(30) Production(s) de GUIRAMAND J.
Potentiation of glutamatergic agonist-induced inositol phosphate formation by basic fibroblast growth factor is related to developmental features in hippocampal cultures: neuronal survival and glial cell proliferation. Auteur(s): Guiramand J. (Article) Publié: European Journal Of Neuroscience, vol. 11 p.3377-86 (1999) Ref HAL: hal-00397704_v1 PMID 10564345 Résumé: We investigated the modulation by growth factors of phospholipase C (PLC)-linked glutamate receptors during in vitro development of hippocampal cultures. In defined medium, glial cells represent between 3 and 14% of total cell number. When we added basic fibroblast growth factor (bFGF) 2 h after plating, we found: (i) a neuroprotection from naturally occurring death for up to 5 days; (ii) a proliferation of glial cells from day 3; and (iii) a potentiation of quisqualate (QA)-induced inositol phosphate (IP) formation from 1 to 10 days in vitro (DIV) and 1S, 3R-amino-cyclopentane-1,3-dicarboxylate (ACPD) response from 3 to 10 DIV. The antimitotic cytosine-beta,D-arabinofuranoside (AraC) blocked glial cell proliferation induced by bFGF, but not neuroprotection. Under these conditions, the early potentiation of the QA response (1-3 DIV) was not changed, while the ACPD and late QA response potentiations were prevented (5-10 DIV). Epidermal growth factor was not neuroprotective but it induced both glial cell proliferation and late QA or ACPD potentiation. Surprisingly, the early bFGF-potentiated QA-induced IP response was blocked by 6, 7-dinitro-quinoxaline-2,3-dione (DNQX), suggesting the participation of ionotropic (RS)-alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionate (AMPA)/kainate (KA) receptors. The delayed bFGF-potentiated ACPD-induced IP response is inhibited by (S)-alpha-methyl-4-carboxyphenylglycine (MCPG), indicating possible activation of glial metabotropic receptors. These results suggest that, in hippocampal cultures, bFGF modulates AMPA and metabotropic glutamate receptors linked to the IP cascade, possibly in relation to the regulation of neuronal survival and glial cell proliferation, respectively. |
A modulation of glutamate-induced phosphoinositide breakdown by intracellular pH changes. Auteur(s): Vignes M., Blanc Emmanuelle, Guiramand J., Gonzalez Elsa, Sassetti Isabelle, Récasens Max (Article) Publié: Neuropharmacology, vol. 35 p.1595-604 (1996) Ref HAL: hal-00397711_v1 PMID 9025107 Résumé: The influence of intracellular pH (pHi) changes on the formation of inositol phosphate metabolites (IPs) produced by glutamatergic stimulation was studied in 8-day-old rat brain synaptoneurosomes. For this purpose pHi was measured using 2',7'-bis-(2-carboxyl)-5,6-carboxyfluorescein (BCECF) fluorimetric assay in parallel with the basal and receptor-mediated formations of inositol monophosphate (IP1) and inositol bisphosphate (IP2). We found that glutamate (1 mM), which induces a transient acidification (delta pH = -0.05), produces an identical accumulation of IP1 and IP2. K+ (30 mM), which provokes an alkalinization of the internal medium (delta pH = +0.22), mainly leads to the formation of IP1 metabolites. Paired combinations of glutamate with 1, 5 and 10 mM NH4+ finally result in an alkalinization of the intrasynaptoneurosomal medium. These combinations produce a strong decrease of the IP2 level concomitant with an increase of the IP1 formation, compared to the levels of IP1 and IP2 evoked by glutamate alone. The total amount of IPs (IP1 + IP2) produced by these combinations is not different from that obtained with glutamate alone. Paired combinations of carbachol with NH4+ produce an identical alkalinization to that produced by NH4+ alone. These combinations produce an increased IP1 accumulation, while the IP2 formation is slightly decreased. When the internal medium is acidified by diminishing the external concentration of Na+, the ratio IP1/IP2 produced after metabotropic glutamate receptor (mGluR) activation is shifted to lower values, while it is not affected for the muscarinic stimulation. These data suggest that the mGluR-associated pathway in synaptoneurosomes is sensitive to pHi shifts, while the muscarinic receptor-associated pathway is less altered when pHi is manipulated. It may be proposed that pH-sensitive inositol phosphate dephosphorylating systems, i.e. phosphatases, are associated with mGluRs in this preparation. |
Cadmium rapidly and irreversibly blocks presynaptic phospholipase C-linked metabotropic glutamate receptors. Auteur(s): Vignes M., Blanc Emmanuelle, Davos Frédérique, Guiramand J., Récasens Max (Article) Publié: Neurochemistry International, vol. 29 p.371-81 (1996) Ref HAL: hal-00397709_v1 PMID 8939445 Résumé: Calcium ions (Cd2+) inhibit inositol phosphate (IP) formation elicited by glutamate (GLU) or K+ ions, without affecting carbachol (Carb)-induced IP response in 8-day-old rat forebrain synaptoneurosomes and synaptosomes. On the contrary, Cd2+ was almost ineffective in blocking GLU- and K(+)-responses in hippocampal neurones in culture. The mechanism of Cd2+ inhibition was thus examined in synaptoneurosomes. Extensive washing of synaptoneurosomes pretreated for 1, 5, 15, or 30 min by 100 microM Cd2+ did not modify the inhibitory effect of Cd2+ on GLU-, K(+)- and A23187-evoked IP formation or its lack of effect on Carb response. The later addition of a high affinity Cd2+ chelator (100 microM), N,N,N',N'-tetrakis(2-pyridylmethyl)ethylenediamine (TPEN) also did not reverse the inhibitory effect. TPEN, however, penetrates into synaptoneurosomes and efficiently displaces Cd2+ from the Fura-2-Cd2+ complex as shown by Fura-2 fluorescence recordings. TPEN is not easily removed from the intracellular space, as demonstrated by its ability to still block Cd(2+)-induced Fura-2 fluorescence increase after extensive washing. Pretreatment of synaptoneurosomes by this chelator did not prevent Cd2+ inhibition of GLU-induced IP formation. These data indicate that Cd2+ ions rapidly, irreversibly and extracellularly inhibit GLU-elicited IP formation in synaptoneurosomes or synaptosomes, but not in hippocampal neurones in culture. It is speculated that Cd2+ ions could allow one to distinguish the activity of presynaptic metabotropic glutamate receptors (mGLURs) linked to phosphoinositide metabolism from that of mGLURs located postsynaptically. |
Alternative splicing of the dopamine D2 receptor directs specificity of coupling to G-proteins. Auteur(s): Guiramand J., Montmayeur Jean-Pierre, Ceraline Jocelyn, Bhatia M., Borrelli Emiliana (Article) Publié: The Journal Of Biological Chemistry, vol. 270 p.7354-8 (1995) Ref HAL: hal-00422122_v1 PMID 7706278 Résumé: Two isoforms of the dopamine D2 receptor have been characterized, D2L (long) and D2S (short), generated by alternative splicing from the same gene. They differ by an in-frame insert of 29 amino acids specific to D2L within the putative third intracytoplasmic loop of the receptor. We have previously demonstrated (Montmayeur, J.-P., Guiramand, J., and Borelli, E. (1993) Mol. Endocrinol. 7, 161-170) that D2S and D2L, although presenting very similar pharmacological profiles, couple differently to the alpha-subunit of guanine nucleotide-binding regulatory proteins (G-proteins). In particular, D2L, but not D2S, requires the presence of the alpha-subunit of the inhibitory G-protein (G alpha i2) to elicit greater inhibition of adenylyl cyclase activity. The insert present in D2L must therefore confer the specificity of interaction with G alpha i2. Thus, we introduced substitution mutations within the D2L insert. These mutant receptors were expressed in JEG3 cells, a G alpha i2-deficient cell line, scoring for those presenting an increased inhibition of adenylyl cyclase by dopamine. Our analysis identified two mutants, S259/262A and D249V, with these properties. These results clearly show that the insert present in D2L plays a critical role in the selectivity for the G-proteins interacting with the receptor. |
Preferential coupling between dopamine D2 receptors and G-proteins. Auteur(s): Montmayeur Jean-Pierre, Guiramand J., Borrelli Emiliana (Article) Publié: Molecular Endocrinology -Baltimore-, vol. 7 p.161-70 (1993) Ref HAL: hal-00422125_v1 PMID 7682286 Résumé: The D2 dopamine receptor, an inhibitor of adenylyl cyclase, belongs to the family of seven transmembrane domain G-protein-coupled receptors. This receptor is encoded by two mRNAs produced from the same gene by alternative splicing, here referred to as D2L and D2S. The resultant proteins are identical except for an insertion of 29 amino acids in the putative third intracytoplasmic domain. This domain has been shown to be important for the coupling of this family of receptors to G-proteins. We have previously shown that there is differential inhibition of the adenylyl cyclase activity when these two receptors are produced in JEG3 cells; D2S is more efficient than D2L. To understand the molecular basis of such differential activity, we analyzed the G-proteins expressed in these cells. Here we show that G alpha i2 is absent in this cell line. Moreover, it is possible to restore the same inhibitory activity obtained by D2S when an expression vector encoding this alpha-subunit is cotransfected with D2L. In addition, transfections of the two receptors in a recipient cell line containing the three G alpha i subtypes confirm that the two receptors behave similarly. We conclude that the 29-amino acid insertion present in D2L allows it to interact specifically with G alpha i2. These data suggest that in vivo the function of activated D2 receptors is exerted by specific interactions with Gi-protein subtypes. |
Effect of thiol reagents on phosphoinositide hydrolysis in rat brain synaptoneurosomes. Auteur(s): Vignes M., Guiramand J., Sassetti Isabelle, Récasens Max (Article) Publié: European Journal Of Neuroscience, vol. 5 p.327-34 (1993) Ref HAL: hal-00422123_v1 PMID 8261113 Résumé: Some divalent ions, such as Cd2+ and Zn2+, are able to stimulate phosphoinositide (PI) breakdown and to inhibit receptor-mediated PI metabolism. These ions are also known to react with the free -SH groups of proteins. This prompted us to investigate the effects of more potent sulphydryl reagents, Hg2+ and p-chloromercuric benzosulphonic acid (PCMBS), on the inositol phosphate (IP) accumulation triggered by the neuroactive substances: glutamate, carbachol and K+, using synaptoneurosomes from 8-day-old rat forebrains. Hg2+ and PCMBS, depending on their concentration, had two distinct effects on IP accumulation: at low doses, Hg2+ (from 1 to 10 microM) and PCMBS (0.1 mM) by themselves stimulated PI breakdown, inhibited glutamate-elicited IP accumulation and had additive effects with respect to carbachol-induced IP stimulation. At higher doses, Hg2+ (from 0.01 to 1 mM) inhibited both basal and neuroactive substance-stimulated IP accumulation. PCMBS (1 mM), provoked only an inhibition of the agonist-stimulated IP formation. Monitoring membrane potential and intracellular Ca2+ with the fluorescent dyes diSC2(5) and fura2, respectively, indicated that these mercurials could strongly depolarize the synaptoneurosomal membrane and produce a Ca2+ influx dependent on extracellular Ca2+. The stimulatory effects of low concentrations of mercurials on PI turnover could be linked to the depolarization they provoke and the subsequent Ca2+ rise, which in turn is known to stimulate some phospholipase C enzymes. The inhibitory effects observed at high concentrations might be due to a loss of activity of proteins involved in PI breakdown, as all receptor-mediated IP accumulations were inhibited. |
Binding properties of 3-[125I]iodophencyclidine, a new radioligand for N-methyl-D-aspartate-gated ionic channels. Auteur(s): Chicheportiche Robert, Guiramand J., Kamenka Jean- Marc, Ponchant M., Beaucourt J. P. (Article) Publié: Journal Of Neurochemistry, vol. 59 p.492-9 (1992) Ref HAL: hal-00422127_v1 PMID 1378488 Résumé: The binding properties of the 125I-labeled phencyclidine derivative N-[1-(3-[125I]iodophenyl)cyclohexyl]piperidine (3-[125I]iodo-PCP), a new ligand of the N-methyl-D-aspartate (NMDA)-gated ionic channel, were investigated. Association and dissociation kinetic curves of 3-[125I]iodo-PCP with rat brain homogenates were well described by two components. About 32% of the binding was of fast association and fast dissociation, and the remaining binding was of slow association and slow dissociation. Saturation curves of 3-[125I]iodo-PCP also were well described using two binding sites: one of a high affinity (KDH = 15.8 +/- 2.3 nM) and the other of a low affinity (KDL = 250 +/- 40 nM). 3-Iodo-PCP inhibited the binding of 3-[125I]iodo-PCP with inhibition curves that were well fitted by a two-site model. The binding constants (KiH, BmaxH; KiL, BmaxL) so obtained were close to those obtained in saturation experiments. Ligands of NMDA-gated ionic channels also inhibited the binding of 3-[125I]iodo-PCP with two constants, KiH and KiL. There was a very good correlation (r = 0.987) between the affinities of these ligands to bind to NMDA-gated ionic channels and their potencies to inhibit the binding of 3-[125I]iodo-PCP with a high affinity. Moreover, the regional distribution of the high-affinity binding of 3-[125I]-iodo-PCP paralleled that of tritiated N-[1-(2-thienyl)cyclohexyl]piperidine ([3H]TCP). In contrast to that of [3H] TCP, the binding of 3-[125I]iodo-PCP to well-washed rat brain membranes was fast and insensitive to glutamate and glycine.(ABSTRACT TRUNCATED AT 250 WORDS) |