title: GABAergic and glutamatergic axons innervate the axon initial segment and organize GABA(A) receptor clusters of cultured hippocampal pyramidal cells.
abstract: We have studied gamma-aminobutyric acid (GABA)(A) receptor (GABA(A)R) clustering within the axon initial segment (AIS) in low-density cultures of hippocampal pyramidal cells following GABAergic and glutamatergic innervation of the AIS. Large, intensely fluorescent, and postsynaptic GABA(A)R clusters were present in the AIS. More than 95% of these clusters colocalized with presynaptic GABAergic or glutamatergic terminals, forming matched or mismatched synapses, respectively. Less than 5% of the GABA(A)R clusters of the AIS did not colocalize with GABAergic or glutamatergic terminals, suggesting that GABA(A)Rs normally do not form clusters unless the AIS received GABAergic or glutamatergic innervation. Few or no clusters of the alpha-amino-3-hydroxy-5-methyl-4-isoxazole-propionate (AMPA) receptors or the postsynaptic density-95 protein (PSD-95) were found in the AIS, even when the AIS was innervated by glutamatergic axons. Glutamatergic innervation of the AIS that formed mismatched synapses with postsynaptic GABA(A)R clusters mainly occurred when the AIS did not receive GABAergic innervation. However, when the AIS was innervated by GABAergic axons, the formation of matched GABAergic synapses predominated and coincided with large reductions in both the density of glutamatergic terminals from the AIS and the mismatching of GABA(A)R clusters. A similar effect was observed at axo-dendritic synapses, where GABAergic innervation also led to a large decrease in mismatched GABA(A)R clusters and a smaller, but significant, decrease in glutamatergic terminal density in dendrites that received GABAergic innervation. We hypothesize that competition between GABAergic and glutamatergic innervation of the AIS in the intact hippocampus leads to the exclusive presence of GABAergic inhibitory synapses in the AIS of pyramidal cells.
authors.affiliation: Department of Physiology and Neurobiology, University of Connecticut, Storrs, Connecticut 06269, USA.
authors.forename: Sean B
authors.initials: SB
authors.lastname: Christie
authors.forename: Angel L
authors.initials: AL
authors.lastname: De Blas
mesh_terms: D000818:Animals; D001369:Axons / Q000378:metabolism*; D002352:Carrier Proteins / Q000378:metabolism; D018929:Cell Culture Techniques; D000076122:Disks Large Homolog 4 Protein; D005455:Fluorescent Antibody Technique; D005968:Glutamate Decarboxylase / Q000378:metabolism; D018698:Glutamic Acid / Q000378:metabolism*; D006624:Hippocampus / Q000166:cytology* / Q000378:metabolism*; D047908:Intracellular Signaling Peptides and Proteins; D008565:Membrane Proteins; D026901:Membrane Transport Proteins; D009419:Nerve Tissue Proteins / Q000378:metabolism; D017729:Presynaptic Terminals; D017966:Pyramidal Cells / Q000378:metabolism*; D051381:Rats; D017207:Rats, Sprague-Dawley; D018091:Receptors, AMPA / Q000378:metabolism; D011963:Receptors, GABA-A / Q000378:metabolism*; D013569:Synapses / Q000378:metabolism; D050598:Vesicular Glutamate Transport Protein 1; D050599:Vesicular Glutamate Transport Protein 2; D033921:Vesicular Transport Proteins; D005680:gamma-Aminobutyric Acid / Q000378:metabolism*
chemical_list: D002352:Carrier Proteins; D000076122:Disks Large Homolog 4 Protein; C501449:Dlg4 protein, rat; D047908:Intracellular Signaling Peptides and Proteins; D008565:Membrane Proteins; D026901:Membrane Transport Proteins; D009419:Nerve Tissue Proteins; D018091:Receptors, AMPA; D011963:Receptors, GABA-A; C493475:Slc17a6 protein, rat; D050598:Vesicular Glutamate Transport Protein 1; D050599:Vesicular Glutamate Transport Protein 2; D033921:Vesicular Transport Proteins; C033652:postsynaptic density proteins; D018698:Glutamic Acid; D005680:gamma-Aminobutyric Acid; D005968:Glutamate Decarboxylase