ABSTRACT Collybistin (CB) is a guanine nucleotide exchange factor selectively localized to γ‐aminobutyric acid (GABA)ergic and glycinergic postsynapses. Active CB interacts with gephyrin, inducing the submembranous clustering and the postsynaptic accumulation of gephyrin, which is a scaffold protein that recruits GABAA receptors (GABAARs) at the postsynapse. CB is expressed with or without a src homology 3 (SH3) domain. We have previously reported the effects on GABAergic synapses of the acute overexpression of CBSH3− or CBSH3+ in cultured hippocampal (HP) neurons. In the present communication, we are studying the effects on GABAergic synapses after chronic in vivo transgenic expression of CB2SH3− or CB2SH3+ in neurons of the adult rat cerebral cortex. The embryonic precursors of these cortical neurons were in utero electroporated with CBSH3− or CBSH3+ DNAs, migrated to the appropriate cortical layer, and became integrated in cortical circuits. The results show that: 1) the strength of inhibitory synapses in vivo can be enhanced by increasing the expression of CB in neurons; and 2) there are significant differences in the results between in vivo and in culture studies. J. Comp. Neurol. 525:1291–1311, 2017. © 2016 Wiley Periodicals, Inc. Using in utero electroporation, the authors show that transgenic overexpression of collybistin in neurons of the cerebral cortex in the adult rat increases the postsynaptic clustering of gephyrin and GABAA receptors and the strength of GABAergic synapses in vivo.