Voltage Dependence of Transepithelial Guanidine Permeation Across Caco-2 Epithelia Allows Determination of the Paracellular Flux Component

Carr, Georgina
Haslam, Iain S.
Simmons, Nicholas L.

¹Institute for Cell and Molecular Biosciences, Medical School, University of Newcastle upon Tyne, Framlington Place, Newcastle upon Tyne, NE2 4HH, UK.

²To whom correspondence should be addressed. (e-mail: [email protected])

Pharmaceutical Research 23(3):p 540-548, March 2006.

Purpose

The aim of this study was to investigate transepithelial ionic permeation via the paracellular pathway of human Caco-2 epithelial monolayers and its contribution to absorption of the base guanidine.

Methods

Confluent monolayers of Caco-2 epithelial cells were mounted in Ussing chambers and the transepithelial conductance and electrical potential difference (p.d.) determined after NaCl dilution or medium Na substitution (bi-ionic conditions). Guanidine absorption (J_a-b) was measured ± transepithelial potential gradients using bi-ionic p.d.'s.

Results

Basal NaCl replacement with mannitol gives a transepithelial dilution p.d. of 28.0 ± 3.1 mV basal solution electropositive (P_Cl/P_Na = 0.34). Bi-ionic p.d.'s (basal replacements) indicate a cation selectivity of NH₄⁺ > K⁺∼Cs⁺ > Na⁺ > Li⁺ > tetraethylammonium⁺ > N-methyl-D-glucamine⁺∼ choline⁺. Transepithelial conductances show good correspondence with bi-ionic potential data. Guanidine J_a-b was markedly sensitive to imposed transepithelial potential difference. The ratio of guanidine to mannitol permeability (measured simultaneously) increased from 3.6 in the absence of an imposed p.d. to 13.8 (basolateral negative p.d.).

Conclusions

Hydrated monovalent ions preferentially permeate the paracellular pathway (Eisenman sequence 2 or 3). Guanidine may access the paracellular pathway because absorptive flux is sensitive to the transepithelial potential difference. An alternative method to assess paracellular-mediated flux of charged organic molecules is suggested.