The present study provides an account of a sensitive and rapid experimental approach for MRI visualization and analysis of spinal cord (SC) laminar activity in normal and injured animals. This approach is based upon neuronal activity-dependant manganese (Mn) uptake after focal SC injection of MnCl(2), and subsequent ex-vivo magnetic resonance imaging (MRI) of activated SC pathways. The method was designed as an alternative to time-intensive histochemical and behavioral approaches typically used for analysis of spinal cord injury (SCI) and our results provide both anatomical and functional insights. We show that ex vivo imaging can determine layer-specific activity over an extended region of the rat SC. In addition, we demonstrate that the Mn concentration profile along the SC axis accurately reflects the type of SC injury. The approach is flexible since MRI analysis can be done immediately after animal sacrifice, or alternatively several days later, without a loss of sensitivity. Moreover, the integrity and functional state of SC circuitry can be analyzed in less than 1 h whereas several days and weeks are necessary to perform classical histochemical and behavioral analysis. Thus our method can be used for precise assessment of the extent of dysfunction or change in SC disorders and may facilitate the screening of molecules with therapeutic potential after SC injury.