PURPOSE: The purpose of this research was to determine the polarity and directionality of the PEPT2-mediated uptake and transepithelial transport of the neuropeptide glycyl-L-glutamine (GlyGln) in choroid plexus. METHODS: The transport kinetics of [3H]GlyGln was studied in neonatal rat choroid plexus epithelial cells in primary culture grown on laminin-coated Transwell filter inserts. Using a bicarbonate artificial cerebrospinal fluid (CSF) buffer (pH 7.4) at 37 degrees C, GlyGln studies were performed as a function of time, substrate concentration, and the presence of potential inhibitors (at 1 mM). RESULTS: GlyGln (2 microM) accumulation was about three to four times greater when introduced from the apical (CSF-facing) as opposed to the basal (blood-facing) side of the cell monolayer, and transepithelial transport was about two times greater in the apical-to-basal direction. The apical uptake of radiolabeled GlyGln (2 microM) was inhibited significantly by dipeptides (i.e., unlabeled GlyGln and cysteinylglycine) and some neuropeptides (i.e., carnosine, N-acetylaspartylglutamate, kyotorphin), but was unaffected by amino acids (i.e., glycine, glutamine) as well as by [D-Arg2]-kyotorphin and glutathione. The concentration-dependent apical uptake of GlyGln (2-1000 microM) was characterized by a high-affinity process (i.e., Vmax of 72 pmol/mg/min; Km of 136 microM), consistent with the properties of PEPT2. The intracellular hydrolysis of GlyGln was extensive, however, with only 40% of the dipeptide remaining intact after 1 h. CONCLUSIONS: The results demonstrate that PEPT2 plays an important role in regulating the apical uptake of GlyGln at the blood-CSF interface. Once inside the cell, GlyGln is rapidly degraded to its constitutive amino acids for further processing.
PURPOSE: The purpose of this research was to determine the polarity and directionality of the PEPT2-mediated uptake and transepithelial transport of the neuropeptide glycyl-L-glutamine (GlyGln) in choroid plexus. METHODS: The transport kinetics of [3H]GlyGln was studied in neonatal rat choroid plexus epithelial cells in primary culture grown on laminin-coated Transwell filter inserts. Using a bicarbonate artificial cerebrospinal fluid (CSF) buffer (pH 7.4) at 37 degrees C, GlyGln studies were performed as a function of time, substrate concentration, and the presence of potential inhibitors (at 1 mM). RESULTS:GlyGln (2 microM) accumulation was about three to four times greater when introduced from the apical (CSF-facing) as opposed to the basal (blood-facing) side of the cell monolayer, and transepithelial transport was about two times greater in the apical-to-basal direction. The apical uptake of radiolabeled GlyGln (2 microM) was inhibited significantly by dipeptides (i.e., unlabeled GlyGln and cysteinylglycine) and some neuropeptides (i.e., carnosine, N-acetylaspartylglutamate, kyotorphin), but was unaffected by amino acids (i.e., glycine, glutamine) as well as by [D-Arg2]-kyotorphin and glutathione. The concentration-dependent apical uptake of GlyGln (2-1000 microM) was characterized by a high-affinity process (i.e., Vmax of 72 pmol/mg/min; Km of 136 microM), consistent with the properties of PEPT2. The intracellular hydrolysis of GlyGln was extensive, however, with only 40% of the dipeptide remaining intact after 1 h. CONCLUSIONS: The results demonstrate that PEPT2 plays an important role in regulating the apical uptake of GlyGln at the blood-CSF interface. Once inside the cell, GlyGln is rapidly degraded to its constitutive amino acids for further processing.
Authors: M D Owen; C B Unal; M F Callahan; K Trivedi; C York; W R Millington Journal: Am J Physiol Regul Integr Comp Physiol Date: 2000-11 Impact factor: 3.619
Authors: Scott M Ocheltree; Hong Shen; Yongjun Hu; Jianming Xiang; Richard F Keep; David E Smith Journal: J Pharmacol Exp Ther Date: 2003-11-04 Impact factor: 4.030
Authors: Mohamed A Kamal; Huidi Jiang; Yongjun Hu; Richard F Keep; David E Smith Journal: Am J Physiol Regul Integr Comp Physiol Date: 2009-02-18 Impact factor: 3.619