Synergistic Inhibition of Glycinergic Transmission In Vitro and In Vivo by Flavonoids and Strychnine

Abstract

The inhibitory glycine receptor (GlyR) is a key mediator of synaptic signaling in spinal cord, brain stem, and higher central nervous system regions. The flavonoids quercetin and genistein have been identified previously as promising GlyR antagonists in vitro, but their detailed mechanism of action was not known. Here, inhibition of recombinant human α1 GlyRs in HEK 293 cells by genistein, quercetin, and strychnine was studied using whole-cell recording techniques. The interaction of several inhibitors applied alone or in combination was analyzed using a minimum mechanism of receptor activation and inhibition. Receptor inhibition in vivo was studied in a mouse model of strychnine toxicity. Genistein, quercetin, and strychnine were noncompetitive GlyR inhibitors. The inhibitory potency of one flavonoid (either genistein or quercetin) was not affected by simultaneous application of the other, suggesting that both flavonoids target the same site on the receptor. In combination with strychnine, flavonoid inhibition was augmented, indicating that strychnine binds to a position on the receptor physically distant from the flavonoid site. Potentiation of strychnine inhibition by flavonoids was also observed in vivo, where harmless doses of flavonoids enhanced strychnine toxicity in mice. Thus, in vitro and in vivo studies demonstrated a true synergism between flavonoids and strychnine for GlyR inhibition. The mechanism-based approach used here allows a rapid analysis of the effects of single drugs versus drug combinations.

Amelioration of Diabetes and Painful Diabetic Neuropathy by Punica granatum L. Extract and Its Spray Dried Biopolymeric Dispersions

“…The environment consisted of standard mouse cages with a 12 h light/dark cycle. The temperature was 22 ± 1°C; animals had open access to water and standard laboratory pellets (20% proteins, 5% fats, and 1% multivitamins) [26, 28]. The mice were kept in those conditions for a seven-day period of adaptation prior to the beginning of the experiment.…”

Flavonoid nutraceuticals and ionotropic receptors for the inhibitory neurotransmitter GABA

“…Dietary flavonoids are known to induce cytoprotective proteins (Leonardo and Dore, 2011). They interact with many neurotransmitter systems including adenosine (Alexander, 2006), dopamine (Kita et al, 2014), GABA (Hanrahan et al, 2011) and glycine (Raafat et al, 2010) that may be involved in neuroprotection.…”

“…It is not easy to study as it involves the doseedependant interactions between three ligands and thus may be difficult to observe. Synergistic interactions have been described between other flavonoids and GABA receptors , and between flavonoids, strychnine and glycine receptors (Raafat et al, 2010). Thus complex tertiary interactions between flavonoids and other substances may be a subtle feature of cys-loop ligand gated ion channels.…”

Drug Synergy – Mechanisms and Methods of Analysis

“…T h i s m a y b e a n i m p o r t a n t r e s u l t f o r d r u g development, and is obtained without need of structural data. (Note: strictly speaking, the result only tells whether binding of two inhibitors is mutually exclusive or nonexclusive) The method has originally been presented for ligand-gated ion channels by Karpen and Hess (Karpen, Aoshima et al 1982;Karpen and Hess 1986), and subsequently been used for the study of action of multiple inhibitors on ion channels (Karpen, Aoshima et al 1982;Karpen and Hess 1986;Breitinger, Geetha et al 2001;Raafat, Breitinger et al 2010). The basic mechanism presented here is by far not sufficient to describe multimeric enzymes, enzynmes requiring cofactors, and various modes of inhibition.…”