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PHT1 (SLC15A4) is responsible for translocating L-histidine (L-His), di/tripeptides and peptide-like drugs across biological membranes. Previous studies have indicated that PHT1 is located in brain parenchyma, however, its role and significance in brain, along with its impact on the biodistribution of substrates is unknown. In the present study, adult gender-matched Pht1-competent (wildtype) and Pht1-deficient (null) mice were used to investigate the effect of PHT1 on L-His brain disposition via in vitro slice and in vivo pharmacokinetic approaches. Initial phenotyping of the two genotypes and expression measurements of select transporters/enzymes were also performed. No significant differences were observed between genotypes in serum chemistry, body weight, viability and fertility. Polymerase chain reaction (PCR) analyses indicated that Pept2 had a compensatory up-regulation in Pht1 null mice (about 2-fold) as compared to wildtype animals, which was consistent in different brain regions and confirmed by immunoblots. The uptake of L-His was reduced in brain slices by 50% during PHT1 ablation. The L-amino acid transporters accounted for 30% of the uptake, and passive (other) pathways for 20% of the uptake. During the in vivo revealed that, when sampled 5 min after dosing, L-His values were 28–48% lower in Pht1 null mice as compared to wildtype animals, in brain parenchyma but not cerebrospinal fluid. Concentration-time profiles of the in vivo samples were then analyzed using nonlinear mixed effects modeling with NONMEM v7.3. In addition to active PHT1- mediated uptake into brain parenchyma, influx and efflux rate constants of L-His between plasma, brain parenchyma and CSF were modeled as first-order processes. Diffusion between brain parenchyma and CSF, CSF bulk flow and tissue volumes were obtained from the literature. The disposition kinetics of L-His in plasma, CSF and brain parenchyma was best described by a four-compartment model. We observed that the plasma and CSF PK profiles of L-His were comparable in WT and KO mice. However, a more rapid uptake of L-His occurred in the brain parenchyma of WT mice due to active transport by PHT1, which was modeled with a Michaelis- Menten term (Km = 39.9 μM and Vmax = 0.140 nmol/min). Our model quantitatively described the transport kinetics of PHT1-mediated uptake of L-His in brain, for the first time, under in vivo conditions. The results suggest that PHT1 may play an important role in histidine transport in brain, and resultant effects on histidine/histamine homeostasis and neuropeptide regulation. The findings also provide a valuable tool in predicting the disposition of L-His in brain and the potential of PHT1 as a drug target to treat serious CNS diseases.pharmacokinetic (PK) studies, plasma concentration-time profiles of L-His were comparable between the two genotypes after intravenous administration. Still, biodistribution studies

myo-Inositol synthesis and catabolism are crucial in many multicellular eukaryotes for production of phosphatidylinositol and inositol phosphate signaling molecules. myo-inositol monophosphatase (IMP) is a major enzyme required for the synthesis of myo-inositol and breakdown of inositol (1,4,5)-trisphosphate (InsP3), a potent second messenger involved in many biological activities. Arabidopsis contains a single canonical IMP gene, which was previously shown in our lab to encode a bifuntional enzyme with both IMP and L-galactose 1-phosphatase activity. Analysis of metabolite levels in imp mutants showed only slight modifications with less myo-inositol and ascorbate accumulation in these mutants. This result suggests the presence of other functional IMP enzymes in plants. Two other genes in Arabidopsis encode chloroplast proteins, which we have classified as IMP-like (IMPL), because of their greater homology to the prokaryotic IMPs such as the SuhB, and CysQ proteins. Prokaryotic IMP enzymes are known to dephosphorylate D-Inositol 1-P (D-Ins 1-P) and other substrates in vitro, however their in vivo substrates are not characterized. A recent study revealed the ability of IMPL2 to complement a bacterial histidinol 1-phosphate phosphatase mutant defective in histidine synthesis, which suggested an important role for IMPL2 in amino acid synthesis. The research presented here focuses on the characterization of IMPL functional roles in plant growth and development. To accomplish this I performed kinetic comparisons of the Arabidopsis recombinant IMPL1 and IMPL2 enzymes with various inositol phosphate substrates and with L-histidinol 1-phosphate, respectively. The data supports that IMPL2 gene encodes an active histidinol 1-phosphate phosphatase enzyme in contrast to the IMPL1 enzyme which has the ability to hydrolyze D-Ins 1-P substrate and may be involved in the recycling of inositol from the second messenger, InsP3. Analysis of metabolite levels in impl2 mutant plants reveals that impl2 mutant growth is impacted by alterations in the histidine biosynthesis pathway. Together these data solidify the catalytic role of IMPL2 in histidine synthesis in plants and highlight its importance in plant growth and development.