title: Gephyrin regulates GABAergic and glutamatergic synaptic transmission in hippocampal cell cultures.
abstract: Gephyrin is a scaffold protein essential for stabilizing glycine and GABA(A) receptors at inhibitory synapses. Here, recombinant intrabodies against gephyrin (scFv-gephyrin) were used to assess whether this protein exerts a transynaptic action on GABA and glutamate release. Pair recordings from interconnected hippocampal cells in culture revealed a reduced probability of GABA release in scFv-gephyrin-transfected neurons compared with controls. This effect was associated with a significant decrease in VGAT, the vesicular GABA transporter, and in neuroligin 2 (NLG2), a protein that, interacting with neurexins, ensures the cross-talk between the post- and presynaptic sites. Interestingly, hampering gephyrin function also produced a significant reduction in VGLUT, the vesicular glutamate transporter, an effect accompanied by a significant decrease in frequency of miniature excitatory postsynaptic currents. Overexpressing NLG2 in gephyrin-deprived neurons rescued GABAergic but not glutamatergic innervation, suggesting that the observed changes in the latter were not due to a homeostatic compensatory mechanism. Pulldown experiments demonstrated that gephyrin interacts not only with NLG2 but also with NLG1, the isoform enriched at excitatory synapses. These results suggest a key role of gephyrin in regulating transynaptic signaling at both inhibitory and excitatory synapses.
authors.affiliation: Neurobiology Department and Italian Institute of Technology Unit, International School for Advanced Studies (SISSA), Via Bonomea 265, 34136 Trieste, Italy.
authors.forename: Zeynep Kasap
authors.initials: ZK
authors.lastname: Varley
authors.forename: Rocco
authors.initials: R
authors.lastname: Pizzarelli
authors.forename: Roberta
authors.initials: R
authors.lastname: Antonelli
authors.forename: Stefka H
authors.initials: SH
authors.lastname: Stancheva
authors.forename: Matthias
authors.initials: M
authors.lastname: Kneussel
authors.forename: Enrico
authors.initials: E
authors.lastname: Cherubini
authors.forename: Paola
authors.initials: P
authors.lastname: Zacchi
mesh_terms: D000818:Animals; D002352:Carrier Proteins / Q000096:biosynthesis* / Q000235:genetics; D015816:Cell Adhesion Molecules, Neuronal / Q000235:genetics / Q000378:metabolism; D057809:HEK293 Cells; D006624:Hippocampus / Q000166:cytology / Q000378:metabolism*; D006801:Humans; D008565:Membrane Proteins / Q000096:biosynthesis* / Q000235:genetics; D009419:Nerve Tissue Proteins / Q000235:genetics / Q000378:metabolism; D017729:Presynaptic Terminals / Q000378:metabolism*; D051381:Rats; D017208:Rats, Wistar; D011963:Receptors, GABA-A / Q000235:genetics / Q000378:metabolism; D018009:Receptors, Glycine / Q000235:genetics / Q000378:metabolism; D009435:Synaptic Transmission / Q000502:physiology*; D050495:Vesicular Inhibitory Amino Acid Transport Proteins / Q000235:genetics / Q000378:metabolism; D005680:gamma-Aminobutyric Acid / Q000235:genetics / Q000378:metabolism
chemical_list: D002352:Carrier Proteins; D015816:Cell Adhesion Molecules, Neuronal; D008565:Membrane Proteins; D009419:Nerve Tissue Proteins; D011963:Receptors, GABA-A; D018009:Receptors, Glycine; C118623:Slc32a1 protein, rat; D050495:Vesicular Inhibitory Amino Acid Transport Proteins; C073441:gephyrin; C093438:neuroligin 1; C098003:neuroligin 2; D005680:gamma-Aminobutyric Acid