Microsoft Word - NEF097BF

Prof. José M. López-Novoa, Departamento de Fisiología y Farmacología, Facultad de Medicina, Universidad de Salamanca, Avenida Campo Charro s/n, E-37007 Salamanca (Spain) Dear Sir, NO is synthesized from L-arginine [1] in a reaction catalyzed by nitric oxide synthase (NOS) and is oxidized to nitrite and nitrate in a constant ratio, at a constant pH [2]. Nitrite production has been used as an index of glomerular NO production [3]. Arginine analogs, such as NG-mono-methyl-L-arginine (L-NMMA), NG, NG-di-methyl-Larginine (ADMA) and No-nitro-L-arginine methyl ester (L-NAME), are commonly used as ‘specific’ inhibitors of NO synthesis; however, their effects on NO levels are rarely measured. Some investigators have reported that incubation of arterial rings with L-NMMA causes increases in NO levels [4]. L-NAME has been widely used to investigate the effect of NO synthesis inhibition on renal function. There is no information about the effect of LNAME on glomerular NO synthesis in long-term incubations. Thus, the aim of this study was to investigate the effect of L-NAME on glomerular NO synthesis, measured by nitrite production. Female Wistar rats were anesthetized with ether and after washing the kidneys with ice-cold saline through the abdominal aorta, glomeruli were isolated by mechanical sieving [5] in sterile conditions, and plated out in 4 × 6 well plates at a concentration of 7,000 glomeruli per well. Glomeruli were incubated at 37 °C in endotoxin-free RPMI 1640 culture medium (Gibco) supplemented with 10% fetal calf serum. Some wells were added L-NAME 10-4M (Sigma) and some others were used as controls. Additionally, some empty wells were added RPMI medium alone and some others L-NAME and RPMI medium, to assess the effect of RPMI on L-NAME molecule.