Distinct alpha subunit subtypes in the molecular assembly of GABA(A) receptors are a critical determinant of the functional properties of inhibitory synapses and their modulation by a range of pharmacological agents. We investigated the contribution of these subunits to the developmental changes of inhibitory synapses in cerebellar granule neurons in primary cultures from wild-type and alpha1 subunit -/- mice. The decay time of miniature inhibitory postsynaptic currents (mIPSCs) halved between 6 days in vitro (DIV6) and DIV12. This was paralleled by the decrease of alpha2 and alpha3 subunits, the increase of alpha1 and alpha6 subunits expression at synapses, and changes in the action of selective alpha subunit modulators. A small but significant shortening of mIPSCs was observed with development in cells from -/- mice together with a decrease in the expression of alpha3 subunit. In contrast, the expression of alpha2 subunit at inhibitory synapses in -/- cells was significantly higher than in +/+ cells at DIV11-12. alpha5 subunit was not detected, and increased sensitivity to a selective alpha4/alpha6 subunit agonist suggests increased expression of extrasynaptic receptors in -/- mice. beta2/beta3 subunit expression and loreclezole sensitivity increased with development in +/+ but not in -/- cells, supporting the preferential association of the alpha1 with the beta2 subunit. Synaptic charge transfer strongly decreased with development but was not different between cells in the +/+ and -/- groups until DIV11-12. Our results uncover a pattern of sequential expression of alpha subunits underlying the changes in functional efficacy of GABAergic networks with development.