Single dose of MDMA causes extensive decrement of serotoninergic fibre density without blockage of the fast axonal transport in Dark Agouti rat brain and spinal cord

Abstract

Prolonged neurotoxicity of the recreational drug, MDMA (3,4-methylenedioxymethamphetamine) on serotoninergic axon terminals has been suggested. The effect of a single (15 mg/kg) dose of intraperitoneally administered MDMA on serotoninergic fibre density, defined by tryptophan hydroxylase (TpH) and serotonin transporter (5-HTT) immunoreactivity, has been evaluated in the spinal cord and brain areas in Dark Agouti rats, 7 and 180 days after MDMA applications. Immunostaining for amyloid precursor protein (APP) has been performed to examine possible defects of the fast axonal transport, and 5-HTT mRNA expressions were quantified in neurones of medullary raphe nuclei. Seven days after MDMA treatment, a substantial decrease in the density of TpH-immunoreactive fibres was detectable in the frontal cortex, the caudate-putamen, the CA1 region of the hippocampus, and marked decreases were found in the spinal cord. These changes in TpH density showed a high correlation with 5-HTT densities. In contrast, APP-immunoreactive axonal bulbs were not detected in any of the brain regions studied. Seven days after MDMA administrations, significantly elevated 5-HTT mRNA expressions were found in the raphe pallidus and obscurus. Our results suggest that a single dose of MDMA elicits widespread depletion of TpH and 5-HTT immunoreactivity in serotoninergic axons without morphological sign of the blockage of the fast anterograde axonal transport. Our results do not support the notion of MDMA-induced axotomy of serotoninergic neurones. The up-regulation of 5-HTT mRNA expressions 1 week after MDMA injections might indicate the potential recovery of the serotonin system.

Recovery and aging of serotonergic fibers after single and intermittent MDMA treatment in dark agouti rat

“…Tph immunohistochemistry is an early and sensitive method of demonstrating vulnerability caused by MDMA, reflecting the morphology of serotonergic axons as well as the actual level of Tph protein (Adori et al, 2006;Xie et al, 2006;Kovacs et al, 2007). Moreover, the Tph immunohistochemistry results are in agreement with findings from densitometry analysis of 5-HTT immunoreactivity, which is another marker of the damage to serotonergic fibers after MDMA treatment (Kovacs et al, 2007;Kirilly et al, 2008;Ando et al, 2010).…”

“…2,4,5,[7][8][9], mouse monoclonal Tph antibody exclusively stained the raphe nuclei and some other brainstem neuronal perikarya corresponding anatomically to the serotonergic neuronal population (not shown). Moreover, rabbit polyclonal 5-HT transporter (5-HTT), another serotonergic marker, showed co-localization with mouse monoclonal anti-Tph, providing further evidence that the mouse monoclonal Tph antibody is appropriate for the detection of serotonergic axons in our sections (Adori et al, 2006;Kovacs et al, 2007).…”

“…Therefore, this antibody stains Tph, TyH, and PaH in Western blot and frozen sections. However, in fixed and paraffin-embedded sections, it selectively stains Tph but not TyH and PaH (manufacturer's technical information; Haan et al, 1987;Kloppel et al, 2001;Kovacs et al, 2007;Adori et al, 2006Adori et al, , 2010aAdori et al, , 2010b. In our sections, in addition to the widespread axonal immunostaining (see Figs.…”

“…) in Dark Agouti rats have already been characterized (O'Shea et al, 1998;Balogh et al, 2004;Kovacs et al, 2007). In the Dark Agouti rat strain, the CYP2D1 enzyme, which is an important factor in MDMA metabolism and is the equivalent of human CYP2D6 (debrisoquine hydroxylase), shows decreased activity.…”

Long‐lasting neuroprotective effect of sildenafil against 3,4‐methylenedioxymethamphetamine‐ induced 5‐hydroxytryptamine deficits in the rat brain

“…These brain areas were chosen because the neurotoxic effects of MDMA are limited to the 5-HT terminals of the serotonergic neurotransmitter system, insofar as cell bodies in the raphe nuclei are not damaged by MDMA (O'Hearn et al, 1988;Kovács et al, 2007). Briefly, samples were injected using an automatic sample injector (Waters 717 plus) onto a Spherisorb ODS-2 reverse-phase C18 column (5 lm, 150 3 4.6 mm; Teknokroma, San Cugat del Valles, Spain) connected to a Decade amperometric detector (Antec Leyden, Zoeterwoude, The Netherlands), with a glassy carbon electrode maintained at 0.7 V with respect to an Ag/AgCl reference electrode.…”