To understand the physiological properties and anatomical organization of the spatiotemporal interaction of the responses to multiwhisker stimulation in neurons of the rat barrel cortex, single-unit recordings of 114 neurons were performed across all layers (layer II/III, n = 39; IV, n = 33; V/VI, n = 42) of the posteromedial barrel subfield of the primary somatosensory cortex of anesthetized rats. Two neighboring principal and adjacent whiskers (PW and AW, respectively) in the same row were deflected rostrally or caudally at varying interstimulus intervals (ISIs). In 37% of the neurons, the response to the combined stimulus was significantly larger than the sum of the responses to stimulation of the individual whiskers. In instances in which response facilitation was observed, selectivity was noted for the combination (75%) of the PW with a particular AW or for a particular direction (60%) of whisker deflection. The direction bias of the responses to multiwhisker stimulation was well correlated with that of the sum of the responses to single whisker stimulation (r = 0.83; p < 0.001). The pattern and magnitude of the response interaction in the neurons of the superficial layers were closely related to the location of the recorded cell in the barrel columns. Multiwhisker stimulation at short ISIs (</=3 msec) evoked prominent response facilitation in cells located close to the border between two columns (p < 0.05, one-way ANOVA), where two excitatory inputs were expected to arrive at the same time. Our results suggest that the spatiotemporal patterns of multiwhisker stimulation, such as whisker combination, direction of deflection, and timing, are expressed as different magnitudes of response interaction, which depends on the proximity of cells to home and adjacent barrel columns.