Diversity in excitatory synaptic transmission by cortical pyramidal neurons give rise to the possibility of different neuronal networks that mediate distinct cortical function. Understanding heterogeneity of excitatory input to pyramidal neurons across distinct cortical areas and species will provide insight into cortical specialization and, ultimately, selective vulnerability of cortical areas to neuropathology in humans. In a previous study in our laboratory (Medalla and Luebke, 2015), significant differences in the ultrastructural features of excitatory asymmetric synapses in layers 2-3 (L2-3) neuropil were found between two distinct cortical areas in the rhesus monkey – primary sensory visual (V1) versus higher-order lateral prefrontal (LPFC) cortices. Here, we used serial sectioning electron microscopy to determine whether these differences in synaptic elements also exist in the corresponding visual (V1) and frontal (FC) cortices in the mouse. Multiple analyses of L2-3 neuropil of FC and V1 in mouse revealed three fundamental principles. First, in contrast to the diverse synapses in monkey LPFC and V1, asymmetric axospinous synapses in L2-3 neuropil of mouse FC and V1 are remarkably homogenous with regard to presynaptic and postsynaptic entities. Second, asymmetric axospinous synapses in L2-3 neuropil of mouse V1 resemble that of monkey V1 in postsynaptic entities, but differ in presynaptic entity. Third, asymmetric axospinous synapses in L2-3 neuropil of mouse FC and monkey LPFC differ substantially in both presynaptic and postsynaptic entities. These findings have broad implications for extrapolation of excitatory synaptic transmission data from one cortical area to another, and also from one species to another.