Constitutive endocytosis of GABAA receptors by an association with the adaptin AP2 complex modulates inhibitory synaptic currents in hippocampal neurons

Type A GABA receptors (GABA(A)) mediate the majority of fast synaptic inhibition in the brain and are believed to be predominantly composed of alpha, beta, and gamma subunits. Although changes in cell surface GABA(A) receptor number have been postulated to be of importance in modulating inhibitory synaptic transmission, little is currently known on the mechanism used by neurons to modify surface receptor levels at inhibitory synapses. To address this issue, we have studied the cell surface expression and maintenance of GABA(A) receptors. Here we show that constitutive internalization of GABA(A) receptors in hippocampal neurons and recombinant receptors expressed in A293 cells is mediated by clathrin- dependent endocytosis. Furthermore, we identify an interaction between the GABA(A) receptor beta and gamma subunits with the adaptin complex AP2, which is critical for the recruitment of integral membrane proteins into clathrin-coated pits. GABA(A) receptors also colocalize with AP2 in cultured hippocampal neurons. Finally, blocking clathrin-dependant endocytosis with a peptide that disrupts the association between amphiphysin and dynamin causes a large sustained increase in the amplitude of miniature IPSCs in cultured hippocampal neurons. These results suggest that GABA(A) receptors cycle between the synaptic membrane and intracellular sites, and their association with AP2 followed by recruitment into clathrin- coated pits represents an important mechanism in the postsynaptic modulation of inhibitory synaptic transmission.