Scholze, P. | bioRxiv Science

bioRxiv2018

Potential combined pro-cognitive, anxiolytic and antidepressant properties of novel GABAA receptor positive modulators with preferential efficacy at the α5-subunit

Altered {gamma}-aminobutyric acid (GABA) function is consistently reported in psychiatric disorders, normal aging and neurodegenerative disorders, and reduced function of somatostatin - expressing GABA interneurons is associated with both mood and cognitive symptoms. Somatostatin-neurons signal in part through 5-subunit containing GABAA receptors (5-GABAA-Rs) which are localized in brain regions implicated in emotion and cognition. We hypothesize that enhancing 5-GABAA-R activity has therapeutic potential for both mood and cognitive symptoms in stress-based and aging rodent models.\n\nWe synthesized four novel imidazobenzodiazepine (IBZD) amide ligands, tested them for positive allosteric modulation at 5-GABAA-R (5-PAM), pharmacokinetic properties, and for anxiolytic and antidepressant activities in adult mice. Pro-cognitive activity was tested in adult mice submitted to chronic stress and in old mice. Diazepam (DZP), with broad PAM activity at GABAA-Rs, was used as a control.\n\nThree novel IBZD amide ligands (GL-II-73, GL-II-74 and GL-II-75) demonstrated adequate brain penetration, affinity and 5-PAM activity, and metabolic stability for in vivo studies. GL-II-73/74/75 showed significant anxiolytic and antidepressant efficacies in adult mice. GL-II-73 and GL-II-75 significantly reversed cognitive deficits induced by stress or occurring throughout normal aging. This activity was maintained after sub-chronic administration for GL-II-73. In contrast DZP displayed anxiolytic but no antidepressant or pro-cognitive activities.\n\nWe demonstrate for the first time the potential for combined anxiolytic, antidepressant and pro-cognitive therapeutic, mediated by newly designed IBDZ amide ligands with efficacy at 5-GABAA-Rs. These results suggest a novel therapeutic approach targeting both mood and cognitive symptoms in depression and/or aging.

pharmacology and toxicology↗

bioRxiv2018

Kinase activity simultaneously determines the constitutive and the orthosteric gating in α4β1/3δ GABAA receptors in hippocampal granule cells

A subset of the GABAA receptors expressed in recombinant systems and neurons is known to exhibit both constitutive- and agonist-induced gating. Two such receptors are the {delta}-subunit containing GABAA receptors 4{beta}1{delta} and 4{beta}3{delta}, which are expressed in adult rodent hippocampal dentate gyrus granule cells (DGGCs). Here we show that the GABAA receptor mediated tonic current recorded in the presence of tetrodotoxin in adult rodent DGGCs is almost exclusively mediated by constitutively active {delta}-subunit containing GABAA receptors and that the constitutive current is absent in recordings at 24 {degrees}C or in recordings at 34 {degrees}C including an intracellular inhibitor of protein kinase C. These factors simultaneously govern the efficacy of an orthosteric agonist at 4{beta}1/3{delta} receptors, Thio-THIP, in a reciprocal manner. In the absence of constitutive receptor activity, the efficacy of Thio-THIP was increased approximately four-fold relative to recording conditions that favors constitutive activity. Further, only under conditions of an absent constitutive current, the classified neutral antagonist gabazine (GBZ) alone, induced a tonic current in DGGCs (EC50 2.1 M). This effect of GBZ was not seen in recording conditions of high constitutive activity, was inhibited by picrotoxin (PTX), potentiated by DS2, completely absent in {delta}-/- mice and reduced in {beta}1-/- mice, but could not be replicated in human 4{beta}1/3{delta} receptors expressed recombinantly in HEK cells. We hypothesize that specific intracellular components in neurons interact with receptors to determine constitutive gating and receptor responsiveness to orthosteric ligands.\n\nSignificance statementThe presented data highlight how recording conditions for whole cell patch clamp analysis of 4{beta}1/3{delta} GABAA receptors can mask important pharmacological effects. Specifically, orthosteric agonists appear with reduced efficacy, and other ligands, here exemplified with the well-known antagonist GBZ, are misinterpreted as being inactive/neutral, although they could have effect in constitutively silent receptors. Unmasking of potential hidden effects are easily done using recording conditions of reduced kinase activity in a relevant neuronal context. It follows that in pathologies with changes in phosphorylation level of {delta}-subunit containing GABAA receptors, the efficacy of an agonist of these receptors, measured by whole-cell recordings in vitro, will not match the efficacy of the same agonist in an unperturbed neuron in vivo.

neuroscience↗