γ-Aminobutyric Acid Type A (GABAA) Receptor Activation Modulates Tau Phosphorylation

Background Droperidol is used in neuroleptanesthesia and as an antiemetic. Although its antiemetic effect is thought to be caused by dopaminergic inhibition, the mechanism of droperidol's anesthetic action is unknown. Because gamma-aminobutyric acid type A (GABAA) and neuronal nicotinic acetylcholine receptors (nAChRs) have been implicated as putative targets of other general anesthetic drugs, the authors tested the ability of droperidol to modulate these receptors. Methods gamma-Aminobutyric acid type A alpha1beta1gamma2 receptor, alpha7 and alpha4beta2 nAChRs were expressed in Xenopus oocytes and studied with two-electrode voltage clamp recording. The authors tested the ability of droperidol at concentrations from 1 nm to 100 microm to modulate activation of these receptors by their native agonists. Results Droperidol inhibited the GABA response by a maximum of 24.7 +/- 3.0%. The IC50 for inhibition was 12.6 +/- 0.47 nm droperidol. At high concentrations, droperidol (100 microm) activates the GABAA receptor in the absence of GABA. Inhibition of the GABA response is significantly greater at hyperpolarized membrane potentials. The activation of the alpha7 nAChR is also inhibited by droperidol, with an IC50 of 5.8 +/- 0.53 microm. The Hill coefficient is 0.95 +/- 0.1. Inhibition is noncompetitive, and membrane voltage dependence is insignificant. Conclusions Droperidol inhibits activation of both the GABAA alpha1beta1gamma2 and alpha7 nAChR. The submaximal GABA inhibition occurs within a concentration range such that it might be responsible for the anxiety, dysphoria, and restlessness that limit the clinical utility of high-dose droperidol anesthesia. Inhibition of the alpha7 nAChR might be responsible for the anesthetic action of droperidol.

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Propofol at Clinically Relevant Concentrations Increases Neuronal Differentiation But Is Not Toxic to Hippocampal Neural Precursor Cells In Vitro

Anesthesiology ◽

10.1097/aln.0b013e31826f8d86 ◽

2012 ◽

Vol 117 (5) ◽

pp. 1080-1090 ◽

Cited By ~ 28

Author(s):

Jeffrey W. Sall ◽

Greg Stratmann ◽

Jason Leong ◽

Elliott Woodward ◽

Philip E. Bickler

Keyword(s):

Cell Death ◽

Lactate Dehydrogenase ◽

Neuronal Differentiation ◽

Cell Function ◽

Type A ◽

Receptor Activation ◽

Aminobutyric Acid ◽

Neural Precursor ◽

Acid Type

Background Propofol in the early postnatal period has been shown to cause brain cell death. One proposed mechanism for cognitive dysfunction after anesthesia is alteration of neural stem cell function and neurogenesis. We examined the effect of propofol on neural precursor or stem cells (NPCs) grown in vitro. Methods Hippocampal-derived NPCs from postnatal day 2 rats were exposed to propofol or Diprivan. NPCs were then analyzed for bromodeoxyuridine incorporation to measure proliferation. Cell death was measured by lactate dehydrogenase release. Immunocytochemistry was used to evaluate the expression of neuronal and glial markers in differentiating NPCs exposed to propofol. Results Propofol dose dependently increases the release of lactate dehydrogenase from NPCs under both proliferating and differentiating conditions at supraclinical concentrations (more than 7.1 µM). Both Diprivan and propofol had the same effect on NPCs. Propofol-mediated release of lactate dehydrogenase is not inhibited by blocking the γ-aminobutyric acid type A receptor or extracellular calcium influx and is not mediated by caspase-3/7. Direct γ-aminobutyric acid type A receptor activation did not have the same effect. In differentiating NPCs, 6 h of propofol at 2.1 µM increased the number neurons but not glial cells 4 days later. Increased neuronal differentiation was not blocked by bicuculline. Conclusions Only supraclinical concentrations of propofol or Diprivan kill NPCs in culture by a non-γ-aminobutyric acid type A, noncaspase-3 mechanism. Clinically relevant doses of propofol increase neuronal fate choice by a non-γ-aminobutyric acid type A mechanism.

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A Conserved Tyrosine in the β2Subunit M4 Segment Is a Determinant of γ-Aminobutyric Acid Type A Receptor Sensitivity to Propofol

Anesthesiology ◽

10.1097/01.anes.0000278875.36639.2c ◽

2007 ◽

Vol 107 (3) ◽

pp. 412-418 ◽

Cited By ~ 39

Author(s):

James E. Richardson ◽

Paul S. Garcia ◽

Kate K. O'Toole ◽

Jason M. C. Derry ◽

Shannon V. Bell ◽

...

Keyword(s):

Binding Site ◽

Type A ◽

Receptor Activation ◽

Aminobutyric Acid ◽

Gamma Aminobutyric Acid ◽

General Anesthetics ◽

Receptor Function ◽

293 Cells ◽

Acid Type ◽

Normal Sensitivity

Background The gamma-aminobutyric acid type A receptor (GABAA-R) beta subunits are critical targets for the actions for several intravenous general anesthetics, but the precise nature of the anesthetic binding sites are unknown. In addition, little is known about the role the fourth transmembrane (M4) segment of the receptor plays in receptor function. The aim of this study was to better define the propofol binding site on the GABAA-R by conducting a tryptophan scan in the M4 segment of the beta2 subunit. Methods Seven tryptophan mutations were introduced into the C-terminal end of the M4 segment of the GABAA-R beta2 subunit. GABAA-R subunit complementary DNAs were transfected into human embryonic kidney 293 cells grown on glass coverslips. After transfection (36-72 h), coverslips were transferred to a perfusion chamber to assay receptor function. Cells were whole cell patch clamped and exposed to GABA, propofol, etomidate, and pregnenolone. Chemicals were delivered to the cells using two 10-channel infusion pumps and a rapid solution exchanger. Results All tryptophan mutations were well tolerated, and with one exception, all resulted in minimal changes in receptor activation by GABA. One mutation, beta2(Y444W), selectively suppressed the ability of propofol to enhance receptor function while retaining normal sensitivity to etomidate and pregnenolone. Conclusions This is the first report of a mutation that selectively reduces propofol sensitivity without altering the action of etomidate. The reduction in propofol sensitivity is consistent with the loss of a hydrogen bond within the propofol binding site. These results also suggest a possible orientation of the propofol molecule within its binding site.

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Identification of γ-aminobutyric acid type A (GABAA) receptor modulators: HPLC-based activity profiling of Asarum heterotropoides

Planta Medica ◽

10.1055/s-0030-1264920 ◽

2010 ◽

Vol 76 (12) ◽

Author(s):

D Rueda ◽

J Zaugg ◽

S Hering ◽

M Hamburger

Keyword(s):

Gabaa Receptor ◽

Type A ◽

Aminobutyric Acid ◽

Activity Profiling ◽

Acid Type ◽

Asarum Heterotropoides ◽

Receptor Modulators

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An Allosteric Coagonist Model for Propofol Effects on α1β2γ2L γ-Aminobutyric Acid Type A Receptors

Anesthesiology ◽

10.1097/aln.0b013e31823d0c36 ◽

2012 ◽

Vol 116 (1) ◽

pp. 47-55 ◽

Cited By ~ 37

Author(s):

Dirk Ruesch ◽

Elena Neumann ◽

Hinnerk Wulf ◽

Stuart A. Forman

Keyword(s):

Type A ◽

Receptor Activation ◽

Convincing Evidence ◽

Receptor Activity ◽

Aminobutyric Acid ◽

Single Class ◽

Gaba A ◽

Direct Activation ◽

Receptor Sites ◽

Acid Type

Background Propofol produces its major actions via γ-aminobutyric acid type A (GABA(A)) receptors. At low concentrations, propofol enhances agonist-stimulated GABA(A) receptor activity, and high propofol concentrations directly activate receptors. Etomidate produces similar effects, and there is convincing evidence that a single class of etomidate sites mediate both agonist modulation and direct GABA(A) receptor activation. It is unknown if the propofol binding site(s) on GABA(A) receptors that modulate agonist-induced activity also mediate direct activation. Methods GABA(A) α1β2γ2L receptors were heterologously expressed in Xenopus oocytes and activity was quantified using voltage clamp electrophysiology. We tested whether propofol and etomidate display the same linkage between agonist modulation and direct activation of GABA(A) receptors by identifying equiefficacious drug solutions for direct activation. We then determined whether these drug solutions produce equal modulation of GABA-induced receptor activity. We also measured propofol-dependent direct activation and modulation of low GABA responses. Allosteric coagonist models similar to that established for etomidate, but with variable numbers of propofol sites, were fitted to combined data. Results Solutions of 19 μM propofol and 10 μM etomidate were found to equally activate GABA(A) receptors. These two drug solutions also produced indistinguishable modulation of GABA-induced receptor activity. Combined electrophysiological data behaved in a manner consistent with allosteric coagonist models with more than one propofol site. The best fit was observed when the model assumed three equivalent propofol sites. Conclusions Our results support the hypothesis that propofol, like etomidate, acts at GABA(A) receptor sites mediating both GABA modulation and direct activation.

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Multiple assembly signals in γ-aminobutyric acid (type A) receptor subunits combine to drive receptor construction and composition

Biochemical Society Transactions ◽

10.1042/bst0310875 ◽

2003 ◽

Vol 31 (4) ◽

pp. 875-879 ◽

Cited By ~ 29

Author(s):

K. Bollan ◽

L.A. Robertson ◽

H. Tang ◽

C.N. Connolly

Keyword(s):

Gabaa Receptor ◽

Gabaa Receptors ◽

Type A ◽

Aminobutyric Acid ◽

Single Amino Acid ◽

Functional Surface ◽

Surface Receptors ◽

Acid Type ◽

Absolute Requirement ◽

Receptor Assembly

Mammalian γ-aminobutyric acid type A (GABAA) receptors are constructed from a large repertoire of subunits (α1–α6, β1–β3, γ1–γ3, δ, ∊, θ and π) into a pentameric ion channel. GABAA receptor assembly occurs within the endoplasmic reticulum (ER) and involves interactions with chaperone molecules. Only specific subunit combinations can produce functional surface receptors (with a fixed stoichiometry); other subunit combinations are retained within the ER and degraded. Thus, receptor assembly occurs by defined pathways to limit the diversity of GABAA receptors. The key to understanding how receptor diversity is achieved and controlled is the identification of assembly signals capable of distinguishing between other subunit partners. Analysis of an assembly box in α1 (residues 57–68) has revealed an absolute requirement for this region in the assembly of αβ receptors. Furthermore, a selective requirement for a single amino acid (R66) is observed for the assembly of α1β2, but not α1β1 or α1β3, receptors. In addition, we have characterized an assembly signal in the β3 subunit that is capable of driving the assembly of β3, γ2β3 and α1β3 receptors. Interestingly, this signal does not appear to utilize the α1 assembly box, suggesting the presence of alternative assembly signals within the α1 subunit. Although this β3 signal is sufficient to permit the formation of βγ receptors it is not necessary, suggesting that alternative assembly signals also exist within the β3 subunit. These findings support the belief that GABAA receptor assembly occurs via multiple defined pathways that may be determined by subunit availability.

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