Prefrontal cortical functioning depends on D1 family receptors and their complex signal transduction cascade, including protein phosphatase-1 (PP1). Three PP1 isoforms are prominent in the brain: PP1alpha, PP1beta and PP1gamma1. PP1 localization by a variety of scaffolding proteins is critical for dopamine-mediated modulation of glutamatergic neurotransmission. We have quantified the subcellular distribution of each isoform in primate prefrontal cortex using immunoelectron microscopy. All three are found in spines, dendrites, axon terminals, axons and glia. However, PP1alpha and PP1gamma1 labeling is enriched in spines, whereas PP1beta label is enriched in dendrites. Using post-embedding immunogold labeling, we further examined the distribution of PP1alpha and PP1gamma1 within spines. PP1gamma1 is highly and specifically concentrated in the postsynaptic density (PSD) of these spines, while PP1alpha is enriched in the PSD but also found subjacent to the PSD in moderate amounts. Thus, PP1 isoforms are heterogeneously distributed in the cortical neuropil and within spines. These results suggest that each PP1 isoform has access to a different set of substrates and, furthermore, they demonstrate that the composition of signal transduction proteins varies in different parts of the neuron and even in different regions of a dendritic spine in the primate PFC.