The present study demonstrated a remarkable in vivo effect of 5-MeO-DIPT on brain neurotransmission by showing that 5-MeO-DIPT increased extracellular levels of DA, 5-HT, and glutamate in the rat striatum, nucleus accumbens, and frontal cortex. We also observed changes produced by 5-MeO-DIPT in tissue contents of DA and 5-HT as well as their metabolites DOPAC, HVA, and 5-HIAA in various regions of the rat brain. Furthermore, our data revealed a dose-dependent and progressive oxidative damage of cortical DNA by 5-MeO-DIPT. In addition, 5-MeO-DIPT evoked head twitches and potentiated forepaw treading induced by 8-OH-DPAT, which suggests activation of 5-HT2A and 5-HT1A receptors, respectively.

SERT inhibition by 5-MeO-DIPT (Blough et al. 2014) enhances 5-HT level which then affects all subtypes of serotonin receptors in the brain. In addition, 5-MeO-DIPT having by itself affinity for 5-HT1A, 5-HT2A, and 5-HT2C serotonin receptors (Fantegrossi et al. 2006) may potentiate the effects of endogenous serotonin. This interaction can lead to complex behavioral and neurochemical responses. In our study, 5-MeO-DIPT at the dose of 10 mg/kg elicited head twitches commonly used as a model of a hallucinogenic effect mediated through serotonin 5-HT2A receptors (Halberstadt 2015). The response to 5-MeO-DIPT (10 mg/kg) was similar in potency to the effect of selective 5-HT2A receptor agonist (±)DOI (2.5 mg/kg). In another animal model used in our work, 5-MeO-DIPT (5–10 mg/kg) strongly potentiated forepaw treading induced by 8-OH-DPAT, which is thought to be mediated via activation of postsynaptic 5-HT1A receptors (Sanchez et al. 1996; Sloviter et al. 1978). These data suggest that 5-MeO-DIPT enhances serotonin transmission in the brain and activates 5-HT1A and 5-HT2A receptors.

Blockade of intraneuronal serotonin transport by 5-MeO-DIPT led to a dose-dependent increase in extracellular 5-HT level in the rat striatum, nucleus accumbens, and frontal cortex as found in our study. Previous in vitro data of Sogawa et al. (2007) showed that micromolar concentrations of 5-MeO-DIPT inhibited [3H]5-HT uptake in COS cells transfected with SERT cDNA as well as in rat brain synaptosomes. The range of doses (5–20 mg/kg) used in our study seems to be effective in blocking SERT as submicromolar concentrations of 5-MeO-DIPT and its metabolites were found in rat urine samples after oral administration at the dose 5 mg/kg (Kanamori et al. 2006).

Enhancement of DA content in the mesocorticolimbic dopaminergic neurons is responsible for ability of several psychostimulant drugs to cause drug dependence and addiction. However, hallucinogens are not considered as reinforcing drugs (O’Brien2001). In contrast to LSD, 5-MeO-DIPT, like other tryptamines (e.g., 5-MeO-DMT), does not display affinity for dopamine receptors and has a low activity in blocking dopamine transporter DAT (Halberstadt and Geyer 2011; Sogawa et al. 2007). Nevertheless, we show evidence that 5-MeO-DIPT at doses of 10–20 mg/kg is able to increase DA release in the striatum, nucleus accumbens, and frontal cortex; however, at a dose of 5 mg/kg, it was less effective. The possible mechanism responsible for this activity of 5-MeO-DIPT in increasing DA release may be related to the 5-MeO-DIPT-induced stimulation of presynaptic 5-HT2A receptors located on DA neuronal terminals. The data supporting our results were reported by Pehek et al. (2001) who showed that a stimulation of DA release by potassium in the rat prefrontal cortex was mediated by 5-HT2A receptors. Other researchers demonstrated that the effect of 5-HT2A agonist (±)DOI on DA release in the rat nucleus accumbens and the rat striatum was antagonized by 5-HT2A antagonists ketanserin (Yan 2000) or SR 46349B (Lucas and Spampinato 2000). Alternatively, enhancement of DA release by 5-MeO-DIPT may be mediated through the activation of somatodendritic 5-HT2A receptors in the VTA. Those receptors might directly affect local dendritic release of DA and subsequently increase extracellular DA level in mesolimbic or mesocortical DA terminals as suggested by Celada et al. (2001) and Vazquez-Borsetti et al. (2009). Moreover, high affinity of tryptamine hallucinogens for 5-HT1A receptors reported by deMontigny and Aghajanian (1977), and Titeler et al. (1988) suggests that these receptors may play a role in controlling activity of DA neurons. 5-HT1A receptors localized on GABA-ergic interneurons in limbic and cortical brain regions (Hamon et al. 1990; Pazos and Palacios 1985) may disinhibit descending glutamatergic pathways which can subsequently stimulate DA cells. The data presented by Tanda et al. (1994), Sakaue et al. (2000), and Wędzony et al. (1996) support our conclusion, as they demonstrated that selective 5-HT1A receptor agonists, R(+)-8-OH-DPAT or ipsapirone, increased DA release in the frontal cortex.

We found that 5-MeO-DIPT increased extracellular glutamate level in the striatum at all doses and only at higher doses in the nucleus accumbens and frontal cortex. The enhancement of glutamate release by 5-MeO-DIPT may depend on activation of several subtypes of serotonin receptors, and therefore may vary between brain regions. As noted by other researchers, 5-MeO-DIPT acting at postsynaptic 5-HT2A receptors on pyramidal cells enhances glutamate release (Beique et al. 2007). However, 5-HT2A receptors are co-localized on cortical pyramidal cells with serotonin 5-HT1A receptors (Martin-Ruiz et al. 2001), where the two receptor types have opposing effects (Araneda and Andrade 1991). In our study, the decrease in glutamate release caused by the lowest dose of 5-MeO-DIPT in the nucleus accumbens or lack of effect in the frontal cortex suggests that 5-MeO-DIPT at small concentrations preferentially activates 5-HT1A receptors, causes inhibition of pyramidal cells, and subsequently decreases glutamate release. At higher doses, the effect exerted by 5-HT1A receptors is opposed by 5-HT2A receptors, which results in the stimulation of glutamate release. In fact, in vitro affinity of 5-MeO-DIPT at 5-HT1A receptors was found in nM, while at 5-HT2A receptors in μM range of concentrations (Fantegrossi et al. 2006). Therefore, the effect mediated via 5-HT1A receptor may be counteracted by 5-HT2A receptor activated by higher concentration of 5-MeO-DIPT.

The finding that hallucinogens act as agonists of 5-HT2C receptor suggests that these compounds exert some effects via the 5-HT2C receptor subtype. However, there is now a consensus that ability of (±)DOI to induce head-twitch response is not blocked by 5-HT2A/C antagonists (Fantegrossi et al. 2010; Schreiber et al. 1995; Wettstein et al. 1999). It also appears that activity at the 5-HT2C receptor attenuates many of the behavioral effects of hallucinogens. For instance, the ability of (±)DOI to reduce prepulse inhibition in rats was reversed by the 5-HT2C selective agonist WAY-163,909 (Marquis et al. 2007). Furthermore, Halberstadt et al. (2009) demonstrated that 5-HT2A and 5-HT2C receptors exerted opposing effects on locomotor activity in mice. Similar findings have been reported for head-twitch response in mice (Fantegrossi et al. 2010) or in rats (Vickers et al. 2001). Therefore, some effects observed in our study, such as a decrease in DA or glutamate release by a low dose of 5-MeO-DIPT in the striatum or in the nucleus accumbens, respectively, may result from a modulating role of 5-HT2C receptor. However, exact mechanism of the interaction between serotonin receptor subtypes in their effect on brain neurotransmission needs further studies.

In terms of the 5-MeO-DIPT effect on 5-HT tissue content, our data indicate that an increase or lack of changes in serotonin level and a decrease in 5-HIAA level are consistent with the hypothesis that 5-MeO-DIPT, through blocking intraneuronal transport of 5-HT inhibits serotonin metabolism in all studied brain regions. The observed decrease in DA, DOPAC, and HVA tissue levels by all doses of 5-MeO-DIPT in the rat striatum and two higher does in the nucleus accumbens and the frontal cortex appears to be related to a feedback inhibition of DA synthesis as a response to stimulation of dopamine receptors by an increased synaptic pool of DA. On the other hand, a deficit in tissue content of DA and its metabolites may be associated with neurotoxic effect exerted by 5-MeO-DIPT on presynaptic DA terminals.

The possible neurotoxic effects of 5-MeO-DIPT seem to be supported by our findings obtained with the use of the comet assay. It was demonstrated that 5-MeO-DIPT given at a single dose produced DNA single and double-strand breaks in the rat cortex. The magnitude of tail moment reflecting the extent of DNA damage was time- and dose-dependent when measured 72 h and 60 days after administration. A similar effect on DNA damage was observed after treatment of rats with the 5-HT2A agonist (±)DOI and MDMA. The oxidative damage of DNA was reported in brains of animals treated chronically with high doses of MDMA and methamphetamine (Frenzilli et al. 2007; Johnson et al. 2015). The mechanism of DNA oxidation by amphetamine derivatives is related to an oxidative stress and the formation of highly reactive free radicals. Excessive release of DA and glutamate by MDMA or methamphetamine leads to formation of reactive oxygen and nitrogen species as well as reactive quinones, which can damage DNA (Halliwell and Whiteman 2004). Our study is the first to show genotoxic effect of a tryptamine hallucinogen. An increase in DA and glutamate release by 5-MeO-DIPT reported in the present study suggests that DA and glutamate play a role in the induction of oxidative stress. However, other factors such as protective mechanisms and levels of antioxidants which control free radical generation, may also be affected by 5-MeO-DIPT. Therefore, further studies are needed to elucidate the possible mechanism of 5-MeO-DIPT genotoxicity. It has to be emphasized that oxidative stress which has been implicated in neurodegenerative diseases, such as Parkinson’s and Alzheimer’s diseases (Fahn and Sulzer 2004; Halliwell 2006) and which causes oxidative DNA damage seems to be linked with memory loss and cognitive dysfunction in rats (Liu et al. 2002). Alteration in the ability of rats to perform certain cognitive tasks in Cincinnati water maze (Williams et al. 2007) or impairments in working memory (Compton et al. 2011) by 5-MeO-DIPT may be due to persistent and progressive modification of the cortical cell function. All these observations suggest that tryptamine hallucinogens need further extensive studies as they are among the most popular groups of illicit drugs.

In summary, the results of our study demonstrate that exposure of rats to the tryptamine hallucinogen 5-MeO-DIPT produces changes in extracellular serotonin, dopamine, and glutamate levels in cortical and subcortical rat brain regions. Our findings also support the conclusion that hallucinations after administration of tryptamine analogues may be mediated by changes in glutamatergic neurotransmission. The progressive oxidative damage of DNA produced by a single dose of 5-MeO-DIPT indicates development of oxidative stress and suggests marked neurotoxicity of this drug.