The fine scale connections of prefrontal cortex (PFC) were investigated in the rat brain, in order to determine organizational properties of PFC pathways which were previously undefined. Neuroanatomical tract tracers (Fluro-Gold, Fluoro-Ruby, Fluoro-Emerald, Biotinylated dextran amines; Fluorescein and Texas red) were injected (20 -100 nl) into subdivisions of PFC (prelimbic, infralimbic, medial-orbital, ventral-orbital, ventrolateral-orbital, lateral-orbital and dorsolateral-orbital) and their projections studied. Tracer studies identified clear evidence of significantly ordered projections from PFC to temporal and sensory-motor cortices in three axes of orientation (p<0.001), showing differential ordering of input and output connections (p<0.001). Ordered connections were consistent across PFC (from anterior to posterior) and showed evidence of changes in organisation in anterior compared to posterior PFC, in both the PFC-temporal and PFC-sensory-motor cortex pathways. Detailed analysis revealed evidence for an organizational gradient in the relationship between inputs and outputs from anterior to posterior PFC, in which retrograde and anterograde labelling become increasingly differentiated as PFC injection site is moved from posterior to anterior. Analysis of fine scale tracer injections (20-30 nl) revealed evidence to show underlying complex organizational properties of connections from PFC to temporal and sensory-motor cortices. Taken together, the findings show that PFC displays ordered arrangements of connections to temporal and sensory-motor cortex, input and output connections are consistently not found in the same locations and the relationship between inputs and outputs differs in relation to the anterior-posterior location in PFC.