This study describes cortical recordings in a large animal nerve injury model. We investigated differences in primary somatosensory cortex (S1) hyperexcitability when stimulating injured and uninjured nerves and how different cortical layers contribute to S1 hyperexcitability after spared ulnar nerve injury. We used a multielectrode array to record single-neuron activity in the S1 of ten female Danish landrace pigs. Electrical stimulation of the injured and uninjured nerve evoked brain activity up to 3 h after injury. The peak amplitude and latency of early and late peristimulus time histogram responses were extracted for statistical analysis. Histological investigations determined the layer of the cortex in which each electrode contact was placed. Nerve injury increased the early peak amplitude compared with that of the control group. This difference was significant immediately after nerve injury when the uninjured nerve was stimulated, while it was delayed for the injured nerve. The amplitude of the early peak was increased in layers III–VI after nerve injury compared with the control. In layer III, S1 excitability was also increased compared with preinjury for the early peak. Furthermore, the late peak was significantly larger in layer III than in the other layers in the intervention and control group before and after injury. Thus, the most prominent increase in excitability occurred in layer III, which is responsible for the gain modulation of cortical output through layer V. Therefore, layer III neurons seem to have an important role in altered brain excitability after nerve injury.