Evidence for dual serotonergic projections to neocortex: axons from the dorsal and median raphe nuclei are differentially vulnerable to the neurotoxin p-chloroamphetamine (PCA)

Previous studies have shown that there are morphologically dissimilar serotonergic (5-HT) axon types in rat cerebral cortex which are differentially sensitive to the neurotoxic effects of certain psychotropic drugs: methylenedioxyamphetamines (MDA and MDMA) and p-chloroamphetamine (PCA) cause degeneration of fine 5-HT axon terminals in cortex, while sparing beaded axons. Moreover, a recent anterograde transport study suggests that fine and beaded 5-HT axons arise from the dorsal raphe (DR) and median raphe (MR) nuclei, respectively. These data led us to propose that the DR projection to neocortex is selectively vulnerable to the neurotoxic effects of PCA, while the MR projection is resistant; this hypothesis was tested in the present study by comparing retrograde axonal transport of the fluorescent tracer Fluoro-Gold in PCA-treated and control rats. Using this method, only axons that survive PCA treatment can take up and transport the injected label back to the cell bodies of origin, thus allowing us to determine which raphe-cortical projections remain intact after PCA. The results show that PCA administration produces a loss of fine 5-HT axon terminals in neocortex and a concomitant reduction in the number of retrogradely labeled neurons in the DR (77% decrease), when compared to controls. In contrast, beaded 5-HT axon terminals are spared and the number of labeled neurons in the MR remains unchanged after PCA. These results demonstrate that DR and MR projections to cortex are differentially vulnerable to PCA: fine axon terminals arise from neurons in the DR and are highly sensitive to the neurotoxic effects, whereas beaded axons from the MR are resistant. We therefore propose that there are two anatomically and functionally separate 5-HT projections to cortex having different (1) nuclei of origin, (2) axon morphology, (3) regional distributions, and (4) pharmacological properties. Since the mood-altering substances MDA, MDMA, and PCA act specifically upon 5-HT axon terminals from the dorsal raphe nucleus, DR neurons may be preferentially involved in the control of affective state.     

The serotoninergic innervation of cerebral cortex: different classes of axon terminals arise from dorsal and median raphe nuclei

The objective of the present study was to characterize the morphology of serotoninergic axons in cerebral cortex of the rat and to determine whether dissimilar axon terminals arise from the dorsal vs. the median raphe nuclei. The anterograde tracer PHA-L was administered by iontophoresis into the dorsal (DR) and median (MR) raphe nuclei, and the morphologic features of the respective axonal projections from raphe to forebrain were analyzed. We have observed consistent structural differences between the axons from these two nuclei. Anterogradely labeled axons which arise from cells in the MR are characterized by large, spherical varicosities (type M axons) and by variations in axonal diameter. In contrast, DR fibers are very fine and typically have small, pleomorphic varicosities that are granular or fusiform in shape (type D axons). Similar features of serotonin (5-HT) axon morphology are also evident in 5-HT immunocytochemical preparations. In addition to structural differences, there is a differential topographic distribution of MR vs. DR fibers, with MR axons concentrated in particular areas of limbic cortex such as dentate gyrus, posterior cingulate, and entorhinal areas as well as in parietal cortex. Immunofluorescence with dual labels shows that over two-thirds of the raphe-cortical axons are serotoninergic. The dissimilarities in axon morphology indicate that individual raphe nuclei may form different patterns of synaptic organization. Based on the evidence that the dorsal and median raphe nuclei give rise to morphologically different axon terminals, we conclude that 5-HT axons in cortex may be subdivided into two distinct projections. This proposal is in accord with other, recent data showing that the two axon types have different pharmacologic properties and are likely to be functionally different.     

Changes of serotonin and dopamine metabolism in various forebrain areas of rats injected with morphine either systemically or in the raphe nuclei dorsalis and medianus

The regional brain metabolism of serotonin (5-HT) and dopamine (DA) was studied in rats injected with morphine either systemically or in the nuclei raphe medianus (MR) or dorsalis (DR). A subcutaneous injection of 10 mg/kg morphine significantly raised the levels of 5-hydroxyindoleacetic acid (5-HIAA) in the diencephalon, striatum, nucleus accumbens and cortex with no effect in the hippocampus. Similar changes in 5-HT metabolism were found in animals injected with 5 micrograms/0.5 microliter in the DR whereas morphine injected in the MR raised 5-HIAA levels only in the nucleus accumbens. A subcutaneous or direct injection of morphine in the DR significantly raised the levels of homovanillic acid (HVA) and dihydroxyphenylacetic acid (DOPAC) in the striatum and nucleus accumbens, but injection in the MR was ineffective. All the effects of morphine were blocked by naloxone, injected either intraperitoneally (1 mg/kg) or directly in the raphe nuclei (2 micrograms/0.5 microliter). Pretreatment with parachlorophenylalanine, an inhibitor of serotonin synthesis, significantly reduced the effect of morphine injected in the DR on dopamine metabolism in the striatum and nucleus accumbens. The data suggest that a major mechanism by which morphine increases 5-HT metabolism in the rat forebrain is activation of 5-HT cells in the nucleus raphe dorsalis, and this action may contribute to the increased DA metabolism found in the animal injected with morphine in this brain area.     

MDMA (ecstasy) effects on cultured serotonergic neurons: evidence for Ca-dependent toxicity linked to release

Animal studies have established a correlation between release of 5-hydroxytryptamine (5-HT) and the long-term reduction of 5-HT (toxicity) by 3,4-methylenedioxymethamphetamine (MDMA) with the S(+) enantiomer being more active than the R(-). Using a microculture system of fetal raphe neurons, the enantiomers of MDMA were tested to determine if a similar difference in potency existed. The results showed that the development of the uptake capacity of [3H]5-HT in 4-day cultures was half-maximally inhibited by a single application at time of plating of 5 X 10(-6) M S(+)-MDMA and 5 X 10(-5) M R(-)-MDMA. In order to determine if the Ca2(+)-independent release (chemically induced through the transporter protein and inhibited by reuptake blockers) or the Ca2(+)-dependent release (K(+)-induced and inhibited by presynaptic receptors) contributed to the toxicity, fluoxetine and D1 and alpha 2 agonists were studied. The results showed that both forms of release were involved in the loss of [3H]5-HT uptake capacity, with the direct MDMA-induced Ca2(+)-independent (fluoxetine-sensitive) release being the first step. Evidence from binding studies indicates that MDMA has a micromolar affinity for the 5-HT2 receptor, and our studies in culture showed that ketanserin, a specific 5-HT2 antagonist, was effective at attenuating the effects of S(+)-MDMA on the development of the [3H]5-HT uptake capacity by the cultured raphe neurons. The 5-HT2 receptor is linked to increased intracellular Ca2+ through a second messenger phosphatidylinositol (PI)-hydrolysis mechanism.(ABSTRACT TRUNCATED AT 250 WORDS)     

Role of cholinergic and GABAergic systems in the feedback inhibition of dorsal raphe 5-HT neurons

Several observations indicate that 5-HT1A receptors found on a long neuronal feedback loop, originating from the medial prefrontal cortex, regulate 5-HT neuronal firing. In the present study, the muscarinic (M) receptor antagonists atropine and scopolamine as well as the M2 receptor antagonist AF-DX 116, but not the preferential M1 receptor antagonist pirenzepine, reduced the suppressant effect of the 5-HT1A receptor agonist 8-OH-DPAT on the spontaneous firing activity of rat dorsal raphe 5-HT neurons. Moreover, AF-64A-induced lesions of cholinergic neurons directly in the medial prefrontal cortex and after its i.c.v. injection attenuated the effect of 8-OH-DPAT. Finally, the NMDA receptor antagonist (+)MK-801 and the GABA(B) receptor antagonist SCH-50911, but not the GABA(A) receptor antagonist (-)bicuculline, dampened the latter response. The present study unveiled a key role for the cholinergic and GABAergic systems in the feedback inhibition of dorsal raphe 5-HT neurons.     

Selective 5-HT receptor inhibition of glutamatergic and GABAergic synaptic activity in the rat dorsal and median raphe

The dorsal (DR) and median (MR) raphe nuclei contain 5-hydroxytryptamine (5-HT) cell bodies that give rise to the majority of the ascending 5-HT projections to the forebrain. The DR and MR have differential roles in mediating stress, anxiety and depression. Glutamate and GABA activity sculpt putative 5-HT neuronal firing and 5-HT release in a seemingly differential manner in the MR and DR, yet isolated glutamate and GABA activity within the DR and MR has not been systematically characterized. Visualized whole-cell voltage-clamp techniques were used to record excitatory and inhibitory postsynaptic currents (EPSC and IPSC) in 5-HT-containing neurons. There was a regional variation in action potential-dependent (spontaneous) and basal [miniature (m)] glutamate and GABAergic activity. mEPSC activity was greater than mIPSC activity in the DR, whereas in the MR the mIPSC activity was greater. These differences in EPSC and IPSC frequency indicate that glutamatergic and GABAergic input have distinct cytoarchitectures in the DR and MR. 5-HT(1B) receptor activation decreased mEPSC frequency in the DR and the MR, but selectively inhibited mIPSC activity only in the MR. This finding, in concert with its previously described function as an autoreceptor, suggests that 5-HT(1B) receptors influence the ascending 5-HT system through multiple mechanisms. The disparity in organization and integration of glutamatergic and GABAergic input to DR and MR neurons and their regulation by 5-HT(1B) receptors may contribute to the distinction in MR and DR regulation of forebrain regions and their differential function in the aetiology and pharmacological treatment of psychiatric disease states.     

Evidence that both intragastric and subcutaneous administration of methylenedioxymethylamphetamine (MDMA) produce serotonin neurotoxicity in rhesus monkeys

It has been demonstrated that repeated, subcutaneous administration of 3,4-methylenedioxymethamphetamine (MDMA) to rats, guinea pigs, and squirrel monkeys produces long-lasting depletions of serotonin (5-hydroxytryptamine; 5-HT) in several brain regions. Since evidence of degenerating 5-HT neurons has been observed in the rat brain following MDMA injections, it is likely that these depletions are due to neurotoxicity similar to that observed with other substituted amphetamines. The purpose of the present study was to determine if MDMA produces similar evidence of neurotoxicity in rhesus monkeys when administered by either the intragastric (i.g.) or subcutaneous (s.c.) route. Administration of MDMA (5.0 mg/kg/12 h x 4 days) by either i.g. or s.c. routes depleted 5-HT and 5-hydroxyindoleacetic acid (5-HIAA) in various brain regions 2 weeks after the last injection. Further, a significant decrease in [3H]5-HT uptake sites in the hippocampus was observed in monkeys treated with MDMA by the i.g. route. Reductions in uptake sites did not achieve statistical significance when drug was administered s.c. The results suggest that repeated administration of MDMA produces long-lasting, potentially neurotoxic effects on central 5-HT neurons in primates and does so when given orally.     

The serotonergic projection from the median raphe nucleus to the suprachiasmatic nucleus modulates activity phase onset, but not other circadian rhythm parameters

The suprachiasmatic nucleus (SCN) is densely innervated by serotonergic fibers originating in the median raphe nucleus (MR). Serotonin (5-HT) specific lesions of the MR alter entrainment and eliminate 5-HT fibers in the SCN, as well as in all other MR-recipient areas. The present study used 5-HT specific lesions of the SCN or the MR to determine the role of 5-HT in the SCN as a regulator of entrainment. Neurotoxic lesions of the MR significantly reduced 5-HT cell bodies in that nucleus and eliminated essentially all 5-HT innervation of the SCN. As previously demonstrated, these anatomical changes were associated with an advance in activity onset, delay in offset and expansion of the activity phase (alpha). Neurotoxin directly applied to the SCN caused an advance in the average activity onset, but had no effect on offset or alpha. About half of the SCN lesion animals had onsets equivalent to the MR lesion group, whereas onsets of the remaining animals were normal. Loss of SCN 5-HT innervation was severe for all SCN lesion animals, but significantly greater for those with advanced activity onsets. These results suggest that although the 5-HT projection to the SCN is likely to be responsible for modulating activity onset, the timing of activity offset appears to be regulated by a MR projection to an area outside the SCN. Furthermore, surprisingly few 5-HT fibers in the SCN are sufficient to maintain the normal phase angle of entrainment.     

Methylenedioxyamphetamine (MDA) and methylenedioxymethamphetamine (MDMA) cause selective ablation of serotonergic axon terminals in forebrain: immunocytochemical evidence for neurotoxicity

The psychotropic amphetamine derivatives 3,4-methylenedioxyamphetamine (MDA) and 3,4-methylenedioxymethamphetamine (MDMA) have been used for recreational and therapeutic purposes in man. In rats, these drugs cause large reductions in brain levels of serotonin (5-HT). This study employs immunocytochemistry to characterize the neurotoxic effects of these compounds upon monoaminergic neurons in the rat brain. Two weeks after systemic administration of MDA or MDMA (20 mg/kg, s.c., twice daily for 4 d), there is profound loss of serotonergic (5-HT) axons throughout the forebrain; catecholamine axons are completely spared. Regional differences in drug toxicity are exemplified by partial sparing of 5-HT axons in hippocampus, lateral hypothalamus, basal forebrain, and in some areas of neocortex. The terminals of 5-HT axons are selectively ablated, while axons of passage and raphe cell bodies are spared. Thickened preterminal fibers exhibit increased staining due to damming-up of neurotransmitter and other axonal constituents. Fine 5-HT axon terminals are extremely vulnerable to these drugs, whereas terminal-like axons with large varicosities survive, raising the possibility that some 5-HT axons may be resistant to the neurotoxic effects. At short survivals, visualization of greatly swollen, fragmented 5-HT axons provides anatomic evidence for degeneration of 5-HT projections. The results establish that MDA and MDMA produce structural damage to 5-HT axon terminals followed by lasting denervation of the forebrain. Both drugs have similar effects, but MDA produces a greater reduction of 5-HT axons than does MDMA at the same dosage. The selective degeneration of 5-HT axons indicates that these drugs may serve as experimental tools to analyze the organization and function of 5-HT projections. Caution should be exercised until further studies determine whether these compounds may be hazardous in man.     

Evidence for topographically organized endogenous 5-HT-1A receptor-dependent feedback inhibition of the ascending serotonin system

Raphe and extra-raphe 5-HT-1A receptors contribute to feedback inhibition of serotonin (5-HT) neurons; however, the endogenous function of 5-HT-1A receptor-dependent feedback inhibition remains poorly understood. Here, the possibility that 5-HT-1A-mediated feedback inhibition of the raphe nuclei is topographically organized was examined. This was done by testing the effect of systemic blockade of 5-HT-1A receptors on Fos expression in 5-HT neurons in the dorsal raphe (DR) and median raphe (MR). The premise was that appearance of Fos after 5-HT-1A receptor blockade would implicate endogenous inhibition via 5-HT-1A-dependent processes. 5-HT-1A receptor antagonist administration (WAY-100635) in rats returned to their home cage significantly increased the number of Fos-containing 5-HT cells in the lateral wings and the ventral caudal part of the DR as compared to vehicle-injected controls, suggesting that tonic activity of brain 5-HT-1A receptors impacts on these regions. In rats receiving vehicle injections, swim, a behavior known to influence 5-HT neurotransmission, increased the number of Fos-containing 5-HT cells only in the caudal third of DR. Administration of WAY-100635 preceding a swim did not change the amount of Fos in the caudal DR, but increased the number of Fos-containing 5-HT cells in the rostral DR, lateral wings of the DR, and MR. These results confirm, using an imaging approach, that 5-HT-1A receptor-dependent feedback inhibition depends on behavioral state (return to home cage vs. swim). Moreover, they reveal that the effect of 5-HT-1A receptor blockade in each case is subregionally organized.     

Evidence that the medial and dorsal raphe nuclei mediate serotonergically-induced increases in prolactin release from the pituitary

Electrolytic lesions of the medial (MR) or dorsal (DR) raphe nucleus significantly antagonized serum prolactin elevations produced by 5-hydroxytryptophan (5-HTP) in rats pretreated with fluoxetine or citalopram, (serotonin (5-HT) uptake blockers). Lesioned animals in which total blockade of serum prolactin elevations was observed also had total blockade of 5-HT accumulation in the median eminence. However, the increase in serum prolactin levels produced by 5-HTP plus 5-HT reuptake blockade in lesioned rats was not significantly different from sham-operated rats if as little as 15–20% of control median eminence accumulation was present. Serum prolactin elevations produced by quipazine, a direct acting 5-HT agonist, were not significantly affected by MR lesions. On the basis of these results, we suggest that: (1) serum prolactin elevations following 5-HT reuptake blockade plus 5-HTP are correlated with 5-HT concentration in the median eminence; (2) lesion-induced antagonism of 5-HTP-induced prolactin elevation is critically dependent upon complete blockade of median eminence 5-HT accumulation; and (3) 5-HT neurons arising from cell bodies located in the MR and DR are necessary for endogenous serotonergically mediated effects on prolactin secretion in the rat.     

Enhancement of footshock-induced analgesia by spinal 5,7-dihydroxytryptamine lesions

The analgesia induced by 30-s footshock (1 mA) is enhanced and attenuated by decreasing and increasing the extraneuronal availability of serotonin (5-HT), respectively. In the present work we have shown the effect to be mediated by spinal 5-HT systems as the response was increased by depletion of spinal but not brain 5-HT following injection of 5,7-dihydroxytrptamine into the spinal cord or raphe magnus. Injection of 5,7-DHT into the medial raphe which depleted brain but not spinal 5-HT was without effect.     

Inhibition of 3,4-methylenedioxymethamphetamine metabolism leads to marked decrease in 3,4-dihydroxymethamphetamine formation but no change in serotonin neurotoxicity: implications for mechanisms of neurotoxicity

3,4-Methylenedioxymethamphetamine (MDMA)'s O-demethylenated metabolite, 3,4-dihydroxymethamphetamine (HHMA), has been hypothesized to serve as a precursor for the formation of toxic catechol-thioether metabolites (e.g., 5-N-acetylcystein-S-yl-HHMA) that mediate MDMA neurotoxicity. To further test this hypothesis, HHMA formation was blocked with dextromethorphan (DXM), which competitively inhibits cytochrome P450 enzyme-mediated O-demethylenation of MDMA to HHMA. In particular, rats were randomly assigned to one of four treatment groups (n = 9-12 per group): (1) Saline/MDMA; (2) DXM/MDMA; (3) DXM/Saline; (4) Saline/Saline. During drug exposure, time-concentration profiles of MDMA and its metabolites were determined, along with body temperature. One week later, brain serotonin (5-HT) neuronal markers were measured in the same animals. DXM did not significantly alter core temperature in MDMA-treated animals. A large (greater than 70%) decrease in HHMA formation had no effect on the magnitude of MDMA neurotoxicity. These results cast doubt on the role of HHMA-derived catechol-thioether metabolites in the mechanism of MDMA neurotoxicity.     

Median raphe serotonergic innervation of medial septum/diagonal band of broca (MSDB) parvalbumin-containing neurons: possible involvement of the MSDB in the desynchronization of the hippocampal EEG.

Activation of median raphe serotonergic neurons results in the desynchronization of hippocampal electroencephalographic (EEG) activity. This could be a direct effect, because serotonin (5-HT) fibers terminate on a specific population of hippocampal interneurons. On the other hand, it could be an indirect action through the medial septum/diagonal band of Broca (MSDB) pacemaker cells, because, in addition to previously described inhibitory effects, excitatory actions of 5-HT have been demonstrated on MSDB gamma-aminobutyric acid (GABA)-containing neurons through 5-HT2A receptors. Electron microscopic double immunostaining for Phaseolus vulgaris-leucoagglutinin (PHA-L) injected into the median raphe (MR) and parvalbumin, choline acetyltransferase, or calretinin as well as double immunostaining for 5-HT and parvalbumin, and colocalization for parvalbumin and 5-HT2A receptors were done in rats. The results demonstrated that: 1) MR axons form perisomatic and peridendritic baskets and asymmetric synaptic contacts on MSDB parvalbumin neurons; 2) these fibers do not terminate on septal cholinergic and calretinin neurons; 3) 5-HT fibers form synapses identical to those formed by PHA-L-immunolabeled axons with parvalbumin neurons; and 4) MSDB parvalbumin cells contain 5-HT2A receptors. These observations indicate that 5-HT has a dual action on the activity of hippocampal principal cells: 1) an inhibition of the input sector by activation of hippocampal GABA neurons that terminate exclusively on apical dendrites of pyramidal cells, and 2) a disinhibition of the output sector of principal neurons. MSDB parvalbumin-containing GABAergic neurons specifically innervate hippocampal basket and chandelier cells. Thus, 5-HT-elicited activation of MSDB GABAergic neurons will result in a powerful inhibition of these GABA neurons.     

Evidence for a role of energy dysregulation in the MDMA-induced depletion of brain 5-HT

Although the exact mechanism involved in the long-term depletion of brain serotonin (5-HT) produced by substituted amphetamines is not completely known, evidence suggests that oxidative and/or bioenergetic stress may contribute to 3,4-methylenedioxymethamphetamine (MDMA)-induced 5-HT toxicity. In the present study, the effect of supplementing energy substrates was examined on the long-term depletion of striatal 5-HT and dopamine produced by the local perfusion of MDMA (100 microM) and malonate (100 mM) and the depletion of striatal and hippocampal 5-HT concentrations produced by the systemic administration of MDMA (10 mg/kg i.p. x4). The effect of systemic administration of MDMA on ATP levels in the striatum and hippocampus also was examined. Reverse dialysis of MDMA and malonate directly into the striatum resulted in a 55-70% reduction in striatal concentrations of 5-HT and dopamine, and these reductions were significantly attenuated when MDMA and malonate were co-perfused with nicotinamide (1 mM). Perfusion of nicotinamide or ubiquinone (100 microM) also attenuated the depletion of 5-HT in the striatum and hippocampus produced by the systemic administration of MDMA. Finally, the systemic administration of MDMA produced a 30% decrease in the concentration of ATP in the striatum and hippocampus. These results support the conclusion that MDMA produces a dysregulation of energy metabolism which contributes to the mechanism of MDMA-induced 5-HT neurotoxicity.     

Inhibition of lateral vestibular nucleus neurons by 5-hydroxytryptamine derived from the dorsal raphe nucleus

Electrophysiological studies were performed to elucidate the effect of 5-hydroxytryptamine (5-HT) originating in the dorsal raphe nucleus (DR) on neuronal activity in the lateral vestibular nucleus (LVN) neurons, using cats anesthetized with alpha-chloralose. LVN neurons were classified into monosynaptic and polysynaptic neurons according to their responses to vestibular nerve stimulation. Conditioning stimuli applied to the DR inhibited orthodromic spikes elicited by vestibular nerve stimulation predominantly in polysynaptic neurons of the LVN. The iontophoretic application of 5-HT also inhibited orthodromic spikes of the LVN neurons. A close correlation was observed between the effects of DR conditioning stimulation and iontophoretically applied 5-HT in the same neurons. These inhibitions with both treatments were antagonized during the application of methysergide, a 5-HT antagonist. In the majority of LVN polysynaptic neurons that responded to antidromic stimulation of the ipsilateral or contralateral abducens nucleus, orthodromic spikes elicited by vestibular nerve stimulation were inhibited by DR conditioning stimulation and the iontophoretic application of 5-HT. In contrast, LVN neurons that responded to antidromic stimulation of the vestibulospinal tract were rarely affected by these treatments. These results indicate that 5-HT derived from the DR inhibits the synaptic transmission of LVN polysynaptic neurons ascending to the abducens nucleus, and suggest that 5-HT derived from the DR is involved in the regulation of the vestibulo-ocular reflex.     

Investigations of origins of serotonergic projection to developing rat visual cortex: a combined retrograde tracing and immunohistochemical study.

The present study investigated whether the raphe neurons which give rise to the transient serotonergic fibers in the visual cortex of neonatal rats persist or disappear as the rats mature. Three experiments were performed employing the WGA-apoHRP-Au retrograde transport technique in conjunction with 5-HT or WGA-HRP immunohistochemical staining. WGA-apoHRP-Au was injected into the primary visual cortex of all rats 9 days postnatally. In the first experiment, the animals were examined after 2 days; retrogradely labeled cells were observed in the dorsal raphe nucleus (DR), the median raphe nucleus (MR), and in the B9 and B6 cell groups; the majority (82.5%) of the cells was serotonergic. In the second experiment, the examinations took place following a survival time of 8 weeks: virtually all of the original raphe-visual cortical serotonergic neurons were found to the present. In the third experiment, also performed after 8 weeks relabeling the raphe-visual cortical neurons by WGA-HRP, it was found that 37.2% of the raphe neurons which had projected to the neonatal visual cortex no longer possessed such projections.     

Methamphetamine neurotoxicity and striatal glutamate release: comparison to 3,4-methylenedioxymethamphetamine.

The effect of repeated administration of either methamphetamine (MA), 3,4-methylenedioxymethamphetamine (MDMA) or vehicle on the extracellular concentrations of glutamate (GLU), aspartate, taurine, dopamine (DA) and its metabolite, 3,4-dihydroxyphenylacetic acid (DOPAC), was studied in awake, freely moving rats using in vivo microdialysis. MA (7.5 mg/kg, i.p.) administered every 2 h for a total of 3 injections, increased the extracellular concentration of GLU in the anteromedial striatum. By contrast, neither vehicle nor MDMA (9.2 and 13.8 mg/kg) increased GLU efflux following repeated administration. Both MA and MDMA increased the extracellular concentration of DA in the striatum. However, the cumulative increase in DA was significantly greater in the MDMA treated animals as compared to the MA group. The concentrations of DA, serotonin (5-HT) and their metabolites were determined in the striatum 7 days following the repeated administration of MA, MDMA and vehicle. MA, but not MDMA or vehicle, decreased the concentration of DA in the striatum. Conversely, MDMA (13.8 mg/kg) decreased the concentration of 5-HT, whereas MA, MDMA (9.2 mg/kg) and vehicle had no effect on striatal 5-HT content. These data are suggestive that the long-term (7 day) DA neurotoxicity produced by the repeated administration of MA is mediated, in part, by a delayed increase in extracellular concentrations of GLU. In contrast, repeated administration of MDMA, at a dose which produced a long-term (7 day) depletion of striatal 5-HT content, had no effect on GLU efflux in the striatum.     

Influences of the corticotropic axis and sympathetic activity on neurochemical consequences of 3,4-methylenedioxymethamphetamine (MDMA) administration in Fischer 344 rats

The respective influences of the corticotropic axis and sympathetic activity on 3,4-methylenedioxymethamphetamine (MDMA, ecstasy) immediate effects on body temperature and long-term neurotoxicity, as assessed by decreases in hippocampal and striatal [(3)H]5-hydroxytryptamine ([(3)H]5-HT) reuptake, [(3)H]paroxetine binding at 5-HT transporters (5-HTT), and 5-HT and 5-hydroxyindoleacetic acid (5-HIAA) levels, were examined in Fischer 344 rats. On each of the two injections of MDMA (5 or 10 mg/kg s.c. once a day for 2 consecutive days) body temperature rapidly increased in a dose-dependent manner. Six days after the last injection of 10 mg/kg MDMA, [(3)H]5-HT reuptake, [(3)H]paroxetine binding and 5-HT and 5-HIAA levels were decreased in the hippocampus and, to a lower extent, in striatum. Prior adrenalectomy (1 week beforehand), which weakened the immediate hyperthermic effect of MDMA, prevented the long-term MDMA-elicited reduction in hippocampal and striatal [(3)H]paroxetine binding. Supplementation of adrenalectomised Fischer 344 rats with corticosterone almost reinstated the immediate hyperthermic effect of MDMA and restored MDMA-elicited reduction in hippocampal and striatal [(3)H]paroxetine binding. In a final set of experiments, Fischer 344 rats were pretreated (30 min before each of the two injections of 10 mg/kg MDMA) with the ganglionic blocker chlorisondamine (2.5 mg/kg). This pretreatment markedly reduced the amplitudes of the immediate hyperthermia and long-term declines in hippocampal [(3)H]5-HT reuptake and [(3)H]paroxetine binding at 5-HTT, and in hippocampal and striatal 5-HT and 5-HIAA levels. These results suggest that sympathetic activity (possibly through its control of body temperature), but not corticotropic activity, plays a key role in MDMA-elicited neurotoxicity in Fischer 344 rats.