title: Epileptiform stimulus increases Homer 1a expression to modulate synapse number and activity in hippocampal cultures.
abstract: Neurons adapt to seizure activity structurally and functionally to attenuate hyperactive neural circuits. Homer proteins provide a scaffold in the postsynaptic density (PSD) by binding to ligands through an EVH1 domain and to other Homer proteins by a coiled-coil domain. The short Homer isoform 1a (H1a) has a ligand-binding domain but lacks a coiled-coil domain and thus acts in a dominant-negative manner to uncouple Homer scaffolds. Here, we show that treating rat hippocampal cultures with bicuculline and 4-aminopyridine (Bic+4-AP) evoked epileptiform activity and synchronized Ca(2+) spiking, measured with whole cell current-clamp and fura-2-based digital imaging; Bic+4-AP increased H1a mRNA through the activation of metabotropic glutamate receptor 5 (mGluR5). Treatment with Bic+4-AP for 4 h attenuated burst firing and induced synapse loss. Synaptic changes were measured using a confocal imaging-based assay that quantified clusters of PSD-95 fused to green fluorescent protein. Treatment with an mGluR5 antagonist blocked H1a expression, synapse loss, and burst attenuation. Overexpression of H1a inhibited burst firing similar to Bic+4-AP treatment. Furthermore, knockdown of H1a using a short hairpin RNA (shRNA) strategy reduced synapse loss and burst attenuation induced by Bic+4-AP treatment. Thus an epileptiform stimulus applied to hippocampal neurons in culture induced burst firing and H1a expression through the activation of mGluR5; a 4-h exposure to this stimulus resulted in synapse loss and burst attenuation. These results suggest that H1a expression functions in a negative-feedback manner to reduce network excitability by regulating the number of synapses.
authors.affiliation: Dept. of Pharmacology, Univ. of Minnesota, 6-120 Jackson Hall, 321 Church St. SE, Minneapolis, MN 55455, USA.
authors.forename: Yan
authors.initials: Y
authors.lastname: Li
authors.forename: Jonathan
authors.initials: J
authors.lastname: Popko
authors.forename: Kelly A
authors.initials: KA
authors.lastname: Krogh
authors.forename: Stanley A
authors.initials: SA
authors.lastname: Thayer
mesh_terms: D015761:4-Aminopyridine / Q000494:pharmacology; D000200:Action Potentials / Q000187:drug effects*; D000818:Animals; D001640:Bicuculline / Q000494:pharmacology*; D002118:Calcium / Q000378:metabolism; D020013:Calcium Signaling / Q000187:drug effects; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism*; D003292:Convulsants / Q000494:pharmacology*; D000076122:Disks Large Homolog 4 Protein; D025461:Feedback, Physiological; D006624:Hippocampus / Q000166:cytology / Q000378:metabolism / Q000502:physiology; D000072233:Homer Scaffolding Proteins; D047908:Intracellular Signaling Peptides and Proteins / Q000235:genetics / Q000378:metabolism; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism; D009474:Neurons / Q000166:cytology / Q000378:metabolism / Q000502:physiology*; D026902:Potassium Channel Blockers / Q000494:pharmacology; D020033:Protein Isoforms / Q000235:genetics / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D034741:RNA, Small Interfering; D051381:Rats; D064529:Receptor, Metabotropic Glutamate 5; D018094:Receptors, Metabotropic Glutamate / Q000235:genetics / Q000378:metabolism; D013569:Synapses / Q000187:drug effects*; D014158:Transcription, Genetic / Q000187:drug effects
chemical_list: D002352:Carrier Proteins; D003292:Convulsants; D000076122:Disks Large Homolog 4 Protein; C501449:Dlg4 protein, rat; C578937:Grm5 protein, rat; D000072233:Homer Scaffolding Proteins; D047908:Intracellular Signaling Peptides and Proteins; D008565:Membrane Proteins; D026902:Potassium Channel Blockers; D020033:Protein Isoforms; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D064529:Receptor, Metabotropic Glutamate 5; D018094:Receptors, Metabotropic Glutamate; D015761:4-Aminopyridine; D002118:Calcium; D001640:Bicuculline