Abstract
GABA(A) receptors (GABA(A)-Rs) are localized at both synaptic and extrasynaptic sites, mediating phasic and tonic inhibition, respectively. Previous studies suggest an important role of γ2 and δ subunits in synaptic versus extrasynaptic targeting of GABA(A)-Rs. Here, we demonstrate differential function of α2 and α6 subunits in guiding the localization of GABA(A)-Rs. To study the targeting of specific subtypes of GABA(A)-Rs, we used a molecularly engineered GABAergic synapse model to precisely control the GABA(A)-R subunit composition. We found that in neuron-HEK cell heterosynapses, GABAergic events mediated by α2β3γ2 receptors were very fast (rise time ∼2 ms), whereas events mediated by α6β3δ receptors were very slow (rise time ∼20 ms). Such an order of magnitude difference in rise time could not be attributed to the minute differences in receptor kinetics. Interestingly, synaptic events mediated by α6β3 or α6β3γ2 receptors were significantly slower than those mediated by α2β3 or α2β3γ2 receptors, suggesting a differential role of α subunit in receptor targeting. This was confirmed by differential targeting of the same δ-γ2 chimeric subunits to synaptic or extrasynaptic sites, depending on whether it was co-assembled with the α2 or α6 subunit. In addition, insertion of a gephyrin-binding site into the intracellular domain of α6 and δ subunits brought α6β3δ receptors closer to synaptic sites. Therefore, the α subunits, together with the γ2 and δ subunits, play a critical role in governing synaptic versus extrasynaptic targeting of GABA(A)-Rs, possibly through differential interactions with gephyrin.
Highlights
GABAA receptor ␥2 and ␦ subunits are thought to be responsible for synaptic and extrasynaptic targeting
We employed our recently established hetero-synapse system to investigate the targeting of different subtypes of GABAA-Rs [34]
When Human embryonic kidney (HEK) cells were transfected with GABAA-R subunits and a cell adhesion molecule neuroligin-2 (NL2) and co-cultured with neurons, both spontaneous and action potential-evoked GABAergic events were detected [34]
Summary
GABAA receptor ␥2 and ␦ subunits are thought to be responsible for synaptic and extrasynaptic targeting. Previous studies suggest an important role of ␥2 and ␦ subunits in synaptic versus extrasynaptic targeting of GABAA-Rs. Here, we demonstrate differential function of ␣2 and ␣6 subunits in guiding the localization of GABAA-Rs. To study the targeting of specific subtypes of GABAA-Rs, we used a molecularly engineered GABAergic synapse model to precisely control the GABAA-R subunit composition. Synaptic events mediated by ␣63 or ␣63␥2 receptors were significantly slower than those mediated by ␣23 or ␣23␥2 receptors, suggesting a differential role of ␣ subunit in receptor targeting. This was confirmed by differential targeting of the same ␦-␥2 chimeric subunits to synaptic or extrasynaptic sites, depending on whether it was co-assembled with the ␣2 or ␣6 subunit. The ␣ subunits, together with the ␥2 and ␦ subunits, play a critical role in governing synaptic versus extrasynaptic targeting of GABAA-Rs, possibly through differential interactions with gephyrin
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