Measurement of NO levels in bovine oviductal cells.

(a) Assessment of NO2−/NO3− production. Oviductal cultures were incubated for 60 min at 38.5°C and 5% CO2 in the presence of M199 medium (control) or MetAEA (1.4 nM). Then, once removed the culture medium, cells were incubated in M199 for 24 h at 38.5°C with 5% CO2 to allow the accumulation of NO2− and NO3− in culture supernatants. NO production (NO3− plus NO2−) was measured using the Griess assay; (n = 26; p>0.05).(b) Assessment of NO synthase activity. Confluent cell monolayers were incubated for 30 min at 38.5°C and 5% CO2 in the presence of: 1) M199 medium (control) or 2) MetAEA (1.4 nM). Control and treated cells were removed by trypsinization, centrifuged and incubated for 15 min at 37°C in a HEPES buffer containing L-[14C]-Arginine and NADPH (see material and methods); (n = 7; p>0.05).Data are expressed as mean ± SEM.