GABAA Receptor-Mediated Sleep-Promoting Effect of Saaz-Saphir Hops Mixture Containing Xanthohumol and Humulone.

Hops contain flavonoids that have sedative and sleep-promoting activities such as α-acid, ß-acid, and xanthohumol. In this study, the sleep-enhancing activity of a Saaz-Saphir hops mixture was measured. In the caffeine-induced insomnia model, the administration of a Saaz-Saphir mixture increased the sleep time compared to Saaz or Saphir administration alone, which was attributed to the increase in NREM sleep time by the δ-wave increase. Oral administration of the Saaz-Saphir mixture for 3 weeks increased the γ-amino butyric acid (GABA) content in the brain and increased the expression of the GABAA receptor. As the GABA antagonists picrotoxin and bicuculline showed a decrease in sleep activity, it was confirmed that the GABAA receptor was involved in the Saaz-Saphir mixture activity. In addition, the GABAA receptor antagonist also reduced the sleep activity induced by xanthohumol and humulone contained in the Saaz-Saphir mixture. Therefore, xanthohumol and humulone contained in the Saaz-Saphir mixture showed sleep-promoting activity mediated by the GABAA receptors. The mixture of the Saaz and Saphir hop varieties may thus help mitigate sleep disturbances compared to other hop varieties.

Synthesis of 4,6-diphenylpyrimidin-2-ol derivatives as new benzodiazepine receptor ligands.

Benzodiazepines (BZDs) have been widely used in neurological disorders such as insomnia, anxiety, and epilepsy. The use of classical BZDs, e.g., diazepam, has been limited due to adverse effects such as interaction with alcohol, ataxia, amnesia, psychological and physical dependence, and tolerance. In the quest for new benzodiazepine agonists with more selectivity and low adverse effects, novel derivatives of 4,6-diphenylpyrimidin-2-ol were designed, synthesized, and evaluated. In this series, compound 2, 4-(2-(benzyloxy)phenyl)-6-(4-fluorophenyl)pyrimidin-2-ol, was the most potent analogue in radioligand binding assay with an IC50 value of 19 nM compared to zolpidem (IC50 = 48 nM), a nonbenzodiazepine central BZD receptor (CBR) agonist. Some compounds with a variety of affinities in radioligand receptor binding assay were selected for in vivo evaluations. Compound 3 (IC50 = 25 nM), which possessed chlorine instead of fluorine in position 4 of the phenyl ring, exhibited an excellent ED50 value in most in vivo tests. Proper sedative-hypnotic effects, potent anticonvulsant activity, appropriate antianxiety effect, and no memory impairment probably served compound 3, a desirable candidate as a benzodiazepine agonist. The pharmacological effects of compound 3 were antagonized by flumazenil, a selective BZD receptor antagonist, confirming the BZD receptors' involvement in the biological effects of the novel ligand.

Novel GABAA Agonist Entities: Pharmacological Investigation and Molecular Modeling Study of Thiazolo- and Thiadiazolo-[3,2-a][1,3]diazepine Analogs.

Thiazolo- and thiadiazolo-[3,2-a][1,3]diazepines and their patented derivatives, tested with diverse CNS pharmacological activities, constitute an important class of compounds for new drug development. Therefore, research efforts were continued to design, synthesize, and evaluate compounds for their ultra-short, short-acting hypnotic, anticonvulsant, and neuromuscular blocking activities. The present review provides a summary of the work accomplished by these heterocycles and their biological evaluation.

A multiorganism pipeline for antiseizure drug discovery: Identification of chlorothymol as a novel γ-aminobutyric acidergic anticonvulsant.

OBJECTIVE: Current medicines are ineffective in approximately one-third of people with epilepsy. Therefore, new antiseizure drugs are urgently needed to address this problem of pharmacoresistance. However, traditional rodent seizure and epilepsy models are poorly suited to high-throughput compound screening. Furthermore, testing in a single species increases the chance that therapeutic compounds act on molecular targets that may not be conserved in humans. To address these issues, we developed a pipeline approach using four different organisms. METHODS: We sequentially employed compound library screening in the zebrafish, Danio rerio, chemical genetics in the worm, Caenorhabditis elegans, electrophysiological analysis in mouse and human brain slices, and preclinical validation in mouse seizure models to identify novel antiseizure drugs and their molecular mechanism of action. RESULTS: Initially, a library of 1690 compounds was screened in an acute pentylenetetrazol seizure model using D rerio. From this screen, the compound chlorothymol was identified as an effective anticonvulsant not only in fish, but also in worms. A subsequent genetic screen in C elegans revealed the molecular target of chlorothymol to be LGC-37, a worm γ-aminobutyric acid type A (GABAA ) receptor subunit. This GABAergic effect was confirmed using in vitro brain slice preparations from both mice and humans, as chlorothymol was shown to enhance tonic and phasic inhibition and this action was reversed by the GABAA receptor antagonist, bicuculline. Finally, chlorothymol exhibited in vivo anticonvulsant efficacy in several mouse seizure assays, including the 6-Hz 44-mA model of pharmacoresistant seizures. SIGNIFICANCE: These findings establish a multiorganism approach that can identify compounds with evolutionarily conserved molecular targets and translational potential, and so may be useful in drug discovery for epilepsy and possibly other conditions.

A Structure-Activity Relationship Comparison of Imidazodiazepines Binding at Kappa, Mu, and Delta Opioid Receptors and the GABAA Receptor.

Analgesic and anti-inflammatory properties mediated by the κ opioid receptor (KOR) have been reported for oxadiazole imidazodiazepines. Affinities determined by radioligand competition assays of more than seventy imidazodiazepines using cell homogenates from HEK293 cells that overexpress KOR, µ opioid receptor (MOR), and δ opioid receptor (DOR) are presented. Affinities to synaptic, benzodiazepine-sensitive receptors (BZR) were determined with rat brain extract. The highest affinity for KOR was recorded for GL-I-30 (Ki of 27 nM) and G-protein recruitment was observed with an EC50 of 32 nM. Affinities for MOR and DOR were weak for all compounds. Ester and amide imidazodiazepines were among the most active KOR ligands but also competed with 3H-flunitrazepam for brain extract binding, which is mediated predominately by gamma aminobutyric acid type A receptors (GABAAR) of the α1-3ß2-3γ1-2 subtypes. Imidazodiazepines with carboxylic acid and primary amide groups did not bind KOR but interacted strongly with GABAARs. Pyridine substitution reduced KOR affinity. Oxadiazole imidazodiazepines exhibited good KOR binding and interacted weakly with BZR, whereas oxazole imidazodiazepines were more selective towards BZR. Compounds that lack the imidazole moiety, the pendent phenyl, or pyridine substitutions exhibited insignificant KOR affinities. It can be concluded that a subset of imidazodiazepines represents novel KOR ligands with high selectivity among opioid receptors.

Image-Based Marker-Free Screening of GABAA Agonists, Antagonists, and Modulators.

The ionotropic GABAA receptors represent the main target for different groups of widely used drugs having hypnotic and anxiolytic effects. So far, most approaches used to assess GABA activity involve invasive low -throughput electrophysiological techniques or rely on fluorescent dyes, preventing the ability to conduct noninvasive and thus nonperturbing screens. To address this limitation, we have developed an automated marker-free cell imaging method, based on digital holographic microscopy (DHM). This technology allows the automatically screening of compounds in multiple plates without having to label the cells or use special plates. This methodological approach was first validated by screening the GABAA receptor expressed in HEK cells using a selection of active compounds in agonist, antagonist, and modulator modes. Then, in a second blind screen of a library of 3041 compounds (mostly composed of natural products), 5 compounds having a specific agonist action on the GABAA receptor were identified. The hits validated from this unbiased screen were the natural products muscimol, neurosteroid alphaxalone, and three compounds belonging to the avermectin family, all known for having an agonistic effect on the GABAA receptor. The results obtained were exempt from false negatives (structurally similar unassigned hits), and false-positive hits were detected and discarded without the need for performing electrophysiological measurements. The outcome of the screen demonstrates the applicability of our screening by imaging method for the discovery of new chemical structures, particularly regarding chemicals interacting with the ionotropic GABAA receptor and more generally with any ligand-gated ion channels and transporters.

Subtype Selective γ-Aminobutyric Acid Type A Receptor (GABAAR) Modulators Acting at the Benzodiazepine Binding Site: An Update.

γ-Aminobutyric acid (GABA) is the major inhibitory neurotransmitter within the central nervous system (CNS) with fast, transsynaptic, and modulatory extrasynaptic effects being mediated by the ionotropic GABA type A receptors (GABAARs). These receptors are of particular interest because they are the molecular target of a number of pharmacological agents, of which the benzodiazepines (BZDs), such as diazepam, are the best described. The anxiolytic, sedating, and myorelaxant effects of BZDs are mediated by separate populations of GABAARs containing either α1, α2, α3, or α5 subunits and the molecular dissection of the pharmacology of BZDs indicates that subtype-selective GABAAR modulators will have novel pharmacological profiles. This is best exemplified by α2/α3-GABAAR positive allosteric modulators (PAMs) and α5-GABAAR negative allosteric modulators (NAMs), which were originally developed as nonsedating anxiolytics and cognition enhancers, respectively. This review aims to summarize the current state of the field of subtype-selective GABAAR modulators acting via the BZD binding site and their potential clinical indications.

Steady-state activation of the high-affinity isoform of the α4ß2δ GABAA receptor.

Activation of GABAA receptors consisting of α4, ß2 (or ß3), and δ subunits is a major contributor to tonic inhibition in several brain regions. The goal of this study was to analyze the function of the α4ß2δ receptor in the presence of GABA and other endogenous and clinical activators and modulators under steady-state conditions. We show that the receptor has a high constitutive open probability (~0.1), but is only weakly activated by GABA that has a maximal peak open probability (POpen,peak) of 0.4, taurine (maximal POpen,peak = 0.4), or the endogenous steroid allopregnanolone (maximal POpen,peak = 0.2). The intravenous anesthetic propofol is a full agonist (maximal POpen,peak = 0.99). Analysis of currents using a cyclic three-state Resting-Active-Desensitized model indicates that the maximal steady-state open probability of the α4ß2δ receptor is ~0.45. Steady-state open probability in the presence of combinations of GABA, taurine, propofol, allopregnanolone and/or the inhibitory steroid pregnenolone sulfate closely matched predicted open probability calculated assuming energetic additivity. The results suggest that the receptor is active in the presence of physiological concentrations of GABA and taurine, but, surprisingly, that receptor activity is only weakly potentiated by propofol.

A newly developed anesthetic based on a unique chemical core.

Intravenous anesthetic agents are associated with cardiovascular instability and poorly tolerated in patients with cardiovascular disease, trauma, or acute systemic illness. We hypothesized that a new class of intravenous (IV) anesthetic molecules that is highly selective for the slow type of γ-aminobutyric acid type A receptor (GABAAR) could have potent anesthetic efficacy with limited cardiovascular effects. Through in silico screening using our GABAAR model, we identified a class of lead compounds that are N-arylpyrrole derivatives. Electrophysiological analyses using both an in vitro expression system and intact rodent hippocampal brain slice recordings demonstrate a GABAAR-mediated mechanism. In vivo experiments also demonstrate overt anesthetic activity in both tadpoles and rats with a potency slightly greater than that of propofol. Unlike the clinically approved GABAergic anesthetic etomidate, the chemical structure of our N-arylpyrrole derivative is devoid of the chemical moieties producing adrenal suppression. Our class of compounds also shows minimal to no suppression of blood pressure, in marked contrast to the hemodynamic effects of propofol. These compounds are derived from chemical structures not previously associated with anesthesia and demonstrate that selective targeting of GABAAR-slow subtypes may eliminate the hemodynamic side effects associated with conventional IV anesthetics.

Design, synthesis, in silico ADMET profile and GABA-A docking of novel phthalazines as potent anticonvulsants.

A new series of 2-substituted-2,3-dihydrophthalazine-1,4-diones (2- 9) were designed and synthesized to evaluate their anticonvulsant activity. The neurotoxicity was assessed using the rotarod test. Molecular docking was performed for the synthesized compounds to assess their binding affinities as γ-aminobutyric acid A (GABA-A) receptor agonists as a possible mechanism of their anticonvulsant action, to rationalize their anticonvulsant activity in a qualitative way. The data obtained from the molecular modeling was strongly matched with that obtained from the biological screening, which revealed that compounds 5a , 9b , and 9h showed the highest binding affinities toward the GABA-A receptor and also showed the highest anticonvulsant activities with relative potencies of 1.66, 1.63, and 1.61, respectively, compared with diazepam. The most active compounds 5a , 9b , and 9h were further tested against maximal electroshock seizures. Compounds 5a and 9b showed 100% protection at a dose level of 125 µg/kg, while compound 9h exhibited 83.33% protection at the same dose level. A GABA enzymatic assay was performed for these highly active compounds to confirm the obtained results and to explain the possible mechanism for their anticonvulsant action. These agents exerted low neurotoxicity and a high safety margin compared with valproate as a reference drug. Most of our designed compounds exhibited a good ADMET profile.

Synthesis of New GABAA Receptor Modulator with Pyrazolo[1,5-a]quinazoline (PQ) Scaffold.

We previously published a series of 8-methoxypirazolo[1,5-a]quinazolines (PQs) and their 4,5-dihydro derivatives (4,5(H)PQ) bearing the (hetero)arylalkylester group at position 3 as ligands at the γ-aminobutyric type A (GABAA) subtype receptor. Continuing the study in this field, we report here the design and synthesis of 3-(hetero)arylpyrazolo[1,5-a]quinazoline and 3-(hetero)aroylpyrazolo[1,5-a]quinazoline 8-methoxy substituted as interesting analogs of the above (hetero)arylalkylester, in which the shortening or the removal of the linker between the 3-(hetero)aryl ring and the PQ was performed. Only compounds that are able to inhibit radioligand binding by more than 80% at 10 µM have been selected for electrophysiological studies on recombinant α1ß2γ2L GABAA receptors. Some compounds show a promising profile. For example, compounds 6a and 6b are able to modulate the GABAAR in an opposite manner, since 6b enhances and 6a reduces the variation of the chlorine current, suggesting that they act as a partial agonist and an inverse partial agonist, respectively. The most potent derivative was 3-(4-methoxyphenylcarbonyl)-8-methoxy-4,5-dihydropyrazolo[1,5-a] quinazoline 11d, which reaches a maximal activity at 1 µM (+54%), and it enhances the chlorine current at ≥0.01 µM. Finally, compound 6g, acting as a null modulator at α1ß2γ2L, shows the ability to antagonize the full agonist diazepam and the potentiation of CGS 9895 on the new α+/ß- 'non-traditional' benzodiazepine site.

TP003 is a non-selective benzodiazepine site agonist that induces anxiolysis via α2GABAA receptors.

Benzodiazepines (BDZ), which potentiate the action of GABA at four subtypes of GABAA receptors (α1, α2, α3, and α5GABAARs), are highly effective against anxiety disorders, but also cause severe side effects greatly limiting their clinical application. Both, preclinical studies in genetically engineered mice, and preclinical and clinical trials with subtype-selective compounds indicate that undesired effects can in principle be avoided by targeting specific GABAAR subtypes. While there is general consensus that activity at α1GABAARs should be avoided, controversy exists as to whether α2 or α3GABAARs need to be targeted for anxiolysis. While previous experiments in GABAAR point-mutated mice demonstrated a critical role of α2GABAARs, studies solely relying on pharmacological approaches suggested a dominant contribution of α3GABAARs. As most α1GABAAR-sparing BDZ site agonists discriminate little between α2 and α3GABAARs, these claims rest almost exclusively on a single compound, TP003, that has been reported to be a selective α3GABAAR modulator. Here, we have revisited the in vitro pharmacological profile of TP003 and, in addition, tested TP003 in GABAAR triple point-mutated mice, in which only either α1, α2, or α3GABAARs were left BDZ sensitive. These experiments revealed that TP003 behaves as a partial, rather non-selective BDZ site agonist in vitro that acts in vivo through α1, α2, and α3GABAARs (α5GABAAR-mediated effects were not tested). With respect to anxiolysis, our results support a critical contribution of α2GABAARs, but not of α3GABAARs. TP003 should therefore not be considered an α3GABAAR selective agent. Previously published studies using TP003 should be interpreted with caution.

Identification of chemically diverse GABAA agonists as potential anti-epileptic agents using structure-guided virtual screening, ADMET, quantum mechanics and clinical validation through off-target analysis.

Development of resistance against existing anti-epileptic drugs has alarmed the scientific innovators to find novel potential chemical starting points for the treatment of epilepsy and GABAA inhibition is a promising drug target strategy against epilepsy. The crystal structure of a subtype-selective ß3-homopentameric ligand-gated ion channel of GABAA receptor has been used for the first time for screening the Asinex library for discovery of GABAA agonists as potential anti-epileptic agents. Co-crystallized ligand established the involvement of part of the ß7-ß8 loop (Glu155 and Tyr157) and ß9-ß10 loop (Phe200 and Tyr205) residues as the crucial amino acids in effective binding, an essential feature, being hydrogen bond or ionic interaction with Glu155 residue. Top ranked hits were further subjected to binding energy estimation, ADMET analysis and ligand efficiency matric calculations as consecutive filters. About 19 compounds qualifying all parameters possessed interaction of one positively charged group with Glu155 with good CNS drug-like properties. Simulation studies were performed on the apo protein, its complex with co-crystallized ligand and the best hit qualifying all screening parameters. The best hit was also analyzed using Quantum mechanical studies, off-target analysis and hit modification. The off-target analysis emphasized that these agents did not have any other predicted side-effects.

Structure-Dependent Activity of Natural GABA(A) Receptor Modulators.

GABA(A) receptors are ligand-gated ion channels consisting of five subunits from eight subfamilies, each assembled in four hydrophobic transmembrane domains. This pentameric structure not only allows different receptor binding sites, but also various types of ligands, such as orthosteric agonists and antagonists, positive and negative allosteric modulators, as well as second-order modulators and non-competitive channel blockers. A fact, that is also displayed by the variety of chemical structures found for both, synthetic as well as nature-derived GABA(A)-receptor modulators. This review covers the literature for natural GABA(A)-receptor modulators until the end of 2017 and discusses their structure-activity relationship.

Bioisosteres of ethyl 8-ethynyl-6-(pyridin-2-yl)-4H-benzo[f]imidazo [1,5-a][1,4]diazepine-3-carboxylate (HZ-166) as novel alpha 2,3 selective potentiators of GABAA receptors: Improved bioavailability enhances anticonvulsant efficacy.

HZ-166 has previously been characterized as an α2,3-selective GABAA receptor modulator with anticonvulsant, anxiolytic, and anti-nociceptive properties but reduced motor effects. We discovered a series of ester bioisosteres with reduced metabolic liabilities, leading to improved efficacy as anxiolytic-like compounds in rats. In the present study, we evaluated the anticonvulsant effects KRM-II-81 across several rodent models. In some models we also evaluated key structural analogs. KRM-II-81 suppressed hyper-excitation in a network of cultured cortical neurons without affecting the basal neuronal activity. KRM-II-81 was active against electroshock-induced convulsions in mice, pentylenetetrazole (PTZ)-induced convulsions in rats, elevations in PTZ-seizure thresholds, and amygdala-kindled seizures in rats with efficacies greater than that of diazepam. KRM-II-81 was also active in the 6 Hz seizure model in mice. Structural analogs of KRM-II-81 but not the ester, HZ-166, were active in all models in which they were evaluated. We further evaluated KRM-II-81 in human cortical epileptic tissue where it was found to significantly-attenuate picrotoxin- and AP-4-induced increases in firing rate across an electrode array. These molecules generally had a wider margin of separation in potencies to produce anticonvulsant effects vs. motor impairment on an inverted screen test than did diazepam. Ester bioisosters of HZ-166 are thus presented as novel agents for the potential treatment of epilepsy acting via selective positive allosteric amplification of GABAA signaling through α2/α3-containing GABA receptors. The in vivo data from the present study can serve as a guide to dosing parameters that predict engagement of central GABAA receptors.

Delineation of the functional properties and the mechanism of action of AA29504, an allosteric agonist and positive allosteric modulator of GABAA receptors.

The retigabine analog 2-amino-4-[(2,4,6-trimethylbenzylamino)-phenyl]-carbamic acid ethyl ester (AA29504) is a positive allosteric modulator (PAM) of γ-aminobutyric acidA receptors (GABAARs), and the modulator has been used in ex vivo/in vivo studies to probe the physiological roles of native δ-containing GABAARs. In this study, the functional properties and mode of action of AA29504 were investigated at human GABAARs expressed in Xenopus oocytes by two-electrode voltage clamp electrophysiology. AA29504 was found to be an allosteric GABAAR agonist displaying low intrinsic activities at 3-30 µM. AA29504 was essentially equipotent as a PAM at the 13 GABAAR subtypes tested (EC50: 0.45-5.2 µM), however GABA EC5-evoked currents through αßδ subtypes were modulated to substantially higher levels than those through αßγ2S subtypes (relative to GABA Imax). While the δ/γ2S-difference clearly was key for this differential GABA efficacy modulation, studies of the AA29504-mediated modulation of different α4,5,6-containing αß, αßγ2S and αßδ GABAARs revealed the α-subunit identity to be another important determinant. Based on its functional properties at numerous mutant GABAARs and on in silico analysis of its low-energy conformations, AA29504 is proposed to act through an allosteric site in the transmembrane ß(+)/α(-) interface in the GABAAR also targeted by etomidate and several other modulators. In contrast to these modulators, however, AA29504 did not display substantial ß2/ß3-over-ß1 GABAAR preference, which challenges the notion of ligands targeting this site always possessing this subtype-selectivity profile. Hence, the detailed pharmacological profiling of AA29504 both highlights the complexity of allosteric GABAAR modulation and provides valuable information about this modulator as a pharmacological tool.

Path to the Desensitized State of Ligand-Gated Ion Channels: Why Are Inhibitory and Excitatory Receptors Different?

The family of pentameric ligand-gated ion channels (pLGICs) includes both inhibitory and excitatory receptors. Electrophysiological methods have explored the time-dependent ion currents induced by their neurotransmitter agonists. Kinetic modeling requires a minimum of three conformational states: resting, active, and desensitized. However, current traces of inhibitory and excitatory pLGICs differ substantially. Reproducing their basic features requires different state connectivity: whether the desensitized state is accessed from the resting or active state. It is surprising that a property as fundamental as state connectivity would differ within the same family. So, we explore the possibility that the connectivity is the same, but corresponding states differ in function: Analogous states on the free energy landscape have similar structure, but differ in ion conductivity, free energies, and agonist binding affinities. This hypothesis is tested using a kinetic model in which agonist and anesthetics modulate the receptor free energy landscape by adsorbing to the membrane in which the receptor is embedded. It was previously shown that even with only three states, the complex behavior observed for GABAAR is reproduced, including its response to anesthetics. It is demonstrated here that this hypothesis accounts for an important difference between inhibitory and excitatory receptors: their opposite responses to inhalation anesthetics.

Discovery of the first low-shift positive allosteric modulators for the muscarinic M1 receptor.

Positive modulation of the muscarinic M1-receptor has for a long time attracted scientists and drug developers for the potential treatment of Alzheimer's disease or Schizophrenia. The precognitive potential of M1 activation has however not been clinically demonstrated as a result of side effects associated both with agonists and positive allosteric modulators (PAM's) of the M1-receptor. To avoid excessive activation of the M1-receptor we have designed a new screening format and developed the first low-shift positive allosteric modulators for the M1 receptor. Low-shift PAM's offer the potential of "use-dependent" attenuation of transmitter-signaling while avoiding pseudo-agonistic behavior in vivo as a common limitation of the so far described high-shift PAM's. With these novel M1-PAM's, the M1 receptor is potentially the first GPCR for which both, high- and low shift PAM's have become available.

The synthetic neuroactive steroid SGE-516 reduces seizure burden and improves survival in a Dravet syndrome mouse model.

Dravet syndrome is an infant-onset epileptic encephalopathy with multiple seizure types that are often refractory to conventional therapies. Treatment with standard benzodiazepines like clobazam, in combination with valproate and stiripentol, provides only modest seizure control. While benzodiazepines are a first-line therapy for Dravet syndrome, they are limited by their ability to only modulate synaptic receptors. Unlike benzodiazepines, neuroactive steroids potentiate a wider-range of GABAA receptors. The synthetic neuroactive steroid SGE-516 is a potent positive allosteric modulator of both synaptic and extrasynaptic GABAA receptors. Prior work demonstrated anticonvulsant activity of SGE-516 in acute seizure assays in rodents. In this study, we evaluated activity of SGE-516 on epilepsy phenotypes in the Scn1a +/- mouse model that recapitulates many features of Dravet syndrome, including spontaneous seizures, premature death and seizures triggered by hyperthermia. To evaluate SGE-516 in Scn1a +/- mice, we determined the effect of treatment on hyperthermia-induced seizures, spontaneous seizure frequency and survival. SGE-516 treatment protected against hyperthermia-induced seizures, reduced spontaneous seizure frequency and prolonged survival in the Scn1a +/- mice. This provides the first evidence of SGE-516 activity in a mouse model of Dravet syndrome, and supports further investigation of neuroactive steroids as potential anticonvulsant compounds for refractory epilepsies.

Efficient synthesis of the GABAA receptor agonist trans-4-aminocrotonic acid (TACA).

Investigations into the pharmacology of different types of cys-loop GABA receptor have relied for years on the chemical modification of GABA-like compounds. The GABA metabolite GABOB is an attractive molecule to modify due to its convenient chemical structure. In the process of developing new GABA-mimic compounds from GABOB as a starting compound three small molecule GABA derivatives were synthesized using a variety of chemical transformations. Amongst these, a new and reliable method to synthesize TACA (trans-4-aminocrotonic acid) is reported.