Effects of monoamine oxidase inhibitor and cytochrome P450 2D6 status on 5-methoxy-N,N-dimethyltryptamine metabolism and pharmacokinetics.

5-Methoxy-N,N-dimethyltryptamine (5-MeO-DMT) is a natural psychoactive indolealkylamine drug that has been used for recreational purpose. Our previous study revealed that polymorphic cytochrome P450 2D6 (CYP2D6) catalyzed 5-MeO-DMT O-demethylation to produce active metabolite bufotenine, while 5-MeO-DMT is mainly inactivated through deamination pathway mediated by monoamine oxidase (MAO). This study, therefore, aimed to investigate the impact of CYP2D6 genotype/phenotype status and MAO inhibitor (MAOI) on 5-MeO-DMT metabolism and pharmacokinetics. Enzyme kinetic studies using recombinant CYP2D6 allelic isozymes showed that CYP2D6.2 and CYP2D6.10 exhibited 2.6- and 40-fold lower catalytic efficiency (V(max)/K(m)), respectively, in producing bufotenine from 5-MeO-DMT, compared with wild-type CYP2D6.1. When co-incubated with MAOI pargyline, 5-MeO-DMT O-demethylation in 10 human liver microsomes showed significantly strong correlation with bufuralol 1'-hydroxylase activities (R(2)=0.98; P<0.0001) and CYP2D6 contents (R(2)=0.77; P=0.0007), whereas no appreciable correlations with enzymatic activities of other P450 enzymes. Furthermore, concurrent MAOI harmaline sharply reduced 5-MeO-DMT depletion and increased bufotenine formation in human CYP2D6 extensive metabolizer hepatocytes. In vivo studies in wild-type and CYP2D6-humanized (Tg-CYP2D6) mouse models showed that Tg-CYP2D6 mice receiving the same dose of 5-MeO-DMT (20mg/kg, i.p.) had 60% higher systemic exposure to metabolite bufotenine. In addition, pretreatment of harmaline (5mg/kg, i.p.) led to 3.6- and 4.4-fold higher systemic exposure to 5-MeO-DMT (2mg/kg, i.p.), and 9.9- and 6.1-fold higher systemic exposure to bufotenine in Tg-CYP2D6 and wild-type mice, respectively. These findings indicate that MAOI largely affects 5-MeO-DMT metabolism and pharmacokinetics, as well as bufotenine formation that is mediated by CYP2D6.

The antiaggressive potency of (-)-penbutolol involves both 5-HT1A and 5-HT1B receptors and beta-adrenoceptors.

The relative importance of 5-HT1A and beta-adrenergic activities in the antiaggressive effects of (-)-penbutolol was studied in male mice. (-)-Penbutolol had high affinity for 5-HT1A receptors and beta-adrenoceptors, and antagonized the 5-methoxy-N,N-dimethyltryptamine (5-MeODMT)-induced 5-HT syndrome and the 8-hydroxy-2-(di-n-propylamin)tetralin (8-OH-DPAT)-induced discriminatory stimulus in rats. (-)-Penbutolol abolished aggressive behaviour (ED50 = 56 mumol/kg), and reversed the antiaggressive effects of 8-OH-DPAT and 1-(3-trifluoromethylphenyl)piperazine (TFMPP) (ED50 = 8.1 and 2.1 mumol/kg, respectively). (N-[2-[4-(2-Methoxyphenyl)-1-piperazinyl]ethyl-N-(2-pyridinyl) cyclohexanecarboxamide (WAY 100635) reversed the antiaggressive effects of 8-OH-DPAT (ED50 = 0.012 mumol/kg), but did not affect the antiaggressive effects of TFMPP. The antiaggressive effect of a submaximal dose of 8-OH-DPAT was markedly potentiated by beta-adrenoceptor antagonists without 5-HT1A receptor affinity, whereas (-)-penbutolol was effective at only one dose (4.5 mumol/kg). In conclusion, the 5-HT1A receptor antagonistic potency of (-)-penbutolol in aggressive mice is attenuated by beta-adrenoceptor-induced facilitation of serotonergic neurotransmission.

Increased dopamine turnover in the ventral striatum by 8-OH-DPAT administration in the rat.

The administration of the 5-HT1A agonist 8-OH-DPAT (0.8 mumols kg-1 s.c.-40 min) produced an increase in dopamine (DA) turnover, estimated by the quotient (DOPAC + HVA) DA-1, in the ventral striatum of the rat. No statistically significant effects were obtained in the dorsal striatum. The accumulation of 3-MT in pargyline-treated animals (375 mumols kg-1 s.c.-60 min) was not affected by 8-OH-DPAT treatment (0.15-2.4 mumols kg-1 s.c.-30 min). These findings indicate that 8-OH-DPAT has weak antagonist properties at striatal DA receptors in normal rats. Both the 5-HT1A agonist flesinoxan (0.06-17.8 mumos kg-1 s.c. -50 min) and the mixed 5-HT1 and 5-HT2 agonist 5-MeODMT (1.6-26.0 mumols kg-1 s.c.-50 min) produced a decrease in forebrain 5-HTP accumulation (striatum and neocortex), following decarboxylase inhibition by means of NSD-1015 in reserpine treated rats, indicating stimulation of central 5-HT receptors by these two compounds. At the same time, the DOPA accumulation by the ventral striatum was decreased by flesinoxan and increased by 5-MeODMT treatment. These observations show that, under these conditions, the decrease in DA synthesis is not directly coupled to the decreased 5-HT synthesis produced by flesinoxan, as previously demonstrated for 8-OH-DPAT. Taken together with previous observations, the present results suggest that 8-OH-DPAT, depending on the experimental conditions, is an agonist or antagonist at striatal DA receptors, in all probability due to partial DA receptor agonist properties of the compound.

The serotonin (5-HT) autoreceptor in the hippocampus of the rabbit: role of 5-HT biophase concentration.

Slices of hippocampus from the rabbit were preincubated with [3H]5-HT), then superfused continuously and twice stimulated electrically. The stimulation-evoked overflow of tritium was inhibited by the 5-HT autoreceptor ligands 5-carboxamido-tryptamine (5-COHT), 5-HT, 5-methoxy-N,N-dimethyl-tryptamine (5-MeOMT), (+/-)-8-hydroxy-2-(di-n-propylamino)tetralin (8-OH-DPAT), methysergide and (+/-)-cyanopindolol in a concentration-dependent manner. These effects were competitively inhibited by the 5-HT autoreceptor antagonists, metitepin and metergoline. (+/-)-Cyanopindolol also reduced the evoked release of 5-HT from slices of cortex from the rat. The inhibitor of the uptake of 5-HT, 6-nitroquipazine diminished the autoreceptor-mediated depression of release of 5-HT. In cortex tissue from the rat, 6-nitroquipazine reversed the decreased release of 5-HT, due to (+/-)-cyanopindolol, to a facilitation. The disinhibition of the release of 5-HT by autoreceptor antagonists was further enhanced by 6-nitroquipazine. Non-linear regression analysis of concentration-response curves for 5-COHT yielded the following pKd of endogenous 5-HT at the autoreceptor: 7.753 +/- 0.116. This value corresponds to the pKd of 5-HT at the 5-HT1B binding site. The 5-HT biophase concentration at the autoreceptor of 10(-8.220 +/- 0.132)M was markedly enhanced by 6-nitroquipazine (10(-6)M) to 10(-7.476 +/- 0.132)M. It is concluded that the 5-HT autoreceptor belongs to the 5-HT1B subtype of receptor; the corresponding 5-HT biophase concentration can be estimated quantitatively; 8-OH-DPAT decreased the evoked release of 5-HT through both 5-HT autoreceptors and alpha 2-heteroreceptors and (+/-)-cyanopindolol acts as partial agonist at the 5-HT autoreceptor.

Study of metabolism of psychotomimetic indolealkylamines by rat tissue extracts using liquid chromatography.

The use of a series of liquid chromatographic techniques involving cation-exchange, reverse-phase and normal-phase chromatography has permitted the separation and characterisation of a number of metabolites of the psychotomimetic indolealkylamines N,N-dimethyltryptamine and 5-methoxy-N,N-dimethyltryptamine which were isolated following incubation of these compounds with rat tissue extracts. In liver, kidney and brain tissue extracts the routes of metabolism identified included oxidative deamination, N-demethylation, O-demethylation and N-oxidation. The quantitative significance of individual routes of metabolism in these tissues was assessed using N,N-dimethyltryptamine as a substrate.

In vivo metabolism of 5-methoxy-N,N-dimethyltryptamine and N,N-dimethyltryptamine in the rat.

Following intraperitoneal administration, 5-methoxy-N,N-dimethyltryptamine and N,N-dimethyltryptamine are subject to both a very rapid uptake into, and clearance from, all tissues examined. The current studies in vivo confirm previous in vitro observations that the routes involved in the metabolism of these compounds include oxidative deamination, N-demethylation, O-demethylation, and N-oxidation. The analysis of metabolic profiles in various tissues led to the identification of the N-oxides as major metabolites. The successful inhibition and redirection of metabolism away from the indole acids towards the parent compounds and their structurally unique metabolites were demonstrated in animals pretreated with iproniazid.

Evidence for 5-HT2 involvement in the mechanism of action of hallucinogenic agents.

The affinities (Ki values) of twenty two psycho-active agents, including LSD, 5-OMe DMT and a series of phenalkylamine derivatives, for cortical 5-HT1 and 5-HT2 binding sites were compared with two measures of behavioral activity. It was found that a significant correlation (r = 0.938) exists between the 5-HT2 binding affinities of these agents and their ED50 values as determined in tests of stimulus generalization using 1-(2,5-dimethoxy-4-methylphenyl)-2-aminopropane (DOM) as the training drug. Furthermore, for fifteen of these agents where human data were available, a significant correlation (r = 0.924) also exists between 5-HT2 binding affinities and their human hallucinogenic potencies. The results of this study suggest that the mechanism of action of these agents involves 5-HT2-related events.

The distribution of serotonin binding sites in the hippocampal region of the rat brain. An autoradiographic study.

The distribution of serotonin binding sites was studied in the rat hippocampal region by using contact-film autoradiography after in vitro incubations of brain sections with 5-[3H]hydroxytryptamine, [3H]spiperone, and [3H]ketanserin, respectively. Biochemical studies of the 5-[3H]hydroxytryptamine binding to sections cut through the hippocampal region showed that at saturating concentrations of 5-[3H]hydroxytryptamine (2-2.5 nM) the specific binding was at least 50% of the total. The 5-[3H]hydroxytryptamine binding sites were found to be heterogeneously distributed within the hippocampal region with the highest densities present in the following parts: layers I and II and layers IV through VI of the entorhinal area, the radial layer of the subiculum and subfield CA1 of the Ammon's horn and the molecular layer of the area dentata. Moderate to low densities of binding was observed in layer III of the entorhinal area, the pre- and parasubiculum, the stratum pyramidale of the Ammon's horn, and the granular cell layer of the area dentata. Removal of the 5-hydroxytryptamine nerve terminals by systemic injections of the 5-hydroxytryptamine neurotoxin parachloroamphetamine resulted in no detectable reductions of 5-[3H]hydroxytryptamine binding in any brain region. Lesions of hippocampal cell bodies by intrahippocampal injections of ibotenic acid prevented the binding of 5-[3H]hydroxytryptamine within the area of the cell loss. Comparisons between the distribution of 5-hydroxytryptamine immunoreactive nerve terminals and the 5-[3H]hydroxytryptamine binding sites showed that in some areas of sparse 5-hydroxytryptamine innervation the 5-[3H]hydroxytryptamine binding was close to background (e.g. the pyramidal cell layer, the stratum lucidum) whereas in areas with little 5-[3H]hydroxytryptamine binding (e.g. layer III of the lateral entorhinal area, the presubiculum) a very dense 5-hydroxytryptamine innervation was found. The hippocampal 5-[3H]hydroxytryptamine binding was displaced neither by ketanserin (1 microM) nor by spiperone (1 microM), two drugs that bind to cortical 5-hydroxytryptamine2 receptors in the rat brain. Furthermore, the pattern of hippocampal [3H]spiperone binding differed considerably from that of 5-[3H]hydroxytryptamine. The [3H]ketanserin binding in the hippocampal region did not exceed background levels, except in the hilus of area dentata in the ventral hippocampus and entorhinal layer VI at the same level, where moderate binding was found.(ABSTRACT TRUNCATED AT 400 WORDS)

Ontogeny of N,N-dimethyltryptamine and related indolealkylamine levels in neonatal rats.

The present study deals with the measurement of the brain levels of the two potent hallucinogens N,N-dimethyltryptamine (DMT) and 5-methoxy-N,N-dimethyltryptamine (OMB), the biogenic amine tryptamine (TA), and its condensation product 1,2,3,4-tetrahydro-beta-carboline (THBC) in rats of various ages. Using gas chromatography-mass spectrometry with isotope dilution, we detected DMT, OMB, and THBC in neonatal rats from birth. DMT levels remained low until days 12 and 17 at which time they increased significantly and then returned to the initial low levels for all subsequent ages. The levels of OMB were higher than those measured for DMT with the highest levels being observed at days 12 and 17, and also on day 31. However, the levels for OMB showed much more variation. Although elevated levels of DMT and OMB have been correlated with stress, there are no known functions for these compounds. TA levels remained below detection limits until day 19. THBC levels were observed to be highest on days 22 and 31. The role that THBC plays in mammalian tissues is not known.

Effect of 5,7-dihydroxytryptamine on serotonergic control of prolactin secretion and behavior in rats.

The intracisternal administration of 5,7-dihydroxytryptamine (5,7-DHT) to rats resulted in a potentiated response to 5-hydroxytryptophan (5-HTP) when the animals were tested 30 days later. The 5-HTP-induced changes include elevation of serum prolactin, decrease in operant responding, and the magnitude of the "serotonin behavioral syndrome" observed after 5-HTP administration. The serotonin concentration in brains of 5,7-DHT-treated animals reached maximum earlier and remained elevated longer than that of controls following administration of 5-HTP. Brain norepinephrine and dopamine concentration were not affected by 5-HTP in either group of animals. The increase in serum prolactin concentration elicited by administration of the serotonergic agonists quipazine or 5-methoxy-N,N-dimethyltryptamine and by the serotonin uptake inhibitor fenfluramine also was potentiated by pretreating rats with 5,7-DHT. These data suggest that both serotonergic receptor supersensitivity and the absence of presynaptic uptake sites contribute to the enhanced responses to 5-HTP occurring in rats previously treated with 5,7-DHT. The findings further demonstrate that both behavioral and hormonal measures can be used to assess the sensitivity of serotonergic receptors and indicate that 5,7-DHT may be useful in evaluating the role of serotonergic neurons in neuroendocrine function.