[Show abstract][Hide abstract] ABSTRACT: Accumulating evidence show that chemokines can modulate the activity of neurons through various mechanisms. Recently, we demonstrated that CCR2, the main receptor for the chemokine CCL2, is constitutively expressed in dopamine neurons in the rat substantia nigra. Here we show that unilateral intranigral injections of CCL2 (50 ng) in freely moving rats increase extracellular concentrations of dopamine and its metabolites and decrease dopamine content in the ipsilateral dorsal striatum. Furthermore, these CCL2 injections are responsible for an increase in locomotor activity resulting in contralateral circling behavior. Using patch-clamp recordings of dopaminergic neurons in slices of the rat substantia nigra, we observed that a prolonged exposure (>8 min) to 10 nM CCL2 significantly increases the membrane resistance of dopaminergic neurons by closure of background channels mainly selective to potassium ions. This leads to an enhancement of dopaminergic neuron discharge in pacemaker or burst mode necessary for dopamine release. We provide here the first evidence that application of CCL2 on dopaminergic neurons increases their excitability, dopamine release and related locomotor activity.
[Show abstract][Hide abstract] ABSTRACT: Dopaminergic neurons of the substantia nigra constitutively express the CXCR4 receptor for the chemokine stromal-cell-derived factor 1alpha (CXCL12) but, to date, no direct effect of CXCR4 activation by CXCL12 on membrane conductance of dopaminergic neurons has been demonstrated. We tested the effects of CXCL12 on whole-cell currents of dopaminergic neurons recorded in patch clamp in substantia nigra slices and showed that CXCL12 (0.01-10 nm) increased the amplitude of total high-voltage-activated (HVA) Ca currents through CXCR4 activation. This effect was reversibly reduced by varpi-conotoxin-GVIA, suggesting that CXCL12 acted on N-type Ca currents, known to be involved in dopamine (DA) release. We therefore investigated the effects of CXCL12 on DA release from cultured dopaminergic neurons from the rat mesencephalon. In basal conditions, CXCL12 alone had no effect on DA release. When neurons were depolarized with KCl (20 mm), and thus when HVA Ca currents were activated, low CXCL12 concentrations (1-50 nm) increased DA release via CXCR4 stimulation. These data strongly suggest that the chemokine CXCL12 can act directly as a neuromodulator of dopaminergic neuronal electrical activity through the modulation of HVA currents.
European Journal of Neuroscience 08/2008; 28(5):862-70. · 3.67 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: We recently demonstrated that dopaminergic (DA) neurons of the rat substantia nigra constitutively expressed CXCR4, receptor for the chemokine stromal cell-derived factor-1 (SDF-1)/CXCL12 (SDF-1). To check the physiological relevance of such anatomical observation, in vitro and in vivo approaches were used. Patch clamp recording of DA neurons in rat substantia nigra slices revealed that SDF-1 (10 nmol/L) induced: (i) a depolarization and increased action potential frequency; and (ii) switched the firing pattern of depolarized DA neurons from a tonic to a burst firing mode. This suggests that SDF-1 could increase DA release from neurons. Consistent with this hypothesis, unilateral intranigral injection of SDF-1 (50 ng) in freely moving rat decreased DA content and increased extracellular concentrations of DA and metabolites in the ipsilateral dorsal striatum, as shown using microdialysis. Furthermore, intranigral SDF-1 injection induced a contralateral circling behavior. These effects of SDF-1 were mediated via CXCR4 as they were abrogated by administration of a selective CXCR4 antagonist. Altogether, these data demonstrate that SDF-1, via CXCR4, activates nigrostriatal DA transmission. They show that the central functions of chemokines are not restricted, as originally thought, to neuroinflammation, but extend to neuromodulatory actions on well-defined neuronal circuits in non-pathological conditions.
Journal of Neurochemistry 09/2007; 102(4):1175-83. · 4.24 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: In rat substantia nigra (SN), Chemokine (CXC motif) receptor 4 (CXCR4) for the chemokine stromal cell-derived factor (SDF)-1alpha is expressed on dopaminergic (DA) neurones, but also on non-DA cells, suggesting presynaptic actions. Using whole-cell patch-clamp recordings in DA neurones of rat SN slices at a holding potential of -60 mV, we showed here that SDF-1alpha exerts multiple presynaptic effects. First, SDF-1alpha (10 nm) induced an increase in the frequency of spontaneous and miniature GABA(A) postsynaptic currents by presynaptic mechanisms, consistent with the presence of CXCR4 on GABAergic neurones of the SN, as revealed by immunocytochemistry. Second, SDF-1alpha (0.1-1 nm) induced a glutamatergic inward current resistant to tetrodotoxin (TTX), most probably the result of glutamate release from non-neuronal cells. This inward current was not blocked by the CXCR4 antagonist AMD 3100 (1 microm), consistent with the lack of CXCR4 on astrocytes as shown by immunocytochemistry under basal conditions. Finally, SDF-1alpha (10 nm) induced, via CXCR4, an outward G protein-activated inward rectifier (GIRK) current, which was TTX sensitive and prevented by application of the GABA(B) antagonist CGP55845A, suggesting GABA spillover on to GABA(B) receptors. Our results show that SDF-1alpha induces, via presynaptic mechanisms, alterations in the excitability of DA neurones as confirmed by current-clamp experiments.
Journal of Neurochemistry 04/2006; 96(6):1540-50. · 4.24 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The cyclo-oxygenase (COX) and lipoxygenase (LOX) pathways belong to the eicosanoid synthesis pathway, a major component of the chronic inflammatory process occurring in Alzheimer's disease (AD). Clinical studies reported beneficial effects of COX inhibitors, but little is known about the involvement of LOXs in AD pathogenesis. β-amyloid peptide (Aβ) accumulation contributes to neurodegeneration in AD, but mechanisms underlying Aβ toxicity have not been fully elucidated yet. Here, using an antisense oligonucleotide-based strategy, we show that blockade of 12-LOX expression prevents both Aβ-induced apoptosis and overexpression of c-Jun, a factor required for the apoptotic process, in cortical neurons. Conversely, the 12-LOX metabolite, 12(S)-HETE (12(S)-hydroxy-(5Z, 8Z, 10E, 14Z)-eicosatetraenoic acid), promoted c-Jun-dependent apoptosis. Specificity of the 12-LOX involvement was further supported by the observed lack of contribution of 5-LOX in this process. These data indicate that blockade of 12-LOX expression disrupts a c-Jun-dependent apoptosis pathway, and suggest that 12-LOX may represent a new target for the treatment of AD.
Cell Death and Differentiation 04/2004; 11(8):875-884. · 8.39 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Recent studies demonstrated that the chemokine monocyte chemoattractant protein-1 (MCP-1)/CCL2 and its receptor, CCR2, play important roles in various brain diseases. In this study, using quantitative autoradiography, we studied the pharmacological properties of [125l]MCP-1/CCL2 binding in rat brain and we clearly showed the distribution of CCR2 receptors in cerebral cortex, nucleus accumbens, striatum, amygdala, thalamus, hypothalamus, hippocampus, substantia nigra, mammillary bodies and raphe nuclei. Moreover, using double fluorescent immunohistochemistry, we showed that CCR2 receptors were constitutively expressed on neurons and astrocytes. Using RT-PCR methods, we demonstrated that CCR2 mRNA is present in various brain areas described above. Four hours after an acute intraperitoneal lipopolysaccharide injection, we showed that MCP-1/CCL2 binding was up-regulated in several brain structures; this effect took place on both CCR2B labelled neurons and astrocytes and to a lesser extent on activated microglia. To explore neurobiological function of CCR2, actimetric study was carried out. After intracerebroventricular injections of MCP-1/CCL2, we showed that motor activity was markedly decreased. Our results provide the first evidence for constitutive CCR2 receptor expression with precise neuroanatomical and cellular localizations in the brain, and its regulation during an inflammatory process, suggesting that MCP-1/CCL2 and CCR2 play important physiological and pathophysiological role(s) in the CNS.
Journal of Neurochemistry 05/2002; 81(2):257-69. · 4.24 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: We have explored the role of endogenous dopamine in the control of histaminergic neuron activity in mouse brain regions evaluated by changes in tele-methylhistamine (t-MeHA) levels. In vitro, methamphetamine released [(3)H]noradrenaline but failed to release [(3)H]histamine from synaptosomes. In vivo, methamphetamine enhanced t-MeHA levels by about 2-fold with ED(50) values of approximately 1 mg/kg in caudate putamen, nucleus accumbens, cerebral cortex, and hypothalamus. This response selectively involved the D(2) and not the D(3) receptor as indicated by its blockade by haloperidol and by its persistence after administration of nafadotride, a D(3) receptor preferential ligand, or in (-/-) D(3) receptor-deficient mice. The t-MeHA response to methamphetamine was delayed compared with the locomotor-activating effect of this drug, suggesting that it is of compensatory nature. In agreement, ciproxifan, an inverse agonist known to enhance histamine neuron activity, decreased the hyperlocomotion induced by methamphetamine. Repeated methamphetamine administration resulted in the expected sensitization to the hyperlocomotor effect of the drug but did not modify either the ED(50) or the E(max) regarding t-MeHA levels. However, it resulted in an enhanced basal t-MeHA level (+30-40%), which was sustained for at least 11 days after withdrawal in hypothalamus, striatum, and cerebral cortex and suppressed by haloperidol. Hence, both the acute and chronic administration of methamphetamine enhance histamine neuron activity, presumably in a compensatory manner. Repeated methamphetamine administration also resulted in a modified balance in the opposite influences of dopamine and serotonin on histaminergic neurons as revealed by the enhanced response to haloperidol and abolished response to ketanserin, respectively.
Journal of Pharmacology and Experimental Therapeutics 03/2002; 300(2):621-8. · 3.86 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The human neuroblastoma cell line CHP212 was found to express functional high affinity neurotensin (NTS-1) receptor subtype. Based on the functional interactions between neurotensin and dopamine transmission, we have used this cell line to investigate the short- and long-term modulation of tyrosine hydroxylase gene expression by the stable neurotensin agonist JMV 449. After exposure of the cells to 1 microM JMV 449 for 5 or 72 h, tyrosine hydroxylase protein and mRNA levels were significantly increased as detected by western blot analysis and quantitative RT-PCR, respectively. Transfection of CHP212 cells with a plasmid containing the luciferase reporter gene under the control of a limited proximal region of the cloned tyrosine hydroxylase promoter, revealed that the effect of JMV 449 results from an increase in the transcriptional activity of the TH gene. These results indicate that modulation of tyrosine hydroxylase gene expression may constitute one of the mechanisms involved in the control of dopamine transmission by neurotensin. Such neurotensin-mediated changes in tyrosine hydroxylase expression may also participate in multiple adaptation processes within the central nervous system to environmental conditions where neurotensin is released such as stress and food intake.
[Show abstract][Hide abstract] ABSTRACT: Accumulation of amyloid beta peptide (Abeta) has been suggested to contribute to neurodegeneration in Alzheimer's disease (AD). Since chronic inflammation occurs in AD pathogenesis and lipoxygenases are important mediators of inflammatory processes, we evaluated the effect of lipoxygenase inhibitors on apoptosis induced by Abeta on rat cortical cells. The 12-lipoxygenase inhibitor baicalein attenuated both neuronal apoptosis and c-jun protein over-expression induced by Abeta(25- 35), whereas no protection was found with the broad spectrum lipoxygenase inhibitor nordihydroguaiaretic acid or the 5-lipoxygenase inhibitor caffeic acid. These results suggest that 12-lipoxygenase participates in a c-jun-dependent apoptosis pathway triggered by Abeta(25-35), and that specific 12-lipoxygenase inhibitors might be of interest in AD.
[Show abstract][Hide abstract] ABSTRACT: Neurotensin is present in selective mesolimbic dopaminergic projections to the nucleus accumbens (NAc) shell but also is synthesized locally in this region and in the motor-associated NAc core. We examined the electron microscopic immunolabeling of the high-affinity neurotensin receptor (NTR) and neurotensin in these subdivisions of rat NAc to determine the sites for receptor activation and potential regional differences in distribution. Throughout the NAc, NTR immunoreactivity was localized discretely within both neurons and glia. NTR-labeled neuronal profiles were mainly axons and axon terminals with diverse synaptic structures, which resembled dopaminergic and glutamatergic afferents, as well as collaterals of inhibitory projection neurons. These terminals had a significantly higher numerical density in the NAc core than in the shell but were prevalent in both regions, suggesting involvement in both motor and limbic functions. In each region, neurotensin was detected in a few NTR-immunoreactive axon terminals and in terminals that formed symmetric, inhibitory type synapses with NTR-labeled somata and dendrites. The NTR labeling, however, was not seen within these synapses and, instead, was localized to segments of dendritic and glial plasma membranes often near excitatory type synapses. Neuronal NTR immunoreactivity also was associated with cytoplasmic tubulovesicles and nuclear membranes. Our results suggests that, in the NAc shell and core, NTR is targeted mainly to presynaptic sites, playing a role in the regulated secretion and/or retrograde signaling in diverse, neurotransmitter-specific neurons. The findings also support a volume mode of neurotensin actions, specifically affecting excitatory transmission through activation of not only axonal but also dendritic and glial NTR.
The Journal of Comparative Neurology 07/2001; 435(2):142-55. · 3.51 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Plasma neurotensin (NT) was measured by radioimmunoassay in propanol extracted and unextracted plasma from 16 parkinsonian patients (four before treatment) and 16 age and sex matched controls. Mean plasma NT concentrations were consistently higher in parkinsonian patients than in controls and higher in the four untreated patients than in levodopa treated patients suggesting that plasma NT measurement may represent an easy detectable additional index in diagnosing parkinsonism and provides a novel approach to research in this field.
[Show abstract][Hide abstract] ABSTRACT: Much evidence suggests that apoptosis plays a crucial role in cell population homeostasis that depends on the expression of various genes implicated in the control of cell life and death. The sensitivity of human neuroblastoma cells SK-N-SH to undergo apoptosis induced by thapsigargin was examined. SK-N-SH were previously differentiated into neuronal cells by treatments with retinoic acid (RA), 4 beta-phorbol 12-myristate 13-acetate (PMA) which increases protein kinase C (PKC) activity, and staurosporine which decreases PKC activity. Neuronal differentiation was evaluated by gamma-enolase, microtubule associated protein 2 (MAP2) and synaptophysin immunocytochemistry. The sensitivity of the cells to thapsigargin-induced apoptosis was evaluated by cell viability and nuclear fragmentation (Hoechst 33258) and compared with pro-(Bcl-2, Bcl-x(L)) and anti-apoptotic (Bax, Bak) protein expression of the Bcl-2 family. Cells treated with RA and PMA were more resistant to apoptosis than controls. Conversely, the cells treated with staurosporine were more susceptible to apoptosis. In parallel with morphological modifications, the expression of inhibitors and activators of apoptosis was directly dependent upon the differentiating agent used. Bcl-2 expression was strongly increased by PMA and drastically decreased by staurosporine as was Bcl-x(L) expression. Bax and Bak expression were not significantly modified. These results demonstrate that drugs that modulate PKC activity may induce a modification of Bcl-2 expression as well as resistance to the apoptotic process. Furthermore, the expression of Bcl-2 was reduced by toxin B from Clostridium difficile and, to a lesser extent, by wortmannin suggesting a role of small G-protein RhoA and PtdIns3 kinase in the control of Bcl-2 expression. Our data demonstrate a relationship between the continuous activation of PKC, the expression of Bcl-2 protein family and the resistance of differentiated SK-N-SH to apoptosis.
European Journal of Biochemistry 04/2001; 268(5):1352-62. · 3.58 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Exposure to psychostimulants and antipsychotics increases neurotensin (NT) gene expression in the striatum and nucleus accumbens. To investigate the contribution of D(3) receptors to these effects we used mice with targeted disruption of the D(3) receptor gene. Basal NT mRNA expression was similar in D(3) receptor mutant mice and wild-type animals. Acute administration of haloperidol increased NT gene expression in the striatum in D(3)+/+, D(3)+/- and D(3)-/- mice. Similarly, acute cocaine and amphetamine induced NT mRNA expression in the nucleus accumbens shell and olfactory tubercle to a comparable extent in D(3) mutants and wild-type mice. Daily injection of cocaine for seven days increased NT mRNA in a restricted population of neurons in the dorsomedial caudal striatum of D(3)+/+ mice, but not in D(3)-/- and D(3)+/- animals. No differences were observed between D(3) receptor mutant mice and wild-type littermates in the locomotor activity and stereotyped behaviors induced by repeated cocaine administration. These findings demonstrate that dopamine D(3) receptors are not necessary for the acute NT mRNA response to drugs of abuse and antipsychotics but appear to play a role in the regulation of NT gene induction in striatal neurons after repeated cocaine. In addition, our results indicate that the acute locomotor response to cocaine and development of psychostimulant-induced behavioral sensitization do not require functional D(3) receptors.
[Show abstract][Hide abstract] ABSTRACT: To investigate whether human corneal epithelial cells express the glucocorticoid receptor (GR) and to assess the influence of dexamethasone (DEX) on these cells.
Human corneal epithelial cells were cultured in medium supplemented with various concentrations of DEX (ranging from 10(-10) to 10(-4) M). Cell proliferation was analyzed by 3-(4, 5-dimethylthiazol-2-yl)-5-(3-carboxy-methoxyphenyl)-2-(4-sulfop henyl) -2H-tetrazolium inner salt (MTS) assay at 2, 4, and 6 days of culture. Apoptosis was studied by nucleus labeling using a fluorescent dye and immunostaining by APO 2.7 at 6 days of culture. GR mRNA was detected in corneal epithelium and cultured corneal epithelial cells by means of reverse transcription-polymerase chain reaction (RT-PCR). Immunocytochemical staining of the epithelial cells was performed with a monoclonal anti-human GR.
RT-PCR and immunocytochemistry showed the expression of GR (mRNA and protein) in corneal epithelial cells. DEX significantly increased corneal epithelial cell proliferation with concentrations ranging from 10(-10) to 10(-6) M, with a maximum effect at 10(-7) M (P < 0.005). However, DEX also induced apoptosis of cultured corneal epithelial cells at any concentration used.
These results indicate that human corneal epithelial cells express the GR and proliferate in response to DEX stimulation which also induces corneal epithelial cell apoptosis.
[Show abstract][Hide abstract] ABSTRACT: Tibolone and its main derivatives were studied in an original model of cultures of normal human epithelial breast (HBE) cells on proliferation, differentiation and apoptosis, the three mechanisms responsible for breast homeostasis. Tibolone and its Delta4 isomer were antiproliferative, both in the absence and presence of oestradiol (E2). The oestrogenic 3alpha and 3beta hydroxy derivatives did not display any mitogenic activities in HBE cells. Moreover, at 1 microM, they were antiproliferative. Tibolone and its Delta isomer increased the 17beta hydroxysteroid dehydrogenase activity similarly to that observed with progestins . Apoptosis was increased in HBE cells to a similar range as with the pure pregnane progestin, Org2058. We also studied the extent of apoptosis in hormone-dependent breast cancer cell lines. Tibolone and its Delta4 isomer also increased apoptosis, especially in ZR75-1 cells containing progesterone and androgen receptors . We could demonstrate that these pro-apoptotic actions of tibolone and its Delta4 isomer were mediated at least partially through the bcl-2-family of proteins. Moreover, the antiproliferative and pro-apoptotic activities of tibolone, as well as Org2058, were mediated by increasing catalase activities in breast cancer cells. Thus, in breast cells, tibolone slows down the proliferation rate, increases differentiation and apoptosis. These actions seem to be optimal on breast tissue.
European Journal of Cancer 10/2000; 36 Suppl 4:S76-7. · 4.82 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Radioimmunotherapy recently afforded convincing results for B-cell non-Hodgkin's lymphoma treatment with antibody specific for B-cell differentiation antigens. High doses of unlabeled or labeled antibodies are necessary to saturate specific sites on normal B-cells. We thus developed a new targeting strategy, taking advantage of dual binding cooperativity, to enhance the specificity of the radioactive uptake by tumor cells. This approach was evaluated using human Burkitt lymphoma cells (Ramos) which express both CD10 and CD20 antigens. Most normal cells express at most one of these two differentiation antigens but many hematological tumors, including most human B type acute lymphoblastic leukemia cells, express both. Cells pretargeted with two bispecific antibodies, one recognizing CD10 and a histamine derivative (HSG), the other recognizing CD20 and the DTPA-indium complex, bind cooperatively radiolabeled mixed-haptens (DTPA-HSG). Increased binding (about 5-fold compared to binding to only one of CD10 or CD20 antigens) is observed at 37 degrees C, demonstrating the feasibility of the technique. This binding enhancement is a slow process, not observed at 4 degrees C. Such a binding enhancement will increase specificity for targeting isotopes to double antigen positive tumor cells compared to nontumor tissue cells bearing only one of them. This approach might be used to increase tumor irradiation with minimal irradiation of normal cells.