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The cortex is a highly organized structure and this organization is integral to cortical function. However, the circuitry underlying cortical organization is only partially understood, thus limiting our understanding of cortical function. Within the somatosensory cortex, organization is manifest as a map of the body surface. At the level of the cortical circuitry the horizontal connections of Layer 2/3 express a physiological bias that reflects discontinuities within the somatosensory map. Both excitation and inhibition are smaller when evoked from across a representational border, as compared to when they are evoked from within the representation. This physiological bias may be due to a bias in either the strength or number of synapses and/or the number of axons that cross this border and the extent of their arborization. In this study we used both an anterograde (Phaseolus vulgaris leucoagglutinin) and a retrograde (cholera toxin B) tracer to examine Layer 2/3 horizontal projections in rat S1. We determined that there is a bias in the amount of horizontal axonal projections that cross the forepaw/lower jaw border as compared to projections remaining within an individual representation. This bias in axonal projection and the correlated bias in excitation and inhibition may underlie the expression of the representational border. J. Comp. Neurol. 500:634–645, 2007. © 2006 Wiley-Liss, Inc.