Photo-antagonism of the GABAA receptor

Neurotransmitter receptor trafficking is fundamentally important for synaptic transmission and neural network activity. GABAA receptors and inhibitory synapses are vital components of brain function, yet much of our knowledge regarding receptor mobility and function at inhibitory synapses is derived indirectly from using recombinant receptors, antibody-tagged native receptors and pharmacological treatments. Here we describe the use of a set of research tools that can irreversibly bind to and affect the function of recombinant and neuronal GABAA receptors following ultraviolet photoactivation. These compounds are based on the competitive antagonist gabazine and incorporate a variety of photoactive groups. By using site-directed mutagenesis and ligand-docking studies, they reveal new areas of the GABA binding site at the interface between receptor β and α subunits. These compounds enable the selected inactivation of native GABAA receptor populations providing new insight into the function of inhibitory synapses and extrasynaptic receptors in controlling neuronal excitation

Use of interrupter technique in assessment of bronchial responsiveness in normal subjects

BACKGROUND: A number of subjects, especially the very young and the elderly, are unable to cooperate and to perform forced expiratory manoeuvres in the evaluation of bronchial hyperresponsiveness (BHR). The objective of our study was to investigate the use of the interrupter technique as a method to measure the response to provocation and to compare it with the conventional PD(20 )FEV(1). METHODS: We studied 170 normal subjects, 100 male and 70 female (mean ± SD age, 38 ± 8.5 and 35 ± 7.5 years, respectively), non-smoking from healthy families. These subjects had no respiratory symptoms, rhinitis or atopic history. A dosimetric cumulative inhalation of methacholine was used and the response was measured by the dose which increases baseline end interruption resistance by 100% (PD(100)Rint, EI) as well as by percent dose response ratio (DRR). RESULTS: BHR at a cut-off level of 0.8 mg methacholine exhibited 31 (18%) of the subjects (specificity 81.2%), 21 male and 10 female, while 3% showed a response in the asthmatic range. The method was reproducible and showed good correlation with PD(20)FEV(1 )(r = 0.76, p < 0.005), with relatively narrow limits of agreement at -1.39 μmol and 1.27 μmol methacholine, respectively, but the interrupter methodology proved more sensitive than FEV(1 )in terms of reactivity (DRR). CONCLUSIONS: Interrupter methodology is clinically useful and may be used to evaluate bronchial responsiveness in normal subjects and in situations when forced expirations cannot be performed

Theoretically nanoscale study on ionization of muscimol nano drug in aqueous solution

In the present work, acid dissociation constant (pKa) values of muscimol derivatives were calculated using the Density Functional Theory (DFT) method. In this regard, free energy values of neutral, protonated and deprotonated species of muscimol were calculated in water at the B3LYP/6-31G(d) basis sets. The hydrogen bond formation of all species had been analyzed using the Tomasi's method. It was revealed that the theoretically calculated pKa values were in a good agreement with the existing experimental pKa values, which were determined from capillary electrophoresis, potentiometric titration and UV-visible spectrophotometric measurements

A Unified Model of the GABA(A) Receptor Comprising Agonist and Benzodiazepine Binding Sites

We present a full-length α(1)β(2)γ(2) GABA receptor model optimized for agonists and benzodiazepine (BZD) allosteric modulators. We propose binding hypotheses for the agonists GABA, muscimol and THIP and for the allosteric modulator diazepam (DZP). The receptor model is primarily based on the glutamate-gated chloride channel (GluCl) from C. elegans and includes additional structural information from the prokaryotic ligand-gated ion channel ELIC in a few regions. Available mutational data of the binding sites are well explained by the model and the proposed ligand binding poses. We suggest a GABA binding mode similar to the binding mode of glutamate in the GluCl X-ray structure. Key interactions are predicted with residues α(1)R66, β(2)T202, α(1)T129, β(2)E155, β(2)Y205 and the backbone of β(2)S156. Muscimol is predicted to bind similarly, however, with minor differences rationalized with quantum mechanical energy calculations. Muscimol key interactions are predicted to be α(1)R66, β(2)T202, α(1)T129, β(2)E155, β(2)Y205 and β(2)F200. Furthermore, we argue that a water molecule could mediate further interactions between muscimol and the backbone of β(2)S156 and β(2)Y157. DZP is predicted to bind with interactions comparable to those of the agonists in the orthosteric site. The carbonyl group of DZP is predicted to interact with two threonines α(1)T206 and γ(2)T142, similar to the acidic moiety of GABA. The chlorine atom of DZP is placed near the important α(1)H101 and the N-methyl group near α(1)Y159, α(1)T206, and α(1)Y209. We present a binding mode of DZP in which the pending phenyl moiety of DZP is buried in the binding pocket and thus shielded from solvent exposure. Our full length GABA(A) receptor is made available as Model S1