Partial Agonism of Taurine at Gamma-Containing Native and Recombinant GABAA Receptors

Abstract

Taurine is a semi-essential sulfonic acid found at high concentrations in plasma and mammalian tissues which regulates osmolarity, ion channel activity and glucose homeostasis. The structural requirements of GABAA-receptors (GABAAR) gated by taurine are not yet known. We determined taurine potency and efficacy relative to GABA at different types of recombinant GABAAR occurring in central histaminergic neurons of the mouse hypothalamic tuberomamillary nucleus (TMN) which controls arousal. At binary α1/2β1/3 receptors taurine was as efficient as GABA, whereas incorporation of the γ1/2 subunit reduced taurine efficacy to 60–90% of GABA. The mutation γ2F77I, which abolishes zolpidem potentiation, significantly reduced taurine efficacy at recombinant and native receptors compared to the wild type controls. As taurine was a full- or super- agonist at recombinant αxβ1δ-GABAAR, we generated a chimeric γ2 subunit carrying the δ subunit motif around F77 (MTVFLH). At α1/2β1γ2(MTVFLH) receptors taurine became a super-agonist, similar to δ-containing ternary receptors, but remained a partial agonist at β3-containing receptors. In conclusion, using site-directed mutagenesis we found structural determinants of taurine’s partial agonism at γ-containing GABAA receptors. Our study sheds new light on the β1 subunit conferring the widest range of taurine-efficacies modifying GABAAR function under (patho)physiological conditions.

Benzodiazepine‐site pharmacology on GABAA receptors in histaminergic neurons

“…GABA A R lacking γ‐subunits are blocked by ZnCl 2 10 μM, whereas γ‐subunit‐containing receptors are weakly or not affected by this concentration (Draguhn et al ., ; Herb et al ., ). In contrast to the rat TMN neurons which are all zinc resistant (Sergeeva et al ., ), we found that, in about 30% of the mouse TMN neurons identified by the expression of HDC, GABA‐responses are about halved by ZnCl 2 10 μM (Kletke et al ., ). In the present study, pyrazolopyridine CGS 20625, a two to six times more potent positive modulator of α1β2 than of α1β2γ receptors (Khom et al ., ), confirmed γ‐subunit‐deficiency of zinc‐sensitive TMN neurons [CGS 20625's EC 50 = 1.1 ± 0.1 μM ( n = 5, 28% of total cell number], which is significantly different ( P = 0.0055) from the zinc‐resistant cells (EC 50 = 3.4 ± 0.4 μM, n = 13, Supporting Information http://onlinelibrary.wiley.com/doi/10.1111/bph.12280/suppinfo).…”

“…Alternatively, receptors containing the γ1‐subunit (Puia et al ., ; Wafford et al ., ) or γ3‐subunit (Herb et al ., ) could mediate the response to zolpidem. No potentiation of GABA A R by zolpidem (1 or 10 μM) is observed in oocytes transfected with α1/2, β2/3, γ2 F77I ‐subunits (Buhr et al ., ; Ramerstorfer et al ., ; Kletke et al ., ), indicating that the ‘anaesthetics‐binding site’ present on β2 and β3 subunits is not involved. The modulatory potency and efficacy of diazepam and chlordiazepoxide was indifferent between WT and KI neurons in our study, whereas at recombinant α1β2γ2 F77I receptors the efficacy of diazepam is reduced (Ramerstorfer et al ., ).…”

“…Receptors composed of α1, α2 or α5, β1 and γ1‐subunits are positively modulated by diazepam (Puia et al ., ) although to a smaller extent than the corresponding γ2‐containing receptors. We did not detect γ3‐subunit expression in individual TMN neurons of the mouse in this and in our previous studies (Sergeeva et al ., ; Kletke et al ., ), whereas the whole TMN region used as positive control contained γ3‐subunit transcripts. This subunit confers zinc‐resistance to the recombinant GABA A receptors and benzodiazepine‐site ligands modulate them, although with very low potencies and efficacies (Herb et al ., ).…”

“…The α‐type subunits influence this modulation and it was reported that association of the γ1‐subunit with the α1‐ and β‐subunits results in a greater zolpidem modulation of GABA‐evoked currents compared to the α2‐containing receptors (Puia et al ., ). Recombinant receptors composed of α2, β1 and γ1 subunits are not modulated by zolpidem [see Supporting Information http://onlinelibrary.wiley.com/doi/10.1111/bph.12280/suppinfo in Kletke et al ., ()]. The α1‐subunit thus plays an important role for the zolpidem modulation of GABA‐responses in mutant mice.…”

“…Hyperammonemia, the major pathogenic factor of HE, triggers brain taurine release, which is neuroprotective through GABA A and glycine receptors. Our finding that the mutation γ2 F77I impairs GABA A R gating by taurine (Kletke et al ., ) will complicate interpretation of data obtained from this model. The changed benzodiazepine‐site pharmacology in histaminergic neurons from γ2 F77I mice highlights a yet unexplored structural complexity of GABA A R, whose modification under pathological conditions is far from being understood (Sergeeva, ).…”