INTRODUCTION: It is accepted that learning induces plastic changes in adult neocortex. Our previous experiments on mice showed that classical conditioning in which monocular visual stimulation was paired with an electric shock to the tail enhanced GABA immunoreactivity within layer 4 of the monocular part of the primary visual cortex (V1), contralateral to the stimulated eye. AIM(S): In the present study we investigated whether the same classical conditioning paradigm induced changes of neuronal excitability in this cortical area. METHOD(S): We performed patch-clamp whole-cell recordings from ex vivo slices of mouse V1. Two experimental groups were used: mice that had 7-day visual classical conditioning and control animals. The slices were perfused with the modified artificial cerebrospinal fluid, the composition of which better mimics the brain interstitial fluid in situ and induces spontaneous activity in slices. The frequency of spontaneous action potentials was calculated as a general measure of neuronal excitability. RESULTS: We found that layer 4 excitatory cells located in the monocular representation of the “trained” eye in V1 had lower frequency of spontaneous action potentials than neurons from the same cortical region of control animals. CONCLUSIONS: Weaker spontaneous firing indicates decreased general neuronal excitability within layer 4 of the monocular representation of the “trained” eye in V1. Such effect could result from enhanced inhibitory processes in this cortical area. FINANCIAL SUPPORT: Supported by the Polish National Science Center grant Symfonia 1 (2013/08/W/NZ4/00691) and the European Union’s Horizon 2020 research and innovation program under the Marie Sklodowska-Curie grant agreement no 665735 (Bio4Med) and by the funding from Polish Ministry of Science and Higher Education within 2016–2020 funds for the implementation of international projects (agreement no 3548/H2020/COFUND/2016/2).