In vivo effects of a new immunosuppressive sigma ligand, SR 31747, on mouse thymus.

SR 31747 is a new sigma ligand which has immunosuppressive properties. The immunopharmacology of SR 31747 was investigated in vivo by studying its effects on the thymuses of C3H mice. The action of SR 31747 was compared with the reference drugs cyclosporin-A and dexamethasone on the basis of several parameters which were: the thymus weight; the number of thymocytes per organ; the percentages of mature CD4+ or CD8+ thymocytes and of immature CD4+ CD8+ thymocytes. SR 31747 slightly but significantly decreased the thymus weight at the dose of 50 mg/kg whereas the number of thymocytes per organ was significantly decreased from 6.25 mg/kg to the 50 mg/kg dose. It had rather no effect on the percentages of immature and mature subsets. These data led to the conclusion that the effects of SR 31747 on the thymuses of C3H mice were close to those obtained with cyclosporin-A and different from those obtained with dexamethasone.

Evidence for modulation of dopamine-neuronal function by tachykinin NK3 receptor stimulation in gerbil mesencephalic cell cultures.

Primary cultures of gerbil mesencephalon were used for studying the modulation exerted by tachykinin NK(3) receptor activation on the activity of dopamine (DA) neurons. Dopamine neurons were identified by their ability to take up [(3)H]DA in a nomifensine-dependent manner. Moreover, tyrosine hydroxylase immunohistochemistry revealed that these neurons accounted for 5-7% of the total cell population. The NK(3) receptor agonists, senktide (EC(50) = 0.58 nM) and [MePhe(7)]neurokinin B (EC(50) = 3 nM), increased spontaneous [(3)H]DA release in a concentration-dependent manner. In contrast, tested at a supramaximal concentration (IC(50) = 0.89 nM), neither septide nor substance P were found to affect [(3)H]DA release. The senktide-evoked [(3)H]DA release was not observed when extracellular Ca(2+) was chelated, but was unaffected by nomifensine. This indicates that this increase in [(3)H]DA outflow resulted more from an exocytotic process than from reversal of carrier-mediated DA uptake. Moreover, the senktide effect was unaffected by the Na+ channel blocker tetrodotoxin, a result suggesting a direct action of senktide on DA neurons. The non-peptide NK(3) receptor antagonist, SR 142801, shifted or blocked (IC(50) = 0.89 nM) the senktide-evoked [(3)H]DA release, while its (-)-antipode, SR 142806, was 80-fold less potent, in agreement with binding data. Selective antagonists for Nk1 (SR 140333) or Nk2 (SR 48968) receptors failed to reduce the senktide effect. Light scanning microscopic analysis of mesencephalic cells loaded with the Ca(2+) sensitive dye, fluo-3, showed that senktide induced a rise in cytosolic Ca(2+) in 8-10% of the cell population. The senktide-induced elevation in intracellular Ca(2+) was rapid in onset and transient (at 10-8 M) or more sustained with no further increase in fluorescence intensity (at 10(-7) M). The proportion of senktide-responsive cells was not significantly modified when extracellular Ca(2+) was chelated, but was reduced by 87% in the presence of SR 142801 and by 75% in cultures that were pre-treated with the DA neurotoxin 1-methyl-4-phenylpyridinium. The present study shows that enhancement of spontaneous [(3)H]DA release and intracellular Ca(2+) mobilization may be observed after NK(3) receptor stimulation and that both biochemical events are likely to occur in DA neurons.

Involvement of peripheral benzodiazepine receptors in the protection of hematopoietic cells against oxygen radical damage.

Several putative functions have been attributed to the peripheral benzodiazepine receptor (PBR), but its precise physiologic role has not been elucidated. In the present study, we investigated PBR function by quantifying this receptor in leukocyte subsets from healthy donors and in leukemic blasts from lymphoid and myeloid lineages. Using a monoclonal antibody (MoAb) directed against the human PBR and a quantitative flow cytometric assay, we found that phagocytic cells from healthy donors displayed a higher level of PBRs than lymphocytes or natural killer (NK) cells. Among the lymphoid lineage, thymocytes and IgD-negative B cells expressed the lowest levels. However, because of the wide heterogeneity of PBR levels among 42 acute or chronic lymphoid and myeloid leukemias, it was not possible to assign PBR expression to a stage of maturation or a cell lineage. Although the PBR displayed a mitochondrial subcellular localization, its expression was not correlated with the mitochondrial content, suggesting a modulation of PBR density at the level of the mitochondria. This modulation was confirmed when we studied in detail the PBR expression during T-cell development by both flow cytometry and confocal microscopy. We found that the PBR was expressed with a bimodal profile during T-cell development, identical to the one observed with the proto-oncogene, Bcl-2. The high similarity in the expression of both the PBR and the Bcl-2 proto-oncogene in T-cell and B-cell subsets, their common mitochondrial localization, and the observation of high quantities of PBR in phagocytic cells, which are known to produce high levels of radical oxygen species, suggested that PBRs may participate in an antioxidant pathway. Indeed, a strong correlation was established between the ability of hematopoietic cell lines to resist H202 cytotoxicity and their level of PBR expression. Demonstration of the role of PBR in the protection against H202 was obtained by transfecting JURKAT cells with the human PBR cDNA. Transfected cells exhibited increased resistance to H202 compared with wild-type cells, suggesting that PBR may prevent mitochondria from radical damages and thereby modulate apoptosis in the hematopoietic system.

The sigma ligand SR 31747 prevents the development of acute graft-versus-host disease in mice by blocking IFN-gamma and GM-CSF mRNA expression.

SR 31747 is a sigma ligand which prevents the development of acute graft-versus-host reaction (GvHR) in hybrid B6D2F1 mice injected with C57BL/6 parental spleen cells. In the present study, we showed that this drug dramatically impaired the GvHR-associated increase in the numbers of both B-lymphocytes and polymorphonuclear cells (PMNs) in the spleen. Because SR 31747 blocked the GvHR-induced expression of both interleukin-2 and transferrin receptors on T-lymphocytes, it is very likely that this molecule prevented the disease through an inhibition of T-lymphocyte activation. Cytokine messenger RNA analyses on spleen cells revealed that SR 31747 blocked IFN-gamma and GM-CSF but not IL-4 transcription. These effects, which are different from those observed with either cyclosporin-A or dexamethasone, strongly suggest that SR 31747 preferentially inhibits the Th1 lymphocyte subset.