Michel Lesne

Benzodiazepine receptors are involved in tabernanthine-induced tremor: in vitro and in vivo evidence

Tabernanthine, an indol alkaloid, is structurally related to carbolines (harmane, harmaline) which, in vitro, displace specific flunitrazepam binding to brain benzodiazepine receptors. In vivo, both tabernanthine and carbolines cause a fine general tremor, suggesting that a possible interaction with benzodiazepine receptors could be involved in the activity of tabernanthine. This hypothesis was validated by the in vitro and in vivo antagonism of benzodiazepine by tabernanthine. In vitro, tabernanthine inhibited specific flunitrazepam binding in a competitive manner with an affinity (IC50 150 microM) in the same range as harmane. Tabernanthine appeared as a benzodiazepine receptor inverse agonist in a discriminant in vitro binding assay. In vivo, the time course of tremorigenic activity was related to the tabernanthine concentration in brain (half-life = 2 h). Moreover, tabernanthine-induced tremor was inhibited reversibly by flunitrazepam or by Ro-15 1788 (an antagonist of benzodiazepine-receptors). These results suggest that part of the action of tabernanthine may be mediated by an interaction at the benzodiazepine receptor level.

Digoxin Acute Intoxication: Evaluation of the Efficiency of Charcoal Hemoperfusion

Since there is no widely used method of reducing the severity of massive digoxin intoxication, the capacity of hemoperfusion with coated, activated charcoal to remove digoxin was evaluated in a case of suicidal digoxin ingestion (25 mg). Seven hours after ingestion the digoxin plasma level was equal to 8.9 ng/ml. This was decreased to 4.5 ng/ml after 6 hr hemoperfusion. The amount of digoxin adsorbed by the column represents 4.8% of the absorbed dose. At a blood flow rate of 170 ml/min, the mean digoxin clearance by hemoperfusion was 44.5 +/- 26.9 ml/min. From these results we conclude that charcoal hemoperfusion in acute digoxin intoxication is of little value.

Hepatic-clearance of Gitoxin - Pharmacokinetic Study On Rabbit Isolated Liver - Influence of Protein-binding and Comparison With Digoxin

SummaryHepatic clearance of gitoxin has been studied in the rabbit and compared with that of digoxin using an isolated perfused liver technique. During 1.5 hour perfussions with a modified Krebs-Henseleit solution, gitoxin perfusate levels decreased bi-exponentially; the distribution and elimination half-lives were estimated to be 0.14 and 1.25 hour, Vd area to be 95.5 ml.g−1 and intrinsic metabolic clearance to be 1.98 ml.min−1.g−1.During 1.5 hour perfussions with modified Krebs-Henseleit solution containing 2.7% bovine serum albumin, gitoxin perfusate levels decreased monoexponentially. This is probably due to protein binding which moderates hepatic uptake so that distribution is not yet complete after 1.5 hour and it is therefore impossible to discriminate the two phases. This was confirmed by 5 hour perfusion experiments with an emulsion of perfluorocarbon in the modified Krebs-Henseleit solution also containing 2.7% bovine serum albumin, during which gitoxin levels decreased bioexponentially. Distribution and elimination half-lives have been estimated to be 0.31 and 5.54 hours, Vd area to be 139 ml.g−1 and intrinsic metabolic clearance to be 1.36 ml. min−1.g−1. Gitoxin has been compared in these experimental conditions with digoxin, one of the most often used cardiotonic’s. Distribution and elimination half-lives of digoxin were estimated to be 0.34 and 4.52 hours, Vd area to be 46.5 ml.g−1 and intrinsic metabolic clearance to be 0.17 ml.min−1.g−1. Other pharmacokinetic parameters (α, β, V1, V2…) also have been calculated for these three types of perfusion experiments.

The binding of gitoxin to human plasma proteins.

SummaryThe binding of gitoxin, digitoxin and digoxin to human plasma proteins was measured by ultracentrifugation and equilibrium dialysis. At concentrations in the range of therapeutic plasma levels, protein binding amounted, respectively, to 85, 92 and 20%, the last two values being consistent with data reported in the literature. The affinity of purified human albumin was not significantly different for the three cardiac glycosides tested. No other protein than albumin was found to bind gitoxin in human plasma.

A Further Study of the Pharmacokinetics of Gitoxin in Rabbit Isolated Liver - Clearance of 3h-gitoxin

SummaryHepatic clearance of 3H-gitoxin was studied in the rabbit using an isolated perfused liver technique with an emulsion of a perfluorocarbon. The liposoluble material in the perfusion medium was extracted with dichloromethane, and gitoxin was assayed in the extract by high performance liquid chromatography.Pharmacokinetic parameters were estimated for the liposoluble (dichloromethane soluble) material in the water phase obtained by centrifugation of the emulsion, for the liposoluble material and unchanged gitoxin in the total emulsion.Distribution and elimination half-lives of the liposoluble fraction in the water phase, were estimated to be 0.47 and 4.80 hours respectively, Vd to be 148 ml.g−1 and intrinsic clearance to be 1.16 ml.min−1 .g−1; these parameters were compared with those of a previous study with unlabelled gitoxin.Distribution and elimination half-lives of the liposoluble compounds in the emulsion were estimated to be 0.48 a n d 4.62 hours, Vd to be 47 ml.g−1 and intrinsic clearance to be 1.07 ml. min−1. g−1; these data were compared with those of the liposoluble compounds in the water phase.Distribution and elimination half-lives of unchanged gitoxin in the emulsion were estimated to be 0.22 and 0.70 hour, Vd to be 59 ml.g−1 and intrinsic clearance to be 11.4 ml.min−1 .g−1; these data were compared with those of the liposoluble compounds in the emulsion.The subcellular distribution of gitoxin and its metabolites in the liver indicated that 79% of t h e radioactivity was found in the soluble fraction, no significant binding occurring in the mitochondrial and microsomal fractions.

Isolation and Mass-spectrometric Identification of Gitoxin Metabolites Excreted in Bile

SummaryThe TLC isolation as well as the mass spectrometric identification of gitoxin metabolites excreted in bile are described.