In intact adult vertebrates, muscles can be activated with a high degree of specificity, so that even within a single traditionally defined muscle, groups of motor units can be differentially activated. Such differential activation might reflect detailed control by descending systems, potentially resulting from postnatal experience such that activation of motor units is precisely tailored to their mechanical actions. Here we examine the degree to which such specific activation can be seen in the rhythmic patterns produced by isolated spinal motor systems in neonates. We examined motor output produced by the in vitro neonatal rat spinal cord with hindlimb attached. We recorded the activity of different regions within the posterior portion of biceps femoris (BFp; i.e., excluding the anterior/vertebral head). We found that in the rhythms evoked by bath application of serotonin/N-methyl-d-aspartate (5-HT/NMDA), all regions of BFp were active during extension. However, the regions of BFp were activated in a specific sequence, with the activation of rostral regions consistently preceding those of more caudal regions in both afferented and deafferented preparations. In the rhythms evoked by cauda equina (CE) stimulation, rostral and middle regions of BFp remained active in extension, but the caudal region of BFp was usually active in flexion. Stimulation of L5 and S2 dorsal roots typically evoked rhythms with all regions of BFp active during extension; although the same rostral to caudal sequence of activation observed in 5-HT/NMDA evoked rhythms could also be observed in these rhythms, we also observed cases with reversed sequences, with activity proceeding from caudal to rostral. S2 dorsal root stimulation occasionally evoked rhythms with flexor activity in caudal BFp, similar to CE-evoked rhythms. Taken together, these results suggest a high degree of individuated control of muscles by spinal pattern generating networks, even at birth.