KCC2 knockdown impairs glycinergic synapse maturation in cultured spinal cord neurons

Synaptic inhibition in the spinal cord is mediated mainly by strychnine-sensitive glycine (GlyRs) and by gamma-aminobutyric acid type A receptors (GABA(A)R). During neuronal maturation, neonatal GlyRs containing alpha 2 subunits are replaced by adult-type GlyRs harboring alpha 1 and alpha 3 subunits. At the same time period of postnatal development, the transmembrane chloride gradient is changed due to increased expression of the potassium-chloride cotransporter (KCC2), thereby shifting the GABA- and glycine-mediated synaptic currents from mostly excitatory depolarization to inhibitory hyperpolarization. Here, we used RNA interference to suppress KCC2 expression during in vitro maturation of spinal cord neurons. Morphological analysis revealed reduced numbers and size of dendritic GlyR clusters containing alpha 1 subunits but not of clusters harboring neonatal alpha 2 subunits. The morphological changes were accompanied by decreased frequencies and amplitudes of glycinergic miniature inhibitory currents, whereas GABAergic synapses appeared functionally unaltered. Our data indicate that KCC2 exerts specific functions for the maturation of glycinergic synapses in cultured spinal cord neurons.