Gintonin, an exogenous ginseng-derived LPA receptor ligand, promotes corneal wound healing

Ginseng gintonin is an exogenous ligand of lysophosphatidic acid (LPA) receptors. Accumulating evidence shows LPA helps in rapid recovery of corneal damage. The aim of this study was to evaluate the therapeutic efficacy of gintonin in a rabbit model of corneal damage. We investigated the signal transduction pathway of gintonin in human corneal epithelium (HCE) cells to elucidate the underlying molecular mechanism. We next evaluated the therapeutic effects of gintonin, using a rabbit model of corneal damage, by undertaking histochemical analysis. Treatment of gintonin to HCE cells induced transient increases of [Ca²⁺](i) in concentration-dependent and reversible manners. Gintonin-mediated mobilization of [Ca²⁺](i) was attenuated by LPA1/3 receptor antagonist Ki16425, phospholipase C inhibitor U73122, inositol 1,4,5-triphosphate receptor antagonist 2-APB, and intracellular Ca²⁺ chelator BAPTA-AM. Gintonin facilitated in vitro wound healing in a concentration-dependent manner. When applied as an eye-drop to rabbits with corneal damage, gintonin rapidly promoted recovery. Histochemical analysis showed gintonin decreased corneal apoptosis and increased corneal cell proliferation. We demonstrated that LPA receptor activation by gintonin is linked to in vitro and in vivo therapeutic effects against corneal damage. Gintonin can be applied as a clinical agent for the rapid healing of corneal damage.

Resveratrol Inhibits GABAC rho Receptor-Mediated Ion Currents Expressed in Xenopus Oocytes

Resveratrol is a phytoalexin found in grapes, red wine, and berries. Resveratrol has been known to have many beneficial health effects, such as anti-cancer, neuroprotective, anti-inflammatory, and life-prolonging effects. However, relatively little is known about the effects of resveratrol on the regulation of ligand-gated ion channels. We have previously reported that resveratrol regulates subsets of homomeric ligand-gated ion channels such as those of 5-HT3A receptors. The gamma-aminobutyric acidC (GABAC) receptor is mainly expressed in retinal bipolar cells and plays an important role in visual processing. In the present study, we examined the effects of resveratrol on the channel activity of homomeric GABAC receptor expressed in Xenopus oocytes injected with cRNA encoding human GABAC rho subunits. Our data show that the application of GABA elicits an inward peak current (IGABA) in oocytes that express the GABAC receptor. Resveratrol treatment had no effect on oocytes injected with H2O or with GABAC receptor cRNA. Co-treatment with resveratrol and GABA inhibited IGABA in oocytes with GABAC receptors. The inhibition of IGABA by resveratrol was in a reversible and concentration-dependent manner. The IC50 of resveratrol was 28.9+/-2.8 microM in oocytes expressing GABAC receptor. The inhibition of IGABA by resveratrol was in voltage-independent and non-competitive manner. These results indicate that resveratrol might regulate GABAC receptor expression and that this regulation might be one of the pharmacological actions of resveratrol on the nervous system.

Inhibitory Effects of Ginsenoside Metabolites, Compound K and Protopanaxatriol, on GABAC Receptor-Mediated Ion Currents

Ginsenosides, one of the active ingredients of Panax ginseng, show various pharmacological and physiological effects, and they are converted into compound K (CK) or protopanaxatriol (M4) by intestinal microorganisms. CK is a metabolite derived from protopanaxadiol (PD) ginsenosides, whereas M4 is a metabolite derived from protopanaxatriol (PT) ginsenosides. The gamma-aminobutyric acid receptorC (GABAC) is primarily expressed in retinal bipolar cells and several regions of the brain. However, little is known of the effects of ginsenoside metabolites on GABAC receptor channel activity. In the present study, we examined the effects of CK and M4 on the activity of human recombinant GABAC receptor (rho1) channels expressed in Xenopus oocytes by using a 2-electrode voltage clamp technique. In oocytes expressing GABAC receptor cRNA, we found that CK or M4 alone had no effect in oocytes. However, co-application of either CK or M4 with GABA inhibited the GABA-induced inward peak current (IGABA). Interestingly, pre-application of M4 inhibited IGABA more potently than CK in a dose-dependent and reversible manner. The half-inhibitory concentration (IC50) values of CK and M4 were 52.1+/-2.3 and 45.7+/-3.9 microM, respectively. Inhibition of IGABA by CK and M4 was voltage-independent and non-competitive. This study implies that ginsenoside metabolites may regulate GABAC receptor channel activity in the brain, including in the eyes.

Effects of Protopanaxatriol-Ginsenoside Metabolites on Rat N-Methyl-D-Aspartic Acid Receptor-Mediated Ion Currents

Ginsenosides are low molecular weight glycosides found in ginseng that exhibit neuroprotective effects through inhibition of N-methyl-D-aspartic acid (NMDA) receptor channel activity. Ginsenosides, like other natural compounds, are metabolized by gastric juices and intestinal microorganisms to produce ginsenoside metabolites. However, little is known about how ginsenoside metabolites regulate NMDA receptor channel activity. In the present study, we investigated the effects of ginsenoside metabolites, such as compound K (CK), protopanaxadiol (PPD), and protopanaxatriol (PPT), on oocytes that heterologously express the rat NMDA receptor. NMDA receptor-mediated ion current (INMDA) was measured using the 2-electrode voltage clamp technique. In oocytes injected with cRNAs encoding NMDA receptor subunits, PPT, but not CK or PPD, reversibly inhibited INMDA in a concentration-dependent manner. The IC50 for PPT on INMDA was 48.1+/-4.6 microM, was non-competitive with NMDA, and was independent of the membrane holding potential. These results demonstrate the possibility that PPT interacts with the NMDA receptor, although not at the NMDA binding site, and that the inhibitory effects of PPT on INMDA could be related to ginseng-mediated neuroprotection.

Inhibitory Effects of Quercetin on Muscle-type of Nicotinic Acetylcholine Receptor-Mediated Ion Currents Expressed in Xenopus Oocytes

The flavonoid quercetin is a low molecular weight compound generally found in apple, gingko, tomato, onion and other red-colored fruits and vegetables. Like other flavonoids, quercetin has diverse pharmacological actions. However, relatively little is known about the influence of quercetin effects in the regulation of ligand-gated ion channels. Previously, we reported that quercetin regulates subsets of nicotinic acetylcholine receptors such as alpha3beta4, alpha7 and alpha9alpha10. Presently, we investigated the effects of quercetin on muscle-type of nicotinic acetylcholine receptor channel activity expressed in Xenopus oocytes after injection of cRNA encoding human fetal or adult muscle-type of nicotinic acetylcholine receptor subunits. Acetylcholine treatment elicited an inward peak current (IACh) in oocytes expressing both muscle-type of nicotinic acetylcholine receptors and co-treatment of quercetin with acetylcholine inhibited IACh. Pre-treatment of quercetin further inhibited IACh in oocytes expressing adult and fetal muscle-type nicotinic acetylcholine receptors. The inhibition of IACh by quercetin was reversible and concentration-dependent. The IC50 of quercetin was 18.9+/-1.2 microM in oocytes expressing adult muscle-type nicotinic acetylcholine receptor. The inhibition of IACh by quercetin was voltage-independent and non-competitive. These results indicate that quercetin might regulate human muscle-type nicotinic acetylcholine receptor channel activity and that quercetin-mediated regulation of muscle-type nicotinic acetylcholine receptor might be coupled to regulation of neuromuscular junction activity.

Quercetin Inhibits alpha3beta4 Nicotinic Acetylcholine Receptor-Mediated Ion Currents Expressed in Xenopus Oocytes

Quercetin mainly exists in the skin of colored fruits and vegetables as one of flavonoids. Recent studies show that quercetin, like other flavonoids, has diverse pharmacological actions. However, relatively little is known about quercetin effects in the regulations of ligand-gated ion channels. In the previous reports, we have shown that quercetin regulates subsets of homomeric ligand-gated ion channels such as glycine, 5-HT3A and alpha7 nicotinic acetylcholine receptors. In the present study, we examined quercetin effects on heteromeric neuronal alpha3beta4 nicotinic acetylcholine receptor channel activity expressed in Xenopus oocytes after injection of cRNA encoding bovine neuronal alpha3 and beta4 subunits. Treatment with acetylcholine elicited an inward peak current (IACh) in oocytes expressing alpha3beta4 nicotinic acetylcholine receptor. Co-treatment with quercetin and acetylcholine inhibited IACh in oocytes expressing alpha3beta4 nicotinic acetylcholine receptors. The inhibition of IACh by quercetin was reversible and concentration-dependent. The half-inhibitory concentration (IC50) of quercetin was 14.9+/-0.8 microM in oocytes expressing alpha3beta4 nicotinic acetylcholine receptor. The inhibition of IACh by quercetin was voltage-independent and non-competitive. These results indicate that quercetin might regulate alpha3beta4 nicotinic acetylcholine receptor and this regulation might be one of the pharmacological actions of quercetin in nervous systems.