3h l arginine

Total NOS activity was determined in the 10% of aorta homogenates by measuring [3H]-L-citrulline formation from [3H]-L-arginine (MP Biochemicals, Santa Ana, California, USA) using the Quanta Smart Tri-Carb Liquid Scintillation Analyzer (TriCarb, Packard, UK) as described elsewhere [36 (link)]. The results are expressed as picokatal per gram of protein (pkat/g protein).

Total NOS activity was measured in the 20% tissue homogenates of the LHV, aorta, brainstem, and cerebellum by determining [³H]-L-citrulline formation from [³H]-L-arginine (MP Biomedicals, USA) as described previously [33 (link)] and expressed as pmol/min/mg of tissue proteins as determined using the Lowry method. NOS activity was determined in six control and eight Epi-treated rats.

Total NO synthase (NOS) activity was measured in tissue homogenates of the aorta (200 mg/mL) by determination of [³H]-L-citrulline formation from [³H]-L-arginine (MP Biomedicals, USA, 50 Ci/mmol), as described previously [32 (link)] and expressed as pmol/min/mg of tissue proteins as determined by the Lowry method [40 (link)]. All chemicals used were purchased from Sigma-Aldrich (Germany) and Merck Chemicals (Germany).

The total NOS activity was determined in crude homogenates of the aorta by measuring the formation of [3^H]-L-citrulline from [3^H]-L-arginine (MP Biochemicals, California, CA, USA), as described elsewhere [40 (link)]. Briefly, 50 µL of 10% homogenates were incubated in the presence of 50 mmol/L Tris-HCl, pH 7.4, 10 µmol/L [³H]-L-arginine (specific activity 5 GBq/mmol), 30 nmol/L calmodulin, 1 mmol/L β-NADPH, 3 µmol/L BH₄, and 2 mmol/L Ca²⁺ in a total volume of 100 µL. After incubation at 37 °C, the reaction was stopped by the addition of 1 mL of stop buffer containing 20 mmol/L HEPES buffer pH 5.5, 2 mmol/L EDTA, 2 mmol/L EGTA, and 1 mmol/L L-citrulline. The samples were applied to Dowex 50WX-8 columns (Na⁺ form). [³H]-L-citrulline was measured with the Quanta Smart TriCarb Liquid Scintillation Analyzer (TriCarb, Packard, UK). NOS activity was expressed as picokatal per gram of protein (pkat/g protein). Protein levels were determined using the Lowry method.

Total NOS activity was determined in crude tissue homogenates from different parts of the heart – infarcted zone (IZ), injured zone (INZ) and non-infarcted zone (NIZ) by measuring [3H]-L- citrulline formation from [3H]-L- arginine (MP Biochemicals, St. Louis, MO, USA) as described elsewhere [36 (link)]. [3H]-L-citrulline content was measured by liquid scintillation calculation (TriCarb, Packard, UK). NOS activity is expressed as picokatal per gram of protein (pkat/g protein).

Total NOS activity was determined in the 10% of BS homogenates by measuring [3H]-L-citrulline formation from [3H]-L-arginine (MP Biochemicals, Santa Ana, CA, USA) using the Quanta Smart Tri-Carb Liquid Scintillation Analyzer (PerkinElmer, Beaconsfield, UK) as described in previous work [36 (link)]. Total NO synthase activity was determined in the control group (n = 7) and MLN group (n = 7). The results are expressed as picokatal per gram of protein (pkat/g protein).