PEPT2-mediated transport of 5-aminolevulinic acid and carnosine in astrocytes

5-Aminolevulinic acid (ALA) and carnosine have important physiological and pathophysiological roles in the CNS. Both are substrates for the proton-coupled oligopeptide transporter PEPT2. The purpose of the current study was to determine the importance of PEPT2 in the uptake of ALA and carnosine in rat and mouse (PEPT2+/+ and PEPT2−/−) cultured neonatal astrocytes. Although neonatal astrocytes are known to express PEPT2, its quantitative importance in the transport of these compounds is not known. [14C]ALA uptake in neonatal rat astrocytes was inhibited by dipeptides, an α-amino containing cephalosporin (which is a PEPT2 substrate) but was not affected by a non-amino containing cephalosporin (which is not a PEPT2 substrate). Uptake was pH sensitive as expected from a proton-coupled transporter and was saturable (Vmax = 715 ± 29 pmol/mg/min, Km = 606 ± 14 μM). [3H]Carnosine uptake in neonatal rat astrocytes was inhibited by dipeptides but not by histidine (a substrate for the peptide/histidine transporters PHT1 and PHT2) and also showed saturable transport (Vmax = 447 ± 23 pmol/mg/min, Km = 43 ± 5.5 μM). Neonatal astrocytes from PEPT2−/− mice had a 62% reduction in [14C]ALA uptake and a 92% reduction in [3H]carnosine uptake compared to PEPT2+/+ mice. These results demonstrate that PEPT2 is the primary transporter responsible for the astrocytic uptake of ALA and carnosine.