Effects of purinergic agonists (,-meATP and ATP) and cyclophosphamide induced cystitis on bladder afferent nerve (BAN) activity were studied in an in vitro bladder-pelvic nerve preparation. Distension of the bladder induced spontaneous bladder contractions that were accompanied by multiunit afferent firing. Intravesical administration of 40 and 130 µM ,-meATP increased afferent firing from 27 ± 3 spikes/s to 53 ± 6 and 61 ± 2 spikes/s, respectively but did not change the maximum amplitude of spontaneous bladder contractions. Electrical stimulation on the surface of the bladder elicited action potentials (AP) in BAN. ,-meATP decreased the voltage threshold from 9.0 ± 1.2 V to 3.5 ± 0.5 V (0.15 ms pulse duration) and increased the area of the APs (82 % at 80 V stimulus intensity). These effects were blocked by TNP-ATP (30 µM). ATP (2 mM) applied in the bath produced similar changes in BAN activity. These effects were blocked by bath application of PPADS (30 µM). Neither TNP-ATP nor PPADS affected BAN activity induced by distension of the bladder. Cystitis induced by pretreatment of the rats with cyclophosphamide (100 mg/kg, i.p.) increased afferent firing in response to isotonic bladder distension (10-40 cmH2O), decreased the threshold and increased the area of evoked APs. The increase in afferent firing at 10 cmH2O intravesical pressure was reduced 52 % by PPADS. These results indicate that purinergic agonists acting on P2X receptors and cystitis induced by cyclophosphamide can increase excitability of the bladder afferent nerves.