Introduction
Tetrahydroisoquinolines are a group of endogenous compounds that are present in the mammalian brain (Abe et al.
2005; Yamakawa et al.
1999; Yamakawa and Ohta
1997,
1999). Some of these compounds, such as 1-benzyl-1,2,3,4-tetrahydroisoquinoline (1BnTIQ), have shown neurotoxic properties and are considered to be involved in the pathogenesis of Parkinson’s disease (PD) (Abe et al.
2001a; Kotake et al.
1995). Kotake et al. (
1995) indicated that the concentration of 1BnTIQ in the cerebrospinal fluid (CSF) of parkinsonian patients was three times higher than that in the CSF of the control group. Previous studies revealed that chronic treatment with 1BnTIQ induces parkinsonian-like symptoms in mammals (Kotake et al.
1995,
2003,
2010). 1BnTIQ accumulates in dopaminergic neurons and can lead to parkinsonian symptoms. 1BnTIQ evoked strong activation of the oxidative MAO-dependent catabolic pathway (Wąsik et al.
2009). In addition, 1BnTIQ significantly inhibits the COMT-dependent O-methylation pathway. This mechanism of action leads to an increase in dopamine oxidation, and as a consequence, leads to an increase in reactive oxygen species (ROS) formation in dopaminergic neurons. Moreover, in vitro studies revealed that 1BnTIQ induces cell death via apoptosis and produces an increase in the formation of the active caspase-3 protein fragments (Shavali and Ebadi
2003). These data suggest that multiple administrations of 1BnTIQ might serve as an adequate animal model of the progressive process of PD. In contrast, 1,2,3,4-tetrahydroisoquinoline (TIQ), and especially its methyl derivative, 1-methyl-1,2,3,4-tetrahydroisoquinoline (1MeTIQ), have shown neuroprotective effects in the brain (Abe et al.
2001b; Antkiewicz-Michaluk et al.
2003,
2004,
2006; Wąsik et al.
2016). TIQ and 1MeTIQ are reversible MAO inhibitors that strongly block the MAO-dependent oxidative pathway and simultaneously increase the COMT-dependent O-methylation catabolic pathway. Therefore, both substances possess antioxidant properties. These compounds inhibit free radical formation and abolish H
2O
2 generation from dopamine via the Fenton reaction (Singer and Ramsay
1995; Antkiewicz-Michaluk et al.
2006; Patsenka and Antkiewicz-Michaluk
2004). Additionally, 1MeTIQ acts as a natural scavenger of free radicals. From a clinical point of view, the lack of a tolerance for its neuroprotective action after chronic treatment is both interesting and important (Antkiewicz-Michaluk et al.
2001; Wąsik et al.
2016).
To continue our previous studies, we would like to investigate the effects of TIQ and 1MeTIQ, as earlier demonstrated neuroprotective compounds, on the dopamine release in vivo in an animal model of PD induced by chronic administration of 1BnTIQ. Using in vivo microdialysis methodology, we measured the impact of acute and chronic treatment with TIQ and 1MeTIQ on 1BnTIQ-induced disorders of dopamine release in the rat striatum. In addition to the biochemical research, the behavioral test was carried out to check the influence of repeated administration of 1BnTIQ on the motor activity of rats.
Discussion
The chronic administration of 1BnTIQ produced clear and significant disturbances in the function of dopaminergic neurons, which led to an increase in dopamine release in the striatum. Based on our previous experiments, we confirmed that chronic treatment with a low dose of 1BnTIQ (25 mg/kg i.p.) damaged the dopamine storage mechanisms causing its non-physiological rise in the extracellular space in the rat striatum (Wąsik et al.
2009,
2014). The main finding of the present study is that chronic administration of TIQ and 1MeTIQ completely prevented the disorders on dopamine release induced by multiple administrations of 1BnTIQ. Dopamine, the main neurotransmitter involved in motor control and Parkinson’s disease, is metabolized both intra- and extraneuronally. The extraneuronal dopamine metabolite, 3-MT, which is present in the synaptic cleft at relatively low concentrations comparable to dopamine, is considered to be a marker of dopamine release (Karoum et al.
1994). However, in contrast to intraneuronal dopamine metabolite, DOPAC and the final metabolite HVA, 3-MT may be biologically active. It was demonstrated previously that 3-MT has a considerable affinity as antagonist for noradrenergic α1 and dopamine D1 and D2 receptors in rat brain (Antkiewicz-Michaluk et al.
2008). In the behavioral tests chronic administration of 1BnTIQ-induced hyperactivity, while multiple injection of both, TIQ and 1MeTIQ significantly decreased the rats locomotor activity (Fig.
1a, b). Additionally, in the combined groups, TIQ and 1MeTIQ inhibited 1-BnTIQ-induced hyperactivity. In that light, the mechanism of the action of TIQ and 1MeTIQ, leading to an increase the concentration of 3-MT antagonism to 1BnTIQ-produced hyperactivity may be explained by its inhibition to catecholaminergic receptors.
Although all substances used in the present study belong to the same chemical group, their mechanisms of action are completely opposite. 1BnTIQ activates the dopamine oxidation pathway, which leads to the elevation in free radical production (Wąsik et al.
2014). On the other hand, TIQ and 1MeTIQ exhibit a contrasting molecular mechanism, and act as reversible MAO inhibitors, blocking the production of free radicals and activating the COMT-dependent O-methylation pathway (Antkiewicz-Michaluk et al.
2001; Wąsik et al.
2009). Our present in vivo study demonstrated that multiple administrations of 1BnTIQ produced a significant and long-lasting increase (approximately 300 % in comparison to the saline group) in dopamine release in the rat striatum, measured in the basal samples 24 h after its 13th injection (1 h before the last 14th injection). Such permanent increase in dopamine levels in the extracellular space after the chronic administrations of 1BnTIQ may, like the treatment with psychostimulants, leads to neurotoxic effects. After the 14th (last) dose of 1BnTIQ, dopamine release was elevated up to 500 % of the saline group; however, at the same time, no change in 3-MT concentration was observed. Moreover, 3-MT is considered as the most reliable indicator of dopamine release into the synaptic cleft (Egan et al.
1991; Karoum et al.
1994). Additionally, as demonstrated previously, 3-MT, in contrast to DOPAC and HVA, is an active metabolite of dopamine and possesses its own receptor activity (Antkiewicz-Michaluk et al.
2008; Alachkar et al.
2010). It is well known that elevation of dopamine release induces an increase of dopaminergic activity and leads to hyperactivity. The results of the in vivo microdialysis study are in agreement with the behavioral locomotor activity test in which chronic treatment with 1BnTIQ-produced hyperactivity in rats (Fig.
1a, b).
As we demonstrated earlier, 1BnTIQ similarly to reserpine (a specific vesicular monoamine transporter 2 (VMAT2) inhibitor) significantly depleted striatal dopamine (Wąsik et al.
2009). We postulated that 1BnTIQ might damage VMAT2 in dopaminergic neurons, leading to the pathological release of dopamine into the cytosol, and increased its MAO-dependent oxidation and free radical production (Wąsik et al.
2014). On the other hand, 1BnTIQ in low micromolar concentrations significantly inhibited the dopamine reuptake in slices of the rat striatum (Patsenka et al.
2004). Similarly, Okada et al. (
1998) demonstrated that 1BnTIQ inhibited the uptake of [3H]dopamine by the dopamine transporter expressed in HEK293 cells. We suggest that DAT is responsible for the selective transport of 1BnTIQ into the dopaminergic neurons, leading to the 1BnTIQ neurotoxicity correlated with impaired dopamine storage via inhibition of VMAT2 (Wąsik et al.
2009). Since TIQ and 1MeTIQ have also affinity to DAT, both substances may restrict access of 1BnTIQ to the DAT (Patsenka et al.
2004). This mechanism results in a reduction of penetration of 1BnTIQ into the neurons. The present study revealed that acute treatment with TIQ induced an increase in dopamine release (approximately 300 %), while 1MeTIQ showed a similar effect of a lesser magnitude (Figs.
2a,
4a). Taking into account that both investigated compounds, TIQ and 1MeTIQ, act as reversible MAO inhibitors, they shift dopamine catabolism toward the COMT-dependent O-methylation pathway, which leads to a significant increase in the extraneuronal concentration of the dopamine metabolite 3-MT (Antkiewicz-Michaluk et al.
2001; Patsenka and Antkiewicz-Michaluk
2004). In the present paper, such properties of TIQ and 1MeTIQ were confirmed in our in vivo microdialysis studies which demonstrated an increase in 3-MT level by approximately 500 % in the case of acute administration and up to 2500 % versus saline group after chronic treatment. It is worth emphasizing, as previously demonstrated that 3-MT, an extraneuronal dopamine metabolite, shows affinity for α1-adrenergic and D1 and D2 receptors as an antagonist, and may play an important role as an inhibitory regulator, counteracting the excessive stimulation of catecholaminergic neurons (Antkiewicz-Michaluk et al.
2008; Alachkar et al.
2010). Thus, a high concentration of 3-MT may play an important role in locomotor activity as dopamine D2 and noradrenergic α1 receptors antagonists (Antkiewicz-Michaluk et al.
2008). Such mechanism of action of 3-MT leads to the paradoxical effect, where despite the increase in dopamine release, we observed decrease in the locomotor activity in rats after treatment with both, TIQ and 1MeTIQ (Fig.
1a, b).
In the combined treatment group, an acute dose of TIQ given before the last dose of 1BnTIQ only slightly reduced the effect of 1BnTIQ, whereas acute administration of 1MeTIQ produced a clear decrease in dopamine release (Figs.
2a,
4a). In both combined treatment groups, a massive increase in 3-MT levels was observed (2500 and 2000 %, respectively; Figs.
3a,
5a). Such an increase in the 3-MT concentration may be associated with the retention of dopamine in the extracellular space, due to dopamine reuptake blockade by TIQ and 1MeTIQ, and catabolism of all available dopamine by the COMT-dependent O-methylation pathway. It is worth emphasizing that such a mechanism of action for TIQ and 1MeTIQ inhibits the production of free radicals, which are formed during the catabolism of dopamine by MAO-dependent oxidation, and consequently leads to a reduction in oxidative stress (Antkiewicz-Michaluk et al.
2006; Miller et al.
1996; Patsenka and Antkiewicz-Michaluk
2004).
Interestingly, in both combined treatment groups, chronic treatment with TIQ or 1MeTIQ completely antagonized the 1BnTIQ-induced pathological increase in dopamine release in the basal samples (Figs.
2b,
4b). Simultaneously, in these groups, we observed a huge elevation in the concentration of 3-MT after the last dose of TIQ and 1MeTIQ (up to 2500 and 2000 %, respectively) (Figs.
3b,
5b). These results are important because they demonstrate that chronic administration of both TIQ and 1MeTIQ can counteract the disturbances in dopamine release produced by multiple administrations of 1BnTIQ. We suggest that TIQ and 1MeTIQ given before each 1BnTIQ injection inhibit DAT and in this way block penetration of 1BnTIQ into the neurons. Since 1BnTIQ penetrates into cells via DAT, DAT is inhibited by prior injection of TIQ and 1MeTIQ; therefore, in the combined treatment groups, we did not observe the effect of 1BnTIQ (Patsenka et al.
2004). The data from the behavioral tests (locomotor activity) confirmed that a high concentration of 3-MT, which was observed in both joint treatment groups, acted as an endogenous neuroleptic, as previously demonstrated (Antkiewicz-Michaluk et al.
2008), and could block hyperactivity in rats (Fig.
1a, b). It is important to mention that pronounced increases in dopamine O-methylation in the COMT-dependent pathway may provide neuroprotection (Antkiewicz-Michaluk et al.
2001; Miller et al.
1996). It was also indicated that elevation of 3-MT in the process of dopamine O-methylation protected cells against oxidative stress (Miller et al.
1996). The neuroprotective properties of 1MeTIQ against different dopaminergic neurotoxins, including MPTP, 6-OHDA, rotenone, and 1BnTIQ, were also demonstrated in in vitro studies in cultured mesencephalic neurons (Kotake et al.
2005) and in ex vivo studies (Antkiewicz-Michaluk et al.
2011; Wąsik et al.
2016).