Volume 118, Issue 3, Pages 535-543 (March 2000) Polarized transport of hydrophilic compounds across rat colonic mucosa from serosa to mucosa is temperature dependent  Mikio Tomita*, Michael J. Menconi‡, Russell L. Delude‡, Mitchell P. Fink‡  Gastroenterology  Volume 118, Issue 3, Pages 535-543 (March 2000) DOI: 10.1016/S0016-5085(00)70259-9 Copyright © 2000 American Gastroenterological Association Terms and Conditions

Fig. 1 Permeabilities of hydrophilic compounds (FD4, FD70, FS, LY, mannitol, and lactulose) across rat colonic mucosa in the S→M direction vs. M→S direction. Mucosal sheets from rat colon stripped of underlying seromuscular tissue were mounted into Ussing chambers filled with K-H buffer. After a 20-minute preincubation period, fresh buffer containing the permeability probe being studied was added to the donor chamber, and buffer without probe was added to the opposite (receiving) chamber. At successive 10-minute intervals, 100-μL samples of buffer were removed from the receiving chamber and replaced with an equivalent volume of buffer. Concentrations of FD4, FD70, FS, and LY were determined by fluorescence spectrophotometry; concentrations of mannitol (MAN) and lactulose (LAC) were determined by high-performance liquid chromatography. Results are expressed as a clearance (nL · min−1 · cm−2) by calculating the transmucosal flux of the probe and then dividing by the concentration of the probe in the donor chamber. Each result represents the mean ± SEM (FD4: n = 22 for M→S, n = 9 for S→M; FD70, FS, and LY: n = 4 each for both M→S and S→M; MAN and LAC: n = 5 each for both M→S and S→M). *P < 0.01 compared with the M→S direction. Gastroenterology 2000 118, 535-543DOI: (10.1016/S0016-5085(00)70259-9) Copyright © 2000 American Gastroenterological Association Terms and Conditions

Fig. 2 Effect of temperature on the polarized permeation of FD4 across rat colonic mucosa. After mucosal sheets from rat colon were prepared and mounted into Ussing chambers, permeability to FD4 was assessed in the M→S and S→M directions at both 37°C and 4°C, as described in Figure 1. Each data point represents the mean ± SEM (n = at least 4 experiments/condition). Gastroenterology 2000 118, 535-543DOI: (10.1016/S0016-5085(00)70259-9) Copyright © 2000 American Gastroenterological Association Terms and Conditions

Fig. 3 Effect of verapamil on the permeability of FD4 across rat colonic mucosa. Mucosal sheets from rat colon were prepared and mounted into Ussing chambers as described in Figure 2. After preincubating the mucosal tissue for 20 minutes with graded concentrations of verapamil, permeability to (A) FD4 was assessed in the S→M direction over an additional 1-hour incubation period in the continued presence of verapamil. Each data point represents the mean ± SEM (n = at least 4 experiments/condition). *P < 0.05, ***P < 0.001 compared with control group. Gastroenterology 2000 118, 535-543DOI: (10.1016/S0016-5085(00)70259-9) Copyright © 2000 American Gastroenterological Association Terms and Conditions

Fig. 4 Effect of dextran 10,000 (D10) on the S→M permeability of FD4 across rat colonic mucosa. Mucosal sheets from rat colon were prepared and mounted into Ussing chambers. After a 20-minute preincubation period, fresh buffer containing a 5- or 10-fold excess of unlabeled D10 (6.25 or 12.5 mmol/L, respectively) was added to the serosal compartment (2), mucosal compartment (▨), or both compartments (▨). Mucosal permeability to FD4 was subsequently assessed in the S→M direction over an additional 1-hour incubation period, as described in Figure 1. Results are expressed as clearances, and each data point represents the mean ± SEM (n = at least 4 experiments/condition). *P < 0.001 compared with control group. Gastroenterology 2000 118, 535-543DOI: (10.1016/S0016-5085(00)70259-9) Copyright © 2000 American Gastroenterological Association Terms and Conditions

Fig. 5 FD4 clearances across rat colonic mucosa in the S→M (■) and M→S (2) directions as a function of FD4 concentration in the donor chamber. Mucosal sheets from rat colon stripped of underlying seromuscular tissue were mounted into Ussing chambers filled with K-H buffer, and graded concentrations of FD4 were added to the serosal or mucosal compartments. Permeation was measured as described in Figure 1. Gastroenterology 2000 118, 535-543DOI: (10.1016/S0016-5085(00)70259-9) Copyright © 2000 American Gastroenterological Association Terms and Conditions