The deeper part of neocortical layer VI is dominated by non-pyramidal neurons, which lack a prominent vertically ascending dendrite, and predominantly establish cortico-cortical connections. These neurons were studied in rat neocortical slices using patch-clamp, single cell reverse transcription-polymerase chain reaction and biocytin labeling. The majority of these neurons expressed the vesicular glutamate transporter but not glutamic acid decarboxylase, suggesting that a high proportion of layer VI non-pyramidal neurons are glutamatergic. Indeed, they exhibited numerous dendritic spines, and established asymmetrical synapses. Our sample of glutamatergic non-pyramidal neurons displayed a wide variety of somatodendritic morphologies and a subset of these cells expressed the Nurr1 mRNA, a marker for ipsilateral, but not commissural cortico-cortical projection neurons in layer VI. Comparison with spiny stellate and pyramidal neurons from other layers showed that glutamatergic neurons consistently exhibited a low occurrence of GABAergic interneuron markers and regular spiking firing patterns. Analysis of electrophysiological diversity using unsupervised clustering disclosed three groups of cells. Layer V pyramidal neurons were segregated into a first group, while a second group consisted of a subpopulation of layer VI neurons exhibiting tonic firing. A third heterogeneous cluster comprised spiny stellate, layer II/III pyramidal and layer VI neurons exhibiting adaptive firing. The segregation of layer VI neurons in two different clusters did not correlate with their somato-dendritic morphologies or with Nurr1 expression. Our results suggest that electrophysiological similarities between neocortical glutamatergic neurons extend beyond layer positioning, somatodendritic morphology and projection specificity.