Recent Highlights

Publications after retirement

On October 1, 2011, Dr. Sieghart (at that time, age 66) officially retired from his position as Head of the Department of Biochemistry and Molecular Biology of the Nervous System, at the Center for Brain Research, Medical University Vienna, and became Guest Professor at this Center.

Nevertheless he continued to direct his Department until July 25, 2013, when his successor, Tibor Harkany, was appointed Professor at the Center for Brain Research and Head of the Department. At that time, the Department was renamed into „Department of Molecular Neurosciences“.

In the meantime, several scientific papers were published by Dr. Sieghart that were initiated and partially completed before his final retirement on July 25, 2013. The most important of these papers are listed in the section „Main contribution to research„.

Other important papers, however, were completed by Dr. Sieghart only after July 25, 2013. They deal with three different topics:

a) GABA_A receptor subunit distribution in the human brain

b) alpha6-GABA_A receptors and neuropsychiatric disorders

c) up-to-date reviews of the GABA_A receptor field

a) GABA_A receptor subunit distribution in the human brain

The excellent and highly specific antibodies against GABA_A receptor subunits generated by Dr. Sieghart, resulted in multiple important and highly cited papers on the regional, cellular, and subcellular distribution of GABA_A receptor subunits in the brain of rats and mice before his final retirement in 2013 (see „Main contribution to research„).

In the last years before his retirement, Ivan Milenkovic and Harald Stefanits, two MD-PhD students of Dr. Sieghart, investigated whether GABA_A receptor subunit-specific antibodies suitable for immunohistochemistry in the rodent brain, can also be used in the human brain. Some of the antibodies were suitable for that purpose, others were not. This resulted in major efforts of Dr. Sieghart and his students to newly generate and characterize GABA_A receptor-selective antibodies suitable for immunohistochemistry in the human brain. The first applications of these antibodies resulted in 3 publications

– Stojanovic et al., (2016) J Comp Neurol 524, 1805-1824 (22 citations)

This paper for the first time investigated the expression pattern of several GABA_A receptor subunits in human cerebellum and adjacent brain structures not only during brain development but also in the adult human brain.

This study supported previous hints that the distribution of GABA_A receptor subunits might be partially different in rodents and humans. This important finding partially explains some of the discrepancies observed in the behavioral effects of GABA_A receptor subtype-selective drugs in rodents and humans. In addition, these data emphasize that the regional and cellular distribution of GABA_A receptor subtypes in the human brain, but not that in the mouse or rat brain, should be used as a guideline for understanding the human GABAergic system.

– Stefanits et al., (2018) J Comp Neurol 526, 324-348 (28 citations)

This is the first detailed investigation of the immunohistochemical distribution of seven GABA_A receptor subunits in the human amygdala. This important brain region regulates anxiety, stress, reward, mnestic functions, addiction and is involved in epilepsy.

Again there were differences in the GABA_A receptor subunit expression between human and rodent amygdala, supporting the conclusion that the human and not the rodent subunit distribution should be considered when developing subunit-selective drugs that can be used to modulate anxiety without inducing sedation!

– Stefanits et al., (2019) J Neuropathol Exp Neurol 78, 1022-1048 (7 citations)

Here, alterations in GABAA receptor subunit expression in amygdala and entorhinal cortex in human temporal lobe epilepsy are described.

b) alpha6-GABA_A receptors and neuropsychiatric disorders

The identification of the alpha+beta- interface of GABA_A receptors as a novel target for subtype-selective drugs by the group of Dr. Sieghart

– Ramerstorfer et al., 2011, J. Neurosci. 31, 870-877 (116 citations)

resulted in the identification of the first ligands that were highly selective positive allosteric modulators of a6 subunit-containing GABA_A receptors

– Varagic et al., 2013, Br. J. Pharmacol. 169, 384-399 (64 citations)

At that time, the biological function of alpha6-containing GABA_A receptors in the brain was not known. Nevertheless, the identification of alpha6-GABA_A receptor-selective ligands was the starting point for an exciting collaboration with Lih-Chu Chiou from the University of Taiwan. LC Chiou had some evidence indicating that the flavonoid hispidulin isolated from a local herb that remitted a patient´s intractable motor tics, might exert its action via alpha6-containing GABA_A receptors. She read our article and contacted the group of Dr. Sieghart for a possible collaboration.

Together with the group of Jim Cook, who synthesized the required highly selective positive modulators of alpha6-containing GABA_A receptors, Dr. Sieghart started a collaboration with LC Chiou.

Already the first experiments of LC Chiou supported the conclusion that the alpha6-receptor-selective compounds, that in contrast to the classical benzodiazepines were devoid of any sedative, hypnotic, anxiolytic and muscle relaxant properties, are able to excert actions similar to those of hispidulin in assays mimicking tic-diseases. This indicated that alpha6-GABA_A receptor-selective compounds could possibly be used for the treatment of tic-disorders.

In contrast to the highly alpha6-selective compounds identified by Varagic et al., 2013, hispidulin is quite non-selective and also able to interact with most of the other GABA_A receptor subtypes. It was thus decided to concentrate on alpha6-GABA_A receptor-selective compounds for a future drug development.

Based on these preliminary results, an international consortium was assembled, consisting of the groups of Jim Cook (University of Wisconsin, Milwaukee, Wisconsin, USA), LC Chiou (National Taiwan University, Taipei, Taiwan), and Werner Sieghart and Margot Ernst (Center for Brain Research, Vienna, Austria). Margot Ernst, was a previous postdoc and continued the work of W. Sieghart as head of the GABA_A receptor research group at the Center for Brain Research.

The group of Jim Cook synthesized deuterated alpha6-selective compounds to increase their metabolic stability and bioavailability. Margot Ernst investigated their alpha6-selectivity in electrophysiological experiments at various recombinant GABA_A receptor subtypes. LC Chiou investigated whether these compounds could ameliorate a disrupted prepulse inhibition of the startle response (PPI), an animal model mimicking tic-disorders.

alpha6-containing GABAA receptors might also be involved in other neuropsychiatric disorders:

Soon it became clear that a disruption in the prepulse inhibition of the startle response (PPI) not only is involved in tic-disorders, but is also manifested in patients with other neuropsychiatric disorders, such as obsessive-compulsive disorder, attention deficit disorder, panic disorder, nocturnal enuresis, Huntington´s disease, premenstrual dysphoric disorder, mania in bipolar disorder, antisocial personality disorder and schizophrenia. Especially in patients with schizophrenia, PPI disruption is a well-known endophenotype manifestation.

In addition, other evidence indicated that GABA_A receptors containing alpha6-subunits also might play a role in neuropathic pain and in migraine, and possibly also in depression. Experiments investigating a possible role of alpha6-GABA_A receptors in neuropathic pain led to the inclusion of Miroslav M. Savic, Dept. Pharmacology, Faculty of Pharmacy, University of Belgrade, Belgrade, Serbia, into the consortium. Experiments investigating a possible role of alpha6-GABA_A receptors in migraine, led to the inclusion of Pi-Chuan Fan, Division of Neurology, Department of Pediatrics, National Taiwan University Hospital, Taipei, Taiwan, into the consortium.

Finally, the group of Marko M. Mihovilovic from the Institute of Applied Synthetic Chemistry of the Vienna University of Technology also joined the consortium. Based on the results of Varagic et al., 2013, mentioned above, Margot Ernst and Marko Mihovilovic had some idea on how to generate additional compounds that might be selective modulators of alpha6-GABA_A receptors. W. Sieghart financed an initial pilot study to enable their synthesis by the group of Marko Mihovilovic. These and subsequent synthetic efforts led to some other novel alpha6-selective positive allosteric modulators, that later on were included into the patent application.

To coordinate all these different groups, to clarify and communicate their mutual requirements, to determine priorities and ensure a smooth development of the project, Dr. Sieghart was elected as the coordinator of this consortium, aiming for a future patent application on alpha6-selective ligands and their possible clinical application.

The provisional patent application was filed June 3, 2016, in the USA

– PCT/US2016/035761

The patent application was then published on May 17, 2018, Pub. No.: US 2018/0134698 A1

020871-9110-US02 Published Application — 23433488 v1

Work on this patent application resulted in several publications with Dr. Sieghart as a co-author. Dr. Sieghart together with the respective first author and senior author was significantly involved in writing of these publications and strongly contributed to the development of the hypotheses put forward to explain the action of the positive allosteric modulators of alpha6-GABA_A receptors in the respective animal models.

– Knutson et al., (2018) J Med Chem 61, 2422-2446 (38 citations)

In this paper, the design, synthesis, basic pharmacology, and behavioral actions of novel deuterated ligands functionally selective for alpha6-GABA_A receptors, is described. Depending on the position of the introduced deuterium, compounds with improved metabolic stability and enhanced bioavailability were obtained in a non-predictable way.

– Chiou et al., (2018) Br J Pharmacol 175, 2414-2427 (25 citations)

Here, for the first time evidence is provided indicating that cerebellar alpha6-GABA_A receptors are novel therapeutic targets for ameliorating disrupted prepulse inhibition in neuropsychiatric disorders. In addition, a hypothesis on the molecular mechanism of action of alpha6-GABA_A receptors in cerebellar function and in disrupted prepulse inhibition is provided.

– Fan et al., (2018) Neuropharmacol 140, 1-13 (19 citations)

– Tzeng et al., 2020, Neurotherapeutics 18, 569-585 (5 citations)

Here, it is demonstrated that enhancing the activity of alpha6-GABA_A receptors in two different animal models of migraine can inhibit a trigeminal vascular system activation that is involved in the pathogenesis of migraine and reduce pain resembling that of migraine. The effects were similar to those of topiramate, a clinically effective anti-migraine agent. These results suggest that alpha6-GABA_A receptor-selective positive allosteric modulators have the potential to be developed as a novel pharmacotherapy for migraine.

Vasovic et al., (2019) Eur J Pain, 1-12. (21 citations)

Here, it is demonstrated that a deuterated alpha6-selective pyrazoloquinolinone  significantly reduced the signs of neuropathic pain in two protocols of chronic constriction injury of the infraorbital nerve in rats. Neuropathic pain induced by trigeminal nerve damage is poorly controlled by current treatments. The compound investigated is appropriate for repeated administration and might thus represent a novel treatment option against the development and maintenance of trigeminal neuropathic pain.

– Huang et al., 2021, bioRxiv preprint   

c) Important recent reviews of the GABA_A receptor field

Sieghart W (2014) GABA_A receptors in the brain: exciting targets for the development of clinically important drugs. Magyar Tudományos Akademia 2014. ISSN 1419-8959, ISBN 978-963-508-779-2.

•  Sieghart_Werner_jav, Hungarian Academy of Sciences

This is a historical description on how and why Dr. Sieghart entered the field of GABA_A receptor research and how he developed his own research line

• Sieghart W (2015) Allosteric modulation of GABA_A receptors via multiple drug-binding sites. Adv. Pharmacol 72, 53-96 (192 citations)

This review discusses the so far identified drug binding sites at GABA_A receptors and for the first time also available evidence that most if not all drugs interacting with GABA_A receptors can do that via multiple binding sites at the same receptor.

• Sieghart W and Savic M (2018) International Union of Basic and Clinical Pharmacology. VXI: GABA_A receptor subtype- and function-selective ligands: key issues in translation to humans. Pharmacol. Rev. 70, 836-878 (111 citations)

This review critically discusses the methodology used to investigate GABA_A receptor subtype-selectivity and provides the chemical structures and published concentration-response curves from electrophysiological experiments of more than 40 compounds claimed to be GABA_A receptor subtype-selective.

Evidence is provided that most of these compounds are not receptor subtype-selective at all under in vivo conditions. The discrepancy in the behavior of some of these compounds between rodents and man is discussed and tentatively explained and strategies for the development of compounds with much better receptor subtype-selectivity are proposed.

Finally, this comprehensive dataset can now be used for selecting combinations of suitable compounds that can help to draw valid conclusions on the function of GABA_A receptor subtypes in the brain.

This article has been recommended in F1000Prime as being of special significance in its field by F1000 Faculty Members Hanns Möhler and Ryan Hibbs.

https://f1000.com/prime/734202313?bd=1

• Sieghart et al., 2022, Pharmacol. Rev. 74, 238-270 (1 citation)

This is a comprehensive and up-to-date review on GABAA receptors containing a6 subunits. It discusses the different types of these receptors, their subunit composition and electrophysiological and pharmacological properties. It covers their regional, cellular, and subcellular distribution not only in the central nervous system but also in the peripheral nervous system and peripheral tissues, and for the first time summarizes their importance for the functions of the cerebellum and the nervous system.

The cerebellum is not only involved in motor control but also in cognitive, emotional and social behaviors. a6-containing GABAA receptors are of crucial importance for the function of the cerebellum. The synaptically located a6-containing GABAA receptors enhance the precision of inputs required for cerebellar timing of motor activity. They thus significantly contribute to adequate responses to our environment and to cognitive processing. Extrasynaptically located a6-containing GABAA receptors regulate the amount of information entering the cerebellum, and thus allow to concentrate on important tasks.

Evidence from animal models mimicking neuropsychiatric phenotypes suggest that compounds selectively modulating cerebellar a6-containing GABAA receptors can alleviate essential tremor and motor disturbances in Angelman and Down syndrome, as well as impaired prepulse inhibition in multiple neuropsychiatric disorders. a6-containing GABAA receptors located in trigeminal ganglia reduce migraine and trigeminal-related pain.

Genetic studies in humans as well as animal studies suggest an association of the human GABAA receptor a6 subunit gene with stress-associated disorders. Neuroimaging and post-mortem studies in humans further support an involvement of a6-containing GABAA receptors in a large variety of neuropsychiatric disorders, pointing to a broad therapeutic potential of drugs modulating a6-containing GABAA receptors. Depending on their actual effects in human neuropsychiatric disorders, these drugs could cause a major breakthrough in psychopharmacology.