Function and expression of the proton‐coupled amino acid transporter PAT1 along the rat gastrointestinal tract: implications for intestinal absorption of gaboxadol

Abstract

BACKGROUND AND PURPOSE Intestinal absorption via membrane transporters may determine the pharmacokinetics of drug compounds. The hypothesis is that oral absorption of gaboxadol (4,5,6,7‐tetrahydroisoxazolo [5,4‐c] pyridine‐3‐ol) in rats occurs via the proton‐coupled amino acid transporter, rPAT1 (encoded by the gene rSlc36a1). Consequently, we aimed to elucidate the in vivo role of rPAT1 in the absorption of gaboxadol from various intestinal segments obtained from Sprague‐Dawley rats. EXPERIMENTAL APPROACH The absorption of gaboxadol was investigated following its administration into four different intestinal segments. The intestinal expression of rSlc36a1 mRNA was measured by quantitative real‐time PCR. Furthermore, the hPAT1‐/rPAT1‐mediated transport of gaboxadol or L‐proline was studied in hPAT1‐expressing Xenopus laevis oocytes, Caco‐2 cell monolayers and excised segments of the rat intestine. KEY RESULTS The absorption fraction of gaboxadol was high (81.3–91.3%) following its administration into the stomach, duodenum and jejunum, but low (4.2%) after administration into the colon. The pharmacokinetics of gaboxadol were modified by the co‐administration of L‐tryptophan (an hPAT1 inhibitor) and L‐proline (an hPAT1 substrate). The in vitro carrier‐mediated uptake rate of L‐proline in the excised intestinal segments was highest in the mid jejunum and lowest in the colon. The in vitro uptake and the in vivo absorption correlated with the expression of rSlc36a1 mRNA along the rat intestine. CONCLUSIONS AND IMPLICATIONS These results suggest that PAT1 mediates the intestinal absorption of gaboxadol and therefore determines its oral bioavailability. This has implications for the in vivo role of PAT1 and may have an influence on the design of pharmaceutical formulations of PAT1 substrates.

Intestinal Drug Transport via the Proton-Coupled Amino Acid Transporter PAT1 (SLC36A1) Is Inhibited by Gly-Xaa Dipeptides

“…7 The inhibition of the initial absorption was also shown recently, where gaboxadol (10 mg/kg), L-proline (PAT1 substrate), and L-tryptophan (PAT1 inhibitor) were coadministered directly into duodenum, jejunum, or colon of Sprague−Dawley rats. 2 Upon jejunal administration, L-proline and L-tryptophan significantly decreased the initial (≤t max ) and maximal absorption without affecting the absorption fraction. 2 In the present study, there is a nonsignificant tendency toward a Gly-X aa mediated reduction in the initial plasma concentration measured 5 min after gaboxadol dosing ( Figure 4); however, because the gaboxadol absorption is so fast, the time-resolution of the present study is not sufficient for an adequate determination of a Gly-X aa dipeptide induced inhibition of the initial absorption of gaboxadol.…”

“…2 The transepithelial flux, J, of gaboxadol (mass/time/area) across the Caco-2 cell monolayer was calculated from Fick's first law as the amount of gaboxadol, Q, accumulating in the receptor compartment per time, t, and area, A, at steady-state conditions:…”

“…At the same time, the expression of Slc36a1 mRNA (expressing PAT1) is high in the proximal parts of the small intestine, and almost absent in the colon, suggesting that it is indeed PAT1 that mediates the intestinal absorption of gaboxadol. 2 Even though PAT1 is an amino acid transporter, we have recently identified the dipeptidomimetic drug substance δ-aminolevulinic acid as a substrate of hPAT1. 11 δ-Aminolevulinic acid is structurally a Gly-Gly mimetic, and Gly-Gly and related mimetics were subsequently identified as PAT1 substrates.…”

PAT1 (SLC36A1) shows nuclear localization and affects growth of smooth muscle cells from rats

“…Threedimensional data sets were acquired using a confocal laser scanning microscope (CLSM) equipped with an Axiovert 100M microscope under ϫ20 or ϫ63 magnification, using 1. 6.67 nM and 8.5 M, respectively, in all uptake or flux experiments, and the specific activity was kept at 0.5 Ci/ml. The sodium dependency of L-[ 3 H]proline uptake was investigated using the same procedure as described above but applying sodium-free HBSS (1.3 mM CaCl2, 0.…”

“…PAT1 mRNA is expressed in several mammalian tissues, including the heart, brain, placenta, testis, spleen, liver, lung, skeletal muscle, pancreas, intestine, skin, and kidney (1,3,6,8,22). PAT1 has a number of functions in cells and tissues, including luminal uptake of amino acids, sensing the availability of amino acids, being a target for rapamycin complex 1 (TORC1) activation, and regulating cell growth in different models (7,11,15,17,23).…”