Role of PEPT2 in glycylsarcosine transport in astrocyte and glioma cultures

Abstract

The aims of the current study were (1) to quantify the role of PEPT2 in the uptake of glycylsarcosine (GlySar) in cultured neonatal astrocytes and (2) to examine GlySar transport and PEPT2 expression in two glioma cell lines. The uptake of [ 14C]GlySar was measured in astrocytes cultured from neonatal mouse (PEPT2 +/+ and PEPT2 −/−) and rat, as well as rat C6 and F98 glioma cells. PEPT2 expression was examined by reverse transcription-polymerase chain reaction (RT-PCR). Neonatal astrocytes from PEPT2 −/− mice had a 94% reduction in [ 14C]GlySar uptake compared to wild type mice and there was no saturable transport. In PEPT2 +/+ mice, [ 14C]GlySar uptake was saturable ( V max 58 ± 12 pmol/mg/min, K m 107 ± 46 μM, K d 0.043 ± 0.004 μl/mg/min). In neonatal rat astrocytes, kinetic analysis also suggested that [ 14C]GlySar uptake was via a single transporter. The inhibitor profile and pH dependence of that transport process was consistent with PEPT2. In C6 and F98 glioma cells, [ 14C]GlySar uptake was markedly reduced (∼96–98%) compared to that in neonatal astrocytes and this was reflected by an absence of PEPT2 mRNA expression. These results indicate that PEPT2 is the sole transporter involved in the uptake of GlySar into neonatal cultured astrocytes. However, PEPT2 mRNA appears to be absent from two glioma cell lines.