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Objective: To investigate the different expression of three isozymes of nitric oxide synthase (NOS) in diabetic rat colons and the contribution to the colonic dysfunction. Methods: Sprague-Dawley (SD) rats were used in this experiment and diabetes were induced by streptozotocin (65 mg/kg, i.v. ). Three isozymes of NOS (nNOS, iNOS and eNOS) expression in proximal and distal colon were measured in two weeks after diabetes induction using the methods of immunocytochemistry and semi-quantitative reverse transcription-polymerase chain reaction (RT-PCR). Results: Positive immunoreactivity for nNOS was found in intermuscular and submucous plexus neuronal cells, neither eNOS nor iNOS had been found in any layers of colon in the two groups. The expression of nNOS mRNA was significantly increased in diabetic colon than that in control rats as determined by RT-PCR. The eNOS mRNA level of diabetic colon was lower compared to thecontrol rats, while no expression of iNOS mRNA was found in the normal or diabetic rats. Conclusion: This report has demonstrated that nNOS increased and eNOS decreased in rat colon in the early stages of diabetes. NO production by the nNOS might play a key role in colonic dysfunction, as supported by raised nNOS mRNA and enzyme expression in the diabetic colon. Reduced eNOS activity might also contribute to colonic dysfunction in experimental diabetes.