PURPOSE: To determine the importance of PEPT2 in the uptake of glycylsarcosine (GlySar) and 5-aminolevulinic acid (5-ALA) in mouse choroid plexus whole tissue. METHODS: Uptake studies were performed in bicarbonate artificial cerebrospinal fluid buffer using choroid plexuses isolated from PEPT2+/+ and PEPT2-/- mice. [14C]GlySar and [14C]5-ALA were studied as a function of temperature, concentration, potential inhibitors, and low sodium conditions. RESULTS: PEPT2-/- mice exhibited a 90% reduction in GlySar uptake (p < 0.001) and a 92% reduction in 5-ALA uptake (p < 0.001) as compared to wild type animals. At 4 degrees C (vs. 37 degrees C), GlySar uptake was reduced by 95% in PEPT2+/+ mice; no difference was observed in null animals. Unlabeled GlySar inhibited the uptake of [14C]GlySar in PEPT2+/+ mice (p < 0.01); self-inhibition did not occur in PEPT2-/- mice. GlySar demonstrated saturable uptake in PEPT2+/+ mice (Vmax = 16.4 pmol mg(-1) min(-1), Km = 70 microM, Kd = 0.014 microl mg(-1) min(-1)), however, uptake was linear in PEPT2-/- mice (Kd = 0.023 microl mg(-1) min(-1)). Low sodium buffer (1 mM) resulted in 75% and 59% reductions, respectively, in GlySar (p < 0.001) and 5-ALA (p < 0.01) uptake in PEPT2+/+ mice; no differences were observed in PEPT2-/- mice. Overall, about 90-95% of the choroid plexus uptake of GlySar and 5-ALA was mediated by PEPT2, with about 5-10% of the residual uptake occurring by nonspecific mechanisms. CONCLUSIONS: The results demonstrate that PEPT2 is the only transporter responsible for the choroid plexus uptake of GlySar and 5-ALA. They also suggest a role for PEPT2 in the clearance of dipeptides and endogenous peptidomimetics from cerebrospinal fluid.
PURPOSE: To determine the importance of PEPT2 in the uptake of glycylsarcosine (GlySar) and 5-aminolevulinic acid (5-ALA) in mouse choroid plexus whole tissue. METHODS: Uptake studies were performed in bicarbonate artificial cerebrospinal fluid buffer using choroid plexuses isolated from PEPT2+/+ and PEPT2-/- mice. [14C]GlySar and [14C]5-ALA were studied as a function of temperature, concentration, potential inhibitors, and low sodium conditions. RESULTS:PEPT2-/- mice exhibited a 90% reduction in GlySar uptake (p < 0.001) and a 92% reduction in 5-ALA uptake (p < 0.001) as compared to wild type animals. At 4 degrees C (vs. 37 degrees C), GlySar uptake was reduced by 95% in PEPT2+/+ mice; no difference was observed in null animals. Unlabeled GlySar inhibited the uptake of [14C]GlySar in PEPT2+/+ mice (p < 0.01); self-inhibition did not occur in PEPT2-/- mice. GlySar demonstrated saturable uptake in PEPT2+/+ mice (Vmax = 16.4 pmol mg(-1) min(-1), Km = 70 microM, Kd = 0.014 microl mg(-1) min(-1)), however, uptake was linear in PEPT2-/- mice (Kd = 0.023 microl mg(-1) min(-1)). Low sodium buffer (1 mM) resulted in 75% and 59% reductions, respectively, in GlySar (p < 0.001) and 5-ALA (p < 0.01) uptake in PEPT2+/+ mice; no differences were observed in PEPT2-/- mice. Overall, about 90-95% of the choroid plexus uptake of GlySar and 5-ALA was mediated by PEPT2, with about 5-10% of the residual uptake occurring by nonspecific mechanisms. CONCLUSIONS: The results demonstrate that PEPT2 is the only transporter responsible for the choroid plexus uptake of GlySar and 5-ALA. They also suggest a role for PEPT2 in the clearance of dipeptides and endogenous peptidomimetics from cerebrospinal fluid.
Authors: Isabel Rubio-Aliaga; Isabelle Frey; Michael Boll; David A Groneberg; Hans M Eichinger; Rudi Balling; Hannelore Daniel Journal: Mol Cell Biol Date: 2003-05 Impact factor: 4.272