High doses of 3,4-methylenedioxymethamphetamine (MDMA; "Ecstasy") are known to reduce levels of various serotonergic markers outside of the raphe nuclei. To test the hypothesis that these deficits reflect a degeneration of distal axons/terminals, we investigated the effects of an MDMA binge (10mg/kg x 4) on the relative protein and genetic expression of several serotonergic markers in rats, as well as the effects of this compound on the quantity of serotonergic terminals in these animals. In experiment I, we examined whether MDMA alters serotonin transporter (SERT) levels as determined by lysate binding and immunoblotting analyses. Both methods of analysis revealed MDMA-induced reductions in regional SERT content. Experiment II investigated MDMA-induced changes in terminal-specific levels of SERT and the vesicular monoamine transporter 2 (VMAT-2) in the hippocampus, a region with sparse dopaminergic innervation, after lesioning noradrenergic input with N-(2-chloroethyl)-N-ethyl-2-bromobenzylamine (DSP-4). Animals were administered 100 mg/kg DSP-4 or saline 1 week prior to MDMA (or saline). As determined by immunoblotting of synaptosomal tissue, the DSP-4/MDMA group showed little change in hippocampal VMAT-2 protein expression compared to DSP-4/Saline controls, despite large reductions in SERT levels in all regions examined in the MDMA-treated animals. Experiment III examined whether MDMA alters genetic expression of SERT and VMAT-2. When compared to saline-treated controls, animals given MDMA showed a striking decrease in SERT gene expression (and a lesser effect on VMAT-2) in dorsal/median raphe as assessed by quantitative RT-PCR. Experiment IV(a) investigated the effects of MDMA on gene and protein expression of tryptophan hydroxylase (TPH) in the hippocampus. Levels of TPH protein were unchanged between treatment groups, while transcript levels were decreased 15-fold in the dorsal/median raphe. In experiment IV(b), flow cytometry was used to measure whether MDMA alters the quantity of serotonergic terminals in the hippocampus. MDMA-treated animals showed an increase in the number of serotonergic synaptosomes identified by co-labeling for synaptosome-associated protein of 25 kDa (SNAP-25) and TPH. These results demonstrate that MDMA causes substantial regulatory changes in the expression of serotonergic markers with no evidence for synaptic loss, questioning the need to invoke distal axotomy as an explanation of MDMA-related serotonergic deficits.

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<p>Enterochromaffin-like (ECL) and ghrelin cells, in the human gastric mucosa and in gastric endocrine tumours (GETs), were subclassified with respect to immunohistochemical reaction <i>vs.</i> vesicular monoamine transporter 2 (VMAT-2), ghrelin/obestatin, and histidine decarboxylase (HDC). The immunohistochemical expression of ghrelin/obestatin and HDC in GETs was related/correlated to plasma ghrelin/obestatin and urinary methyl imidazole acetic acid (U-MeImAA) excretion respectively, with the intention of identifying markers for these tumour types. </p><p>ECL cells in the gastric mucosa appear either with VMAT-2 only, or with HDC immunoreactivity only, or they can express both proteins; but in GETs the transporter protein and the enzyme were almost always co-expressed in the same cells. Furthermore, ghrelin and obestatin were co-localized in the same cells in the gastric mucosa and in the tumours. In the gastric mucosa, occasional ghrelin/obestatin cells expressed VMAT-2, but in GETs these proteins were always co-localized. Ghrelin expressing cells were non-immunoreactive to HDC. Plasma ghrelin/obestatin concentrations remained low in patients with GETs, irrespective of the relative incidence of these cells in the mucosa and in tumours. The plasma values were not related/correlated to various clinico-pathological parameters. A malignant ghrelinoma was however an exception. The tumour released high total and active ghrelin concentrations into the blood circulation. The patient suffered from diarrhoea, hypothyroidism and diabetes mellitus, but it is not clear if these conditions were due to hyperghrelinaemia. The excretion U-MeImAA was increased in a few patients with GETs, but this increase was not always related to clinical symptoms.</p><p>In conclusion, ECL cells are an heterogeneous group according to VMAT-2 and HDC immunoreactivity. Ghrelin and obestatin are expressed in the same cells in the gastric mucosa, and a few of these cells display VMAT-2 immunoreactivity. Ghrelinoma is a new gastric tumour entity.</p>

Enterochromaffin-like (ECL) and ghrelin cells, in the human gastric mucosa and in gastric endocrine tumours (GETs), were subclassified with respect to immunohistochemical reaction vs. vesicular monoamine transporter 2 (VMAT-2), ghrelin/obestatin, and histidine decarboxylase (HDC). The immunohistochemical expression of ghrelin/obestatin and HDC in GETs was related/correlated to plasma ghrelin/obestatin and urinary methyl imidazole acetic acid (U-MeImAA) excretion respectively, with the intention of identifying markers for these tumour types. ECL cells in the gastric mucosa appear either with VMAT-2 only, or with HDC immunoreactivity only, or they can express both proteins; but in GETs the transporter protein and the enzyme were almost always co-expressed in the same cells. Furthermore, ghrelin and obestatin were co-localized in the same cells in the gastric mucosa and in the tumours. In the gastric mucosa, occasional ghrelin/obestatin cells expressed VMAT-2, but in GETs these proteins were always co-localized. Ghrelin expressing cells were non-immunoreactive to HDC. Plasma ghrelin/obestatin concentrations remained low in patients with GETs, irrespective of the relative incidence of these cells in the mucosa and in tumours. The plasma values were not related/correlated to various clinico-pathological parameters. A malignant ghrelinoma was however an exception. The tumour released high total and active ghrelin concentrations into the blood circulation. The patient suffered from diarrhoea, hypothyroidism and diabetes mellitus, but it is not clear if these conditions were due to hyperghrelinaemia. The excretion U-MeImAA was increased in a few patients with GETs, but this increase was not always related to clinical symptoms. In conclusion, ECL cells are an heterogeneous group according to VMAT-2 and HDC immunoreactivity. Ghrelin and obestatin are expressed in the same cells in the gastric mucosa, and a few of these cells display VMAT-2 immunoreactivity. Ghrelinoma is a new gastric tumour entity.

RCSN-3 cells are a cloned cell line derived from the substantia nigra of an adult rat. The cell line grows in monolayer and does not require differentiation to express catecholaminergic traits, such as (i) tyrosine hydroxylase; (ii) dopamine release; (iii) dopamine transport; (iv) norepinephrine transport; (v) monoamine oxidase (MAO)-A expression, but not MAO-B; (vi) formation of neuromelanin; (vii) vesicular monoamine transporter-2 (VMAT-2) expression. In addition, this cell line expresses serotonin transporters, divalent metal transporter, DMT1, dopamine receptor 1 mRNA under proliferating conditions, and dopamine receptor 5 mRNA after incubation with dopamine or dicoumarol. Expression of dopamine receptors D2, D3 and D4 mRNA were not detected in proliferating cells or when the cells were treated with dopamine, CuSO4, dicoumarol or dopamine-copper complex. Angiotensin II receptor mRNA was also found to be expressed, but it underwent down regulation in the presence of aminochrome. Total quinone reductase activity corresponded 94% to DT-diaphorase. The cells also express antioxidant enzymes such as superoxide dismutase, catalase and glutathione peroxidase. This cell line is a suitable in vitro model for studies of dopamine metabolism, since under proliferating conditions the cells express all the pertinent markers.