A principal challenge of systems neuroscience is to understand the unique characteristics of cortical neurons and circuits that enable area- and species-specific sensory encoding, motor function, cognition, and behavior. To address this issue, we compared properties of layer 3 pyramidal neurons in 2 cortical areas that span a broad range of cortical function—primary sensory (V1), to cognitive (frontal)—in the mouse and the rhesus monkey. Hierarchical clustering and discriminant analyses of 15 physiological and 25 morphological variables revealed 2 fundamental principles. First, V1 and frontal neurons are remarkably similar with regard to nearly every property in the mouse, while the opposite is true in the monkey, with V1 and frontal neurons exhibiting significant differences in nearly every property assessed. Second, neurons within visual and frontal areas differ significantly between the mouse and the monkey. Neurons in mouse and monkey V1 are the same size, but differ in nearly every other way; mouse frontal cortical neurons are smaller than those in the monkey and also differ substantially with regard to most other properties. These findings have broad implications for understanding the differential contributions of heterogeneous neuronal types in construction of cortical microcircuitry in diverse brain areas and species.