothalamic neurons. Regulation of G protein–coupled inwardly rectifying potassium (GIRK) channels by G protein–coupled receptors (GPCRs) via the G protein βγ subunits has been well characterized. However, how Kir channels are regulated is incompletely understood. We report here that Kir7.1 is also regulated by GPCRs, but through a different mechanism. Using Western blotting analysis, we observed that multiple GPCRs tested caused a striking reduction in the complex glycosylation of Kir7.1. Further, GPCR-mediated reduction of Kir7.1 glycosylation in HEK293T cells did not alter its expression at the cell surface but decreased channel activity. Of note, mutagenesis of the sole Kir7.1 glycosylation site reduced conductance and open probability, as indicated by single-channel recording. Additionally, we report that the L241P mutation of Kir7.1 associated with Lebers congenital amaurosis (LCA), an inherited retinal degenerative disease, has significantly reduced complex glycosylation. Collectively, these results suggest that Kir7.1 channel glycosylation is essential for function, and this activity within cells is suppressed by most GPCRs. The melanocortin-4 receptor (MC4R), a GPCR previously reported to induce ligand-regulated activity of this channel, is the only GPCR tested that does not have this effect on Kir7.1.

scholarly journals Subunit Stoichiometry of a Heteromultimeric G protein-coupled Inward-rectifier K+Channel

1996 ◽  
Vol 271 (48) ◽  
pp. 30524-30528 ◽  
Author(s):  
Scott K. Silverman ◽  
Henry A. Lester ◽  
Dennis A. Dougherty
Keyword(s):  
G Protein ◽  
Inward Rectifier ◽  
K Channel ◽  
Subunit Stoichiometry ◽  
G Protein Coupled

scholarly journals Primary structure and functional expression of a mouse inward rectifier K+ channel and rat G-protein-coupled muscarinic K+ channel.

1995 ◽  
Vol 15 (2) ◽  
pp. 106-113
Author(s):  
Yoshihiro Kubo ◽  
Eitan Reuveny ◽  
Paul A Slesinger ◽  
Timothy J Baldwin ◽  
Yuh Nung Jan ◽  
...  
Keyword(s):  
G Protein ◽  
Primary Structure ◽  
Functional Expression ◽  
Inward Rectifier ◽  
K Channel ◽  
G Protein Coupled

scholarly journals A novel ion conducting route besides the central pore in an inherited mutant of G-protein-gated inwardly rectifying K+ channel

2021 ◽  
Author(s):  
I-Shan Chen ◽  
Jodene Eldstrom ◽  
David Fedida ◽  
Yoshihiro Kubo
Keyword(s):  
G Protein ◽  
Structural Changes ◽  
Single Channel ◽  
Ion Selectivity ◽  
K Channel ◽  
Girk Channels ◽  
Electrode Voltage ◽  
Ion Conducting ◽  
Inwardly Rectifying ◽  
Better Than

G-protein-gated inwardly rectifying K+ (GIRK; Kir3.x) channels play important physiological roles in various organs. Some of the disease-associated mutations of GIRK channels are known to induce loss of K+ selectivity but their structural changes remain unclear. In this study, we investigated the mechanisms underlying the abnormal ion selectivity of inherited GIRK mutants. By the two-electrode voltage-clamp analysis of GIRK mutants heterologously expressed in Xenopus oocytes, we observed that Kir3.2 G156S permeates Li+ better than Rb+, while T154del or L173R of Kir3.2 and T158A of Kir3.4 permeate Rb+ better than Li+, suggesting a unique conformational change in the G156S mutant. Applications of blockers of the selectivity filter (SF) pathway, Ba2+ or Tertiapin-Q (TPN-Q), remarkably increased the Li+-selectivity of Kir3.2 G156S but did not alter those of the other mutants. In single-channel recordings of Kir3.2 G156S expressed in mouse fibroblasts, two types of events were observed, one attributable to a TPN-Q sensitive K+ current and the second a TPN-Q resistant Li+ current. The results show that a novel Li+ permeable and blocker-resistant pathway exists in G156S in addition to the SF pathway. Mutations in the pore helix (PH), S148F and T151A, also induced high Li+ permeation. Our results demonstrate that the mechanism underlying the loss of K+ selectivity of Kir3.2 G156S involves formation of a novel ion permeation pathway besides the SF pathway, which allows permeation of various species of cations.

scholarly journals Interaction Sites of the G Protein β Subunit with Brain G Protein-coupled Inward Rectifier K+Channel

2001 ◽  
Vol 276 (16) ◽  
pp. 12712-12717 ◽  
Author(s):  
Abla M. Albsoul-Younes ◽  
Pamela M. Sternweis ◽  
Peng Zhao ◽  
Hiroko Nakata ◽  
Shigehiro Nakajima ◽  
...  
Keyword(s):  
G Protein ◽  
Inward Rectifier ◽  
K Channel ◽  
Β Subunit ◽  
Interaction Sites ◽  
G Protein Coupled

Biophysical and pharmacological characterization of inwardly rectifying K+ currents in rat spinal cord astrocytes

1995 ◽  
Vol 73 (1) ◽  
pp. 333-346 ◽  
Author(s):  
C. B. Ransom ◽  
H. Sontheimer
Keyword(s):  
Spinal Cord ◽  
Single Channel ◽  
Resting Potential ◽  
Equilibrium Potential ◽  
Dependent Manner ◽  
Rat Spinal Cord ◽  
Open Probability ◽  
Kir Channels ◽  
Inwardly Rectifying ◽  
K Currents

1. Whole cell and cell-attached patch-clamp recordings were obtained from rat spinal cord astrocytes maintained in culture for 6-14 days. It was found that the resting conductance in these astrocytes is primarily due to inwardly rectifying K+ (Kir) channels. 2. Two types of astrocytic Kir channels were identified with single-channel conductances of approximately 28 and approximately 80 pS, respectively. Channels displayed some voltage dependence in their open probability, which was largest (0.8-0.9) near the K+ equilibrium potential (Ek) and decreased at more negative potentials. The resting potential closely followed Ek, so it can be assumed that Kir channels have a high open probability at the resting potential. 3. The conductance of inwardly rectifying K+ currents (Kir) depended strongly on [K+]o and was approximately proportional to the square-root of [K+]o. 4. Kir currents inactivated in a time- and voltage-dependent manner. The Na+ dependence of inactivation was studied with ion substitution experiments. Replacement of [Na+]o with choline or Li+ removed inactivation. This dependence of current inactivation on [Na+]o resembles the previously described block of Kir channels in other systems by [Na+]o. 5. Kir currents were also blocked in a dose-dependent manner by Cs+ (Kd = 189 microM at -140 mV), Ba2+ (Kd = 3.5 microM), and tetraethylammonium (TEA; 90% block at 10 mM) but were insensitive to 4-aminopyridine (4-AP; 5 mM). In the current-clamp mode, Ba2+ and TEA inhibition of Kir currents was associated with a marked depolarization, suggesting that Kir channel activity played a role in the establishment of the negative resting potential typical of astrocytes. 6. These biophysical features of astrocyte inwardly rectifying K+ channels are consistent with those properties required for their proposed involvement in [K+]o clearance: 1) high open probability at the resting potential, 2) increasing conductance with increasing [K+]o, and 3) rectification, e.g., channel closure, at positive potentials. It is proposed, therefore, that the dissipation of [K+]o following neuronal activity is mediated primarily by the activity of astrocytic Kir channels.

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