Binding of STW 5 (Iberogast) and its components to intestinal 5-HT, muscarinic M3, and opioid receptors.

Clinical studies with the fixed herbal combination product STW 5 (Iberogast) have indicated an efficacy comparable to metoclopramide (5-HT(3) antagonist) and cisapride (5-HT(4) agonist) in functional gastro-intestinal diseases like functional dyspepsia (FD) and irritable bowel syndrome (IBS). Since serotonin (5-HT(3) and 5-HT(4)) and muscarinic M(3) receptors are known to play a central role in the etiology of FD and IBS, the extracts contained in STW 5 and several of their phytochemical components were studied in vitro for binding affinities to these receptors of the intestine. STW 5 inhibited the binding of (3)H-GR113808 and (3)H-4-DAMP to 5-HT(4) and M(3) receptors, respectively, about 10 times more potently than the binding of (3)H-GR65630 to 5-HT(3) receptors. IC(50) values for STW 5 did correspond to extract dilutions of 1:1000 (M(3) binding) and 1:2000 (5-HT(4) binding). In addition, STW 5 also potently inhibited the binding to opioid receptors with an IC(50) value of 1:2000. Of the nine herbal extracts contained in STW 5, the fresh plant extract of bitter candy tuft (Iberis amara) selectively inhibited binding to M(3) receptors, while ethanolic extracts of celandine herb and chamomile flower were selective to 5-HT(4), and liquorice root to 5-HT(3) receptors. Binding affinities to human recombinant 5-HT(3), 5-HT(4) and M(3) receptors were qualitatively similar to those of the corresponding intestinal receptors. The benzylisoquinoline alkaloid berberine had significant inhibitory action on 5-HT(4) and M(3) binding, showing IC(50) values of 40 ng/ml (100 nM) and 200 ng/ml (500 nM), respectively, but is present in the extract of celandine herb only in traces, so that also for the celandine extract a cooperative effect of several phytochemical constituents can be assumed. These in vitro data indicate that 5-HT(4) (to a lesser degree 5-HT(3)), muscarinic M(3), and opioid receptors represent target sites for the treatment of FD and IBS with STW 5 (Iberogast).

Neurochemical studies with St. John's wort in vitro.

The effect of extracts and constituents of St. John's wort, Hypericum perforatum, at various CNS receptors were studied by radioligand binding techniques in order to determine a profile of pharmacological activity in vitro. Binding inhibition was examined for the G-protein coupled opioid, serotonin (5-HT), histamine, neurokinin and corticotropin releasing factor (CRF) receptors, for the steroid estrogen-alpha receptor and for the ligand-gated ionchannel GABA(A) receptor. Hypericin showed the most potent binding inhibiton of all tested constituents to human CRF1 receptor with an IC50 value of 300 nM. Preliminary GTPgamma35S binding studies to CRF1 coupled G-protein indicated an antagonistic action for hypericin. The acylphloroglucinole hyperforin failed to inhibit 125I-astressin binding to hCRF, receptor up to 10 microM. Hyperforin inhibited binding to opioid and serotonin (5-HT) receptors at IC50 values between 0.4 and 3 microM, while hypericin and pseudohypericin inhibited with weaker potency. The biflavonoid I3,II8-biapigenin inhibited 3H-estradiol binding to the estrogen-alpha receptor with an IC50 value of 1 microM. The inhibition of 3H-muscimol binding to the GABA(A) receptor is likely to be exclusively due to GABA present in the extract. We therefore hypothesize that additive or synergistic actions of several ditsinct compounds may be responsible for the beneficial antidepressant effect of St. John's wort.

Interaction of various Piper methysticum cultivars with CNS receptors in vitro.

Methanolic leaf and root extracts of the Hawaiian kava (Piper methysticum Forst.) cultivars, Mahakea, Nene, Purple Moi and PNG, were tested on binding affinities to CNS receptors including GABAA (GABA and benzodiazepine binding site), dopamine D2, opioid (mu and delta), serotonin (5-HT6 and 5-HT7) and histamine (H1 and H2). HPLC analysis was carried out in order to determine the amount of the main kavalactones kavain, 7,8-dihydrokavain, methysticin, 7,8-dihydromethysticin, yangonin and 5,6-demethoxyyangonin. The most potent binding inhibition was observed for leaf extracts to GABAA receptors (GABA binding site) with IC50 values of approximately 3 micrograms/ml, whereas root extracts were less active with IC50 values ranging from 5 micrograms/ml (Nene) to 87 micrograms/ml (Mahakea). Since the leaf extracts generally contained lower amounts of the kavalactones than the root extracts, there might exist additional substances responsible for these activities. Leaf extracts also inhibited binding to dopamine D2, opioid (mu and delta) and histamine (H1 and H2) receptors more potently than the corresponding root extracts with IC50 values ranging from 1 to 100 micrograms/ml vs. > or = 100 micrograms/l, respectively. Significant differences in the potential of binding inhibition were also observed between cultivars. Binding to serotonin (5-HT6 and 5-HT7) and benzodiazepine receptors was only weakly inhibited by both root and leaf extracts of all four cultivars. In conclusion, our investigation indicates that the GABAA, dopamine D2, opioid (mu and delta) and histamine (H1 and H2) receptors might be involved in the pharmacological action of kava extracts. Since the cultivars contained similar amounts of kavalactones, while their pharmacological activities differed markedly, other constituents may play a role in the observed activities. Additionally, leaves generally exhibited more potent binding inhibition than roots, therefore leaf of P. methysticum might be an interesting subject for further pharmacological studies.

Bisanthraquinone glycosides of Hypericum perforatum with binding inhibition to CRH-1 receptors.

Four new bisanthraquinone glycosides, S-(+)-skyrin-6-O-beta-glucopyranoside (1), R-(-)-skyrin-6-O-beta-glucopyranoside (2), S-(+)-skyrin-6-O-beta-xylopyranoside (3) and S-(+)-skyrin-6-O-beta-alpha-arabinofuranoside (4), have been isolated from an ethanol-water (1:1, v/v) dry extract of the aerial parts of Hypericum perforatum L. The structures were elucidated by spectroscopic methods, mainly NMR and mass spectrometry. Circular dichroism was used to determine their axial stereochemistry revealing 1 and 2 to be atropisomers. 1 and 2 inhibited [125I]sauvagine binding to corticotropin releasing hormone (CRH-1) receptors.

Extracts and constituents of Hypericum perforatum inhibit the binding of various ligands to recombinant receptors expressed with the Semliki Forest virus system.

Extracts, fractions and constituents of Hypericum perforatum were studied for in vitro receptor binding with various ligands to recombinant CNS receptors expressed with the Semliki Forest virus expression system. For this purpose we have prepared membranes of CHO cells with high density of several opioid, serotonin, estrogen, histamine, GABAA, neurokinin and metabotropic glutamate receptors, respectively. A lipophilic Hypericum fraction revealed relatively potent inhibition to the binding of the mu-, delta- and kappa-opioid and the 5-HT6 and 5-HT7 receptors. Moreover, Hypericum constituents such as the naphthodianthrones, hypericin and pseudohypericin, and the phloroglucinole hyperforin inhibited both binding to the opioid and serotonin receptors in the lower micromolar range. Estrogen binding was 50% inhibited by the biflavonoid I3,II8-biapigenin at micromolar concentration. The lipophilic Hypericum fraction provided a less potent inhibition of the neurokinin-1 receptor binding compared to the opioid and serotonin receptors. A total ethanolic Hypericum extract potently inhibited GABAA binding at approximately 3 micrograms/ml. This inhibition is however not specific to Hypericum, since extracts of plants like Valeriana officinalis and Passiflora incarnata showed similar inhibitions. Binding to neither histamine nor metabotropic glutamate receptors was affected by Hypericum extracts. These results support the hypothesis that several active constituents of Hypericum might in a synergistic way contribute to its antidepressant effect in the central nervous system.

Hypericum perforatum inhibits the binding of mu- and kappa-opioid receptor expressed with the Semliki Forest virus system.

The effects of Hypericum perforatum extracts on in vitro [3H]naloxone binding to the human mu- and rat kappa-opioid receptors were studied in chinese hamster ovary (CHO) cells using the Semliki Forest virus (SFV) expression system. Binding of [3H]naloxone to the mu- and kappa-opioid receptor was inhibited in the presence of Hypericum extracts showing IC50 values of approximately 25 and 90 micrograms/ml, respectively. In contrast, extracts of Valeriana officinalis did not inhibit binding to the mu-opioid receptor. Also, single constituents of H. perforatum like the flavonoids quercetin and kaempferol and the glycosilated flavonoid quercitrin did not inhibit [3H]naloxone binding to the mu-opioid receptor up to a concentration of 10 microM. The present in vitro data may suggest a new possible mechanism for the anti-depressant effect of H. perforatum.