1. Volume 127, Issue 5, Pages 1410-1422 (November 2004)
H+/amino acid transporter 1 (PAT1) is the imino acid carrier: An intestinal nutrient/drug transporter in human and rat 
Catriona M.H. Anderson, Danielle S. Grenade, Michael Boll, Martin Foltz, Katherine A. Wake, David J. Kennedy, Lars K. Munck, Seiji Miyauchi, Peter M. Taylor, Frederick Charles Campbell, Bjarne G. Munck, Hannelore Daniel, Vadivel Ganapathy, David T. Thwaites 
Gastroenterology 
Volume 127, Issue 5, Pages (November 2004)
DOI: /j.gastro
Copyright © 2004 American Gastroenterological Association Terms and Conditions

2. Figure 1
Time and Na+ dependence of β-alanine uptake across the apical membrane of Caco-2 cell monolayers. β-Alanine (100 μmol/L) uptake was measured over 0.5–90 minutes at apical pH 6.5 (basolateral pH 7.4) in the presence (filled squares) and absence (open triangles) of Na+. (Insert) Data from the main panel expressed as the percentage of total uptake in the presence of Na+. Na+-dependent uptake (filled bars) was calculated as the difference between uptake in the presence and absence of Na+ (open bars). Results are mean ± SEM (n = 12–18).
Gastroenterology  , DOI: ( /j.gastro )
Copyright © 2004 American Gastroenterological Association Terms and Conditions

3. Figure 2
Pharmacological modulation of PAT-mediated amino acid uptake. Apical β-alanine (100 μmol/L) uptake in Caco-2 cells (15 minutes; apical pH 6.5) in the presence and absence of Na+ and the NHE3 selective inhibitor S1611 (3 μmol/L; open bars), the NHE inhibitor EIPA (100 μmol/L; hatched bars), or the NHE1/NHE2 inhibitor cariporide (10 μmol/L; horizontal striped bars). Data are mean ± SEM (n = 12). *P < .05 vs. control in the presence of Na+. NS, P > .05 vs. control. (Insert) Concentration-dependent β-alanine uptake (Na+ solutions, 15 minutes, apical pH 6.5) in the presence (open squares) or absence (filled circles) of apical S1611 (3 μmol/L). Results are mean ± SEM (n = 11–12). Solid lines are the best-fit curves for the hyperbolic Michaelis-Menten kinetics of the data (saturable plus nonsaturable components; r2 = for both curves). The dotted lines represent the carrier-mediated (saturable) uptake (after subtraction of the nonsaturable component).
Gastroenterology  , DOI: ( /j.gastro )
Copyright © 2004 American Gastroenterological Association Terms and Conditions

4. Figure 3
NHE3-dependent recovery of pHi after amino acid-induced acidification. BCECF-loaded Caco-2 cell monolayers were exposed to β-alanine (20 mmol/L; 300 seconds; open bars) in the apical superfusate (Na+ free, pH 5.5; basolateral Na+ free, pH 7.4 throughout). pHi recovery was then measured under various conditions (hatched bars) before the addition of an Na+-containing, pH 7.4 apical solution (filled bars). S1611 was added before (for 10 minutes) and then during exposure to β-alanine (giving a total incubation of 15 minutes before pHi recovery was measured). Composite responses are shown of single monolayers to (A) successive recoveries at apical pH 5.5–7.4 in the presence of Na+ and (B) successive recoveries (at apical pH 7.4) in Na+-containing (i) and Na+-free (ii) solution and in Na+ after exposure to apical S1611 (3 μmol/L) (iii). (C) Mean data for (A) and (B). H+ efflux was calculated from ΔpHi over the initial 30 seconds of pHi recovery.35,37 Data (mean ± SEM; n = 4–7) are expressed as the percentage of H+ efflux in control conditions (Na+ containing; pH 7.4) and were analyzed by the Student t test (paired data): **P < .01; ***P < .001; both vs. control.
Gastroenterology  , DOI: ( /j.gastro )
Copyright © 2004 American Gastroenterological Association Terms and Conditions

5. Figure 4
PAT1 messenger RNA and protein expression in human and rat tissues. Products of PCR with human multiple tissue (A) and digestive tract tissue cDNA panels (B) by using primers specific for hPAT1 (upper panels) and glyceraldehyde phosphate dehydrogenase (lower panels). (A) Lanes are as follows: 1, heart; 2, brain; 3, placenta; 4, liver; 5, lung; 6, skeletal muscle; 7, pancreas; 8, kidney. (B) Lanes are as follows: 1, ascending colon; 2, descending colon; 3, transverse colon; 4, duodenum; 5, ileocecum; 6, ileum; 7, jejunum; 8, rectum; 9, cecum; 10, stomach; 11, liver; 12, esophagus. Ctrl lane shows the positive control reaction with Caco-2 cDNA. Lane N shows the negative control reaction (no cDNA). DNA size markers (with the 1000-bp band identified) are shown. (C-E) Cross-sectional (xz) sections through a Caco-2 cell monolayer (arrows indicate apical membrane, and arrowheads indicate basal membrane) showing immunofluorescent detection of (C) PAT1 (green), (D) rBAT (red), and (E) apical colocalization (yellow). (F-K) A stack of xy sections [at 1.8-μm intervals moving down from the apical (F) to basal (K) surface] through a Caco-2 cell monolayer showing PAT1 localization to the apical membrane (green), whereas CD98 is solely basolateral (red). PAT1 immunofluorescence in rat ileum (L-N) and human jejunum (O-P) is localized to the brush-border membrane (M and P) (arrows). The PAT1 fluorescence is blocked by preadsorption with the antigenic peptide (N). Some lamina propria (but no brush-border) staining is observed with the secondary antibody alone (L and O). All scale bars are 20 μm.
Gastroenterology  , DOI: ( /j.gastro )
Copyright © 2004 American Gastroenterological Association Terms and Conditions

6. Figure 5
PAT/hPAT1 show the same substrate selectivity as the rat imino acid transporter. Apical MeAIB (20 μmol/L) uptake in Caco-2 cells in the presence (0.1–10 mmol/L) of (A) α-ABA, β-ABA, and GABA and (B) l-cycloserine, d-cycloserine, and MeLeu. Data are mean ± SEM (n = 12–20) and are expressed as percentage of control (the absence of competing amino acids). (C) Competition experiments showing the inhibition of proline (100 μmol/L) uptake into hPAT1-expressing oocytes by various amino acids (all 10 mmol/L). Substrates are as in (F) below: i, GABA; ii, β-ABA; iii, α-ABA; iv, isonipecotic acid; v, nipecotic acid; vi, l-pipecolic acid; vii, d-pipecolic acid; viii, l-cycloserine; ix, d-cycloserine; x, MeLeu. Data are mean ± SEM (n = 9–20) and are expressed as percentage of control (Ctrl, the absence of competing amino acids) after subtraction of uptake into water-injected oocytes. (D) Current induced by various amino acids (all 10 mmol/L; pH 5.5; Na+ free) in 2-electrode voltage-clamped hPAT1-expressing oocytes. (E) ΔpHi in Caco-2 cell monolayers after apical superfusion with various amino acids (10 mmol/L; pH 5.5; Na+ free; open bars). Closed bars represent pH 7.4 Na+-containing apical superfusate. Control (Ctrl) indicates the ΔpHi after exposure to the pH 5.5, Na+-free superfusate alone. (F) Correlation between amino acid-induced current in mPAT1-expressing oocytes and ΔpHi in Caco-2 cells. Data are mean ± SEM and are expressed as percentage of GABA response. The linear relationship (r2 = 0.946) between current and ΔpHi suggests that PAT1 (in oocytes) and PAT (in Caco-2 cells) discriminate between potential substrates in a similar manner.
Gastroenterology  , DOI: ( /j.gastro )
Copyright © 2004 American Gastroenterological Association Terms and Conditions

7. Figure 6
pH-dependent, Na+-independent, PAT-type transport in rat small intestine. MeAIB (100 μmol/L) uptake into rat, rabbit, or guinea pig small intestine is shown. Uptake was measured over 30 seconds in the absence of Na+ at a luminal pH of 7.2 or Data are mean ± SEM (n = 8–12). ***P < .001; NS, P > .05; both vs. pH 7.2.
Gastroenterology  , DOI: ( /j.gastro )
Copyright © 2004 American Gastroenterological Association Terms and Conditions