Postsynaptic gephyrin clustering controls the development of adult‐born granule cells in the olfactory bulb

Abstract

In adult rodent olfactory bulb, GABAergic signaling regulates migration, differentiation, and synaptic integration of newborn granule cells (GCs), migrating from the subventricular zone. Here we show that these effects depend on the formation of a postsynaptic scaffold organized by gephyrin-the main scaffolding protein of GABAergic synapses, which anchors receptors and signaling molecules to the postsynaptic density-and are regulated by the phosphorylation status of gephyrin. Using lentiviral vectors to selectively transfect adult-born GCs, we observed that overexpression of the phospho-deficient gephyrin mutant eGFP-gephyrin(S270A), which facilitates the formation of supernumerary GABAergic synapses in vitro, favors dendritic branching and the formation of transient GABAergic synapses on spines, identified by the presence of 2-GABAA Rs. In contrast, overexpression of the dominant-negative eGFP-gephyrin(L2B) (a chimera that is enzymatically active but clustering defective), curtailed dendritic growth, spine formation, and long-term survival of GCs, pointing to the essential role of gephyrin cluster formation for its function. We could exclude any gephyrin overexpression artifacts, as GCs infected with eGFP-gephyrin were comparable to those infected with eGFP alone. The opposite effects induced by the two gephyrin mutant constructs indicate that the gephyrin scaffold at GABAergic synapses orchestrates signaling cascades acting on the cytoskeleton to regulate neuronal growth and synapse formation. Specifically, gephyrin phosphorylation emerges as a novel mechanism regulating morphological differentiation and long-term survival of adult-born olfactory bulb neurons. 2 AbstractIn adult rodent olfactory bulb, GABAergic signaling regulates migration, differentiation, and synaptic integration of newborn granule cells (GCs), migrating from the subventricular zone.Here, we show that these effects depend on the formation of a postsynaptic scaffold organized by gephyrin -the main scaffolding protein of GABAergic synapses, which anchors receptors and signaling molecules to the postsynaptic density -and are regulated by the phosphorylation status of gephyrin. Using lentiviral vectors to selectively transfect adult-born GCs, we observed that overexpression of the phospho-deficient gephyrin mutant eGFPgephyrin(S270A), which facilitates the formation of supernumerary GABAergic synapses in vitro, favors dendritic branching and formation of transient GABAergic synapses on spines, identified by the presence of 2-GABA A Rs. In contrast, overexpression of the dominantnegative eGFP-gephyrin(L2B) (a chimera that is enzymatically active but clustering defective), curtailed dendritic growth, spine formation, and long-term survival of GCs, pointing to the essential role of gephyrin cluster formation for its function. We could exclude any gephyrin overexpression artefacts, as GCs infected with eGFP-gephyrin were comparable to those infected with eGFP alone. The opposite effects induced by the two gephyrin mutant constructs indi...

Partial inactivation of GABAA receptors containing the α5 subunit affects the development of adult‐born dentate gyrus granule cells

“…Furthermore, two-way ANOVA analysis revealed no significant effect of time (F 1,11 = 4.79; P = 0.051), indicating that maximal spine density is achieved already at 28 dpi. We did not attempt to distinguish among spine types based on their morphology, in particular because we know from previous work (Deprez et al, 2015) that spine filopodia might not be detectable using GFP-immunofluorescence in immature cells. However, these data are in line with our previous results in Gabra2-and Gabra4-null mice (Duveau et al, 2011) and confirm that spine formation in GCs might be governed by extrinsic influences from the perforant path (Frotscher et al, 2000).…”

The role of calretinin-expressing granule cells in olfactory bulb functions and odor behavior

“…Our results showing that there are fewer gephyrin + puncta on CR + GCs are in line with our electrophysiological results. The density of gephyrin + puncta was lower on primary dendrites, but not on basal dendrites and cell soma, which are also targeted by inhibitory synapses in GCs 40 – 42 . Interestingly, olfactory learning affects the density of gephyrin + puncta on the primary dendrites of adult-born GCs but not on basal dendrites or cell soma 5 .…”

Dopamine depletion induces neuron‐specific alterations of GABAergic transmission in the mouse striatum

“…Adult, 8–9 week‐old, mice were anesthetized using 1.5%–2.5% of the inhalation anesthetic isoflurane (Attane TM ; Piramal Healthcare, India) in oxygen and subsequently head‐fixed into a stereotaxic frame. The stereotaxic injection procedure was performed as described in (Deprez et al., 2015). Animals were injected unilaterally into the right dorsal striatum (anteroposterior [AP] = +0.9 mm, mediolateral [ML] = +1.75 mm, dorsoventral [DV] = −4.3 mm, relative to Bregma) with 6 µg (≙ 400 nl) of 6‐OHDA (6‐OHDA; Tocris Bioscience, Cat.…”