Chronic amphetamine intake leads to neurogenic bladder and chronic urinary retention. The mechanism underlying persistent urinary retention is unclear. The pelvic-urethral reflex (PUR) is essential for the urethra to develop sufficient resistance to maintain urine continence, an important function of the urinary system. Recent studies on PUR activities have indicated that repetitive/tetanic stimulation of the pelvic afferent fibers induces spinal reflex potentiation (SRP) in PUR activities, which further increases urinary retention. In this study, results showed that test stimulation (TS, 1/30 Hz) evoked a baseline reflex activity, while repetitive stimulation (RS, 1 Hz) induced reflex potentiation in the external urethral sphincter. Intrathecal d-amphetamine (AMPH, 30 μM) did not but higher AMPH concentration (100 μM) induced SRP in TS-induced reflex activity. H89 (10 μM, a protein kinase A inhibitor), but not chelerythrine chloride (CTC, 10 μM, a protein kinase C inhibitor), prevented the 100 μM AMPH-elicited SRP. At 30 μM, forskolin, an activator of adenylyl cyclase, elicited SRP. The co-administration of 10 μM forskolin and 30 μM AMPH induced SRP in TS-induced reflex activity. These results implied that the repetitive/tetanic stimulation of the pelvic afferent fibers could induce SRP in PUR activities, so that the urethra can produce sufficient resistance and played a significant role in urinary retention. Findings in this study demonstrated that amphetamine could induce bladder dysfunction by triggering protein kinase A activation, and provide a practical basis for the development of treatment for amphetamine-associated urinary retention.