Chronic inflammation of the urinary bladder generates hyperalgesia and allodynia. Growing evidence suggests a role of ERK in mediating somatic and visceral pain processing. In the present studies, we characterized and compared the activation of two ERK isoforms ERK1/2 and ERK5 in micturition pathways including the urinary bladder, lumbosacral dorsal root ganglia (DRG) and spinal cord in adult female and male rats before and after cyclophosphamide (CYP)-induced bladder inflammation. Results showed differential activation of ERK1/2 and ERK5 in these regions following cystitis. The level of phospho-ERK1/2 but not phospho-ERK5 was increased in the urinary bladder; the level of phospho-ERK5 but not phospho-ERK1/2 was increased in DRG; and the level of phospho-ERK1/2 but not phospho-ERK5 was increased in lumbar spinal cord following cystitis compared to control. Cystitis-induced up-regulation of phospho-ERK1/2 and phospho-ERK5 was time-dependent and showed similar pattern in female and male rats. The level of phospho-ERK1/2 in bladder was increased at 2-8 hr after CYP injection; the level of phospho-ERK5 in DRG was increased at 8-48 hr after CYP injection; and the level of phospho-ERK1/2 in lumbar spinal cord was increased at 48 hr after CYP injection. The results that phospho-ERK5 was exclusively increased in DRG neurons while phospho-ERK1/2 was increased in the spinal cord and the urinary bladder after cystitis suggest a region-specific effect of neurotrophins on micturition pathways following bladder inflammation.