Placental Transport of Riboflavin in Mice

Transport activity for [³H]riboflavin across the placenta of pregnant Slc52a3+/− mice, which were mated with Slc52a3+/− male mice, at gestation date 16.5 was determined as previously reported36 (link). Pregnant dams were anesthetized with an intraperitoneal administration of sodium pentobarbital. Surgery was performed on each animal lying on a heating pad to maintain constant body temperature. Then, radioisotope-labeled riboflavin (500 nM of [³H]riboflavin, 0.903 TBq/mmol, Moravek Biochemicals) or glucose (40 μM of D-[U-¹⁴C]glucose, 11.5 GBq/mmol, GE Healthcare) was administrated as a bolus via the femoral vein (10 mL/kg). Five minutes after administration, fetuses and placentas were removed and weighted. Fetal tails were collected, and used for genotyping as described above. Fetuses were homogenized in saline, and the samples were placed into Ultima Gold scintillation cocktail (PerkinElmer) followed by solubilization in SOLVABLE (Packard Instrument Co.). The radioactivity was measured by liquid scintillation counting, and the levels of [³H]riboflavin and D-[U-¹⁴C]glucose in each fetus were determined and their transporter activities were calculated.