The three nitric oxide synthase (NOS) isoforms; neuronal NOS (nNOS), endothelial NOS (eNOS) and inducible NOS (iNOS), play an important role in pathological and physiological conditions in kidneys. Nitric oxide (NO) plays an important role in the pathophysiology of Myoglobinuric Acute Kidney Injury (MAKI). We aimed to investigate the effects of Tetrahydrobiopterin (BH4), L-arginine, L-NAME (NOS, inhibitor) and BH4+L-arginine treatment on renal NOS isoenzymes at different time periods in experimental MAKI model. Spraque Dawley male rats were divided into 4 groups (A, B, C and D) and each group had 6 subgroups (n=8) as Control (saline, 8 ml/kg, im), MAKI (%50 glycerol, 8 ml/kg, im), MAKI+BH4 (20 mg/kg, ip), MAKI+L-Arginine (125 mg/kg, ip), MAKI+BH4+Arginine 20+125 mg/kg, ip) and MAKI+L NAME (10 mg/kg, ip). To form a mild MAKI condition in group D, glycerol was applied at a dose of 5 ml/kg of body weight. Group A, B, C and D rats were received the ip injection of treatments for 1, 2, 4 and 4 times, respectively. The rats were euthanatised after the glycerol injection; 24 hours for group A, 48 hours for group B, 96 hours for group C (8 ml/kg) and 96 hours for group D (5 ml/kg) after. Values are means±SD, compared by Mann Whitney U Test. While the MAKI group compared to the control group for the all sub-groups, kidney tissue nNOS, iNOS and eNOS gene expressions were significantly higher, but they were significantly lower in group D (p<0,05). In the group A and B, the treatment of BH4, L-Arginine and BH4+L-Arginine decreased the gene expression of nNOS (p<0,05). In the group A, while the gene expression of iNOS increased by the treatment in BH4 and BH4+L-Arginine groups, it decreased by BH4 treatment in group B. In the group A, the treatment of BH4, L-Arginine and BH4+L-Arginine decreased the gene expression of eNOS (p<0,05). In group B, treatment of BH4 decreased the eNOS gene expression (p<0,05). While MAKI groups compared with the controls of all sub-groups, quantitative analysis of immunostaining for nNOS not changed, iNOS and 3-nitrotyrosine levels were significantly higher and eNOS was significantly lower (p<0,05). Similarly, in all groups, L-Arginine, BH4+L-Arginine treatment decreased the iNOS and 3-nitrotyrosine immunostaining. And again for the all groups, L-Arginine and BH4+L-Arginine treatment increased the eNOS immunostaining. Our results support the hypothesis that administration of BH4, L-Arginine and BH4+L-Arginine were effective in attenuating nitrosative stress in MAKI. These results also suggest that BH4, L-Arginine and BH4+L-Arginine therapy may have a clinical prevention against MAKI after rhabdomyolysis and may provide further evidence of future work in this regard.