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International audience; α-actinin-2 (α-actn-2) is an F-actin-crosslinking protein, localized in dendritic spines. In vitro studies suggested that it is involved in spinogenesis, morphogenesis, actin organization, cell migration and anchoring of the NR1 subunit of the N-methyl-D-aspartate (NMDA) receptors in dendritic spines. However, little is known regarding its function in vivo. We examined the levels of α-actn-2 expression within the dentate gyrus (DG) during the development of chronic limbic seizures (epileptogenesis) induced by pilocarpine in rats. In this model, plasticity of the DG glutamatergic granule cells including spine loss, spinogenesis, morphogenesis, neo-synaptogenesis, aberrant migration, and alterations of NMDA receptors have been well characterized. We showed that α-actn-2 immunolabeling was reduced in the inner molecular layer at 1-2 weeks post-status epilepticus (SE), when granule cell spinogenesis and morphogenesis occur. This low level persisted at the chronic stage when new functional synapses are established. This decreased of α-actn-2 protein is concomitant with the recovery of drebrin A (DA), another actin-binding protein, at the chronic stage. Indeed, we demonstrated in cultured cells that in contrast to DA, α-actn-2 did not protect F-actin destabilization and DA inhibited α-actn-2 binding to F-actin. Such alteration could affect the anchoring of NR1 in dendritic spines. Furthermore, we showed that the expression of α-actn-2 and NR1 are co-down-regulated in membrane fractions of pilocarpine animals at chronic stage. Last, we showed that α-actn-2 is expressed in migrating newly born granule cells observed within the hilus of pilocarpine-treated rats. Altogether, our results suggest that α-actn-2 is not critical for the structural integrity and stabilization of granule cell dendritic spines. Instead, its expression is regulated when spinogenesis and morphogenesis occur and within migrating granule cells. Our data also suggest that the balance between α-actn-2 and DA expression levels may modulate NR1 anchoring within dendritic spines.

International audience; Translocator protein (TSPO) and voltage dependent anion channels (VDAC) are two proteins forming a macromolecular complex in the outer mitochondrial membrane that is involved in pleiotropic functions. Specifically, these proteins were described to regulate the clearance of damaged mitochondria by selective mitophagy in non-erythroid immortalized cell lines. Although it is well established that erythroblast maturation in mammals depends on organelle clearance, less is known about mechanisms regulating this clearance throughout terminal erythropoiesis. Here, we studied the effect of TSPO1 downregulation and the action of Ro5-4864, a drug ligand known to bind to the TSPO/VDAC complex interface, in ex vivo human terminal erythropoiesis. We found that both treatments delay mitochondrial clearance, a process associated with reduced levels of the PINK1 protein, which is a key protein triggering canonical mitophagy. We also observed that TSPO1 downregulation blocks erythroblast maturation at the orthochromatic stage, decreases the enucleation rate, and increases cell death. Interestingly, TSPO1 downregulation does not modify reactive oxygen species (ROS) production nor intracellular adenosine triphosphate (ATP) levels. Ro5-4864 treatment recapitulates these phenotypes, strongly suggesting an active role of the TSPO/VDAC complex in selective mitophagy throughout human erythropoiesis. The present study links the function of the TSPO/VDAC complex to the PINK1/Parkin-dependent mitophagy induction during terminal erythropoiesis, leading to the proper completion of erythroid maturation.

International audience; The effects of adrenergic receptor stimulation on spontaneous synaptic transmission were investigated in cultured rat hippocampal neurons by recording spontaneous excitatory and inhibitory postsynaptic currents (sEPSC and sIPSC). Noradrenaline (NA) inhibited sEPSC in a concentration-dependent manner, with maximal effect at 10 microM. The alpha(1)- and alpha(2)-adrenoceptor-selective agonists cirazoline and clonidine induced an inhibition of sEPSC appearance, whereas the beta-adrenoceptor agonist isoproterenol elicited an increase. The inhibitory effect of NA was reversed by alpha(1)-adrenoceptor blockade. The participation of gamma-aminobutyric acid (GABA)(B)-receptor stimulation in the inhibitory effect of NA was further examined. GABA(B)-receptor stimulation with baclofen induced a strong inhibition of bursting activity, which was fully reversed by the GABA(B) antagonist CGP 55845. By itself, CGP 55845 exerted a stimulatory effect on sEPSC frequency. In the presence of CGP 55845, the inhibitory effects of cirazoline and clonidine were maintained. NA (1, 10, and 100 microM) and alpha-adrenoceptor agonists decreased miniature EPSC and IPSC occurrence, whereas beta-adrenergic stimulation increased it. In 50% of the cells examined, NA (1, 10 microM) had a stimulatory effect on sIPSC, whereas, in the remaining 50% of cells, NA (1, 10 microM) had an inhibitory effect. In all the cells, 100 microM NA induced an inhibition of sIPSC. The inhibitory effect of NA was due to alpha(1)-receptor stimulation, whereas the excitatory effect was due to beta-receptor stimulation. In cultured hippocampal neurons, spontaneous excitatory and inhibitory synaptic transmissions are both similarly altered by adrenoceptor stimulation. However, in a subset of cells, low concentrations of NA mediate an increase of sIPSC via beta-adrenoceptor activation.

International audience; In the immature brain, GABA (gamma-aminobutyric acid) is excitatory, and GABA-releasing synapses are formed before glutamatergic contacts in a wide range of species and structures. GABA becomes inhibitory by the delayed expression of a chloride exporter, leading to a negative shift in the reversal potential for choride ions. I propose that this mechanism provides a solution to the problem of how to excite developing neurons to promote growth and synapse formation while avoiding the potentially toxic effects of a mismatch between GABA-mediated inhibition and glutamatergic excitation. As key elements of this cascade are activity dependent, the formation of inhibition adds an element of nurture to the construction of cortical networks.

International audience; During development, when synapses start to be established, a primitive form of network-driven activity provides most of the synaptic activity. This pattern enables a high degree of synchrony in immature neurons in spite of the small number of functional synapses and could participate in activity-dependent growth and synapse formation. Relying on the giant depolarizing potentials that provide most of the synaptic activity in the developing hippocampus, this article reviews the common properties and generating mechanisms of these patterns, and particularly the role of the early depolarizing action of GABA(A) and glycine receptors and the sequential expression of GABA and glutamate synapses. Patterns similar to giant depolarizing potentials have been observed in a wide range of structures and species suggesting that there is a temporal template throughout evolution that constitutes an essential step in the formation of functional networks.

International audience; The sequence of events leading to the chemical matching of presynaptic neurotransmitters and postsynaptic transmitter receptors is investigated here in vivo for the spinal glycine receptor (GlyR) by using immunocytochemical methods. In the ventral horn of adult rat spinal cord, GlyRs are only present at glycinergic postsynaptic differentiations where they are stabilized by the associated protein gephyrin. With quantitative confocal microscopy, we found that gephyrin is detected before GlyRs at embryonic day (E)13-E14 and at E15, respectively, inside the cytoplasm and at plasmalemmal loci. Around the time of birth, the number of cell surface gephyrin-immunoreactive (-IR) spots exceeds that of GlyR. They first match 10 days after birth. The densities of postsynaptic gephyrin- and GlyR-IR were quantified between birth and the adult stage with post-embedding immunogold staining. Immunostaining for gephyrin and GlyR was not detected in the extrasynaptic membrane. The density of staining in postsynaptic membrane increased progressively with development. The inhibitory amino-acid content of the presynaptic terminal boutons opposed to gephyrin-IR sites was also analyzed. In the newborn, postnatal day 10, and adult, more than 90% of these boutons were immunostained for glycine. As seen with serial sections, 38% and 51.2% of the terminals also contained gamma-aminobutyric acid (GABA) in neonate and adult, respectively. These data indicate that around the time of birth, most glycine-containing boutons, some also containing GABA, are opposed to gephyrin-IR postsynaptic densities, whereas GlyRs are not present. Our results suggest that gephyrin determines subsynaptic loci on the plasma membrane where GlyR will subsequently accumulate.