Endogenous methylarginines regulate neuronal nitric-oxide synthase and prevent excitotoxic injury

Abstract

Nitric oxide (NO) has a critical role in neuronal function; however, high levels lead to cellular injury. While guanidino-methylated arginines (MA) including asymmetric dimethylarginine (ADMA) and NG-methyl-L-arginine (NMA) are potent competitive inhibitors of nitric oxide synthase (NOS) and are released upon protein degradation, it is unknown whether their intracellular concentrations are sufficient to critically regulate neuronal NO production and secondary cellular function or injury. Therefore, we determine the intrinsic neuronal MA concentrations and their effects on neuronal NOS function and excitotoxic injury. Kinetic studies demonstrated that the Km for L-arginine is 2.38 μM with a Vmax of 0.229 μmol mg-1 min-1, while Ki values of 0.67 μM and 0.50 μM were determined for ADMA and NMA, respectively. Normal neuronal concentrations of all NOS-inhibiting MA were determined to be ≈15 μM, while L-arginine concentration is ≈90 μM. These MA levels result in >50% inhibition of NO generation from neuronal NOS. Down-modulation or up-modulation of these neuronal MA levels, respectively, dramatically enhanced or suppressed NO-mediated excitotoxic injury. Thus, neuronal MA profoundly modulate NOS function and suppress NO mediated injury. Pharmacological modulation of the levels of these intrinsic NOS inhibitors offers a novel approach to modulate neuronal function and injury.