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We investigated the effect of DDAH/ADMA/NOS pathway in penile tissue of rats of different age to better understand the mechanism of age-related erectile dysfunction(ED). The Sprague-Dawley male rats were assigned to young group (3 month old, n=10) and old group (18 month old, n=10). Intracavernous pressure (ICP)was measured before and after papaverine intracaverous injection.Pathology structure of penile tissue was evaluated under transmission electron microscope. The expression amounts of ADMA and cyclic guanosine monophosphate (cGMP) in penile tissue were detected by ELISA; the expression levels of isoform-specific DDAH and NOS was assessed via Western blot. Compared with the young group, the ICP in old group rat decreased significantly (33.46+5.37 vs 39.71±3.67 mmHg,p=0.02) after papaverine injection. Diffused fibrosis and impairment of endothelial cell were observed in corpus cavernosum in old group rat. Higher level of ADMA( 10.83±0.96 vs7.51±1.39μmol/gpro, p=3. 14× 10-4) and lower level of cGMP (29.42±3.84 vs 47.09±6.07 nmol/gpro, p=1.57×l0-6) were detected in penile tissue of old group compared with those of young group. Expression of DDAHI, DDAH2,endothelial NOS(eNOS) and neuronal NOS(nNOS) were all decreased significantly in penile tissue of old group rat. The DDAH/ADMA/NOS regulation pathway change dramatically accompanying with lower ICP in old group rat compared with those of the young group. This finding in rat may be suggestive in understand the mechanism of aged-related ED in human.