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Identification of unitary tight junction pore opening/closing events in MDCK monolayers
Author(s) -
Weber Christopher R,
Liang Guo Hua,
Shen Le,
Wang Yitang,
Nelson Deborah J,
Yu Alan S,
Turner Jerrold R
Publication year - 2013
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.27.1_supplement.256.1
Subject(s) - claudin , tight junction , paracellular transport , chemistry , barrier function , occludin , conductance , biophysics , ion channel , cysteine , patch clamp , permeability (electromagnetism) , microbiology and biotechnology , physics , biology , biochemistry , condensed matter physics , receptor , membrane , enzyme
Analyses of epithelial barrier function have led to the hypothesis that tight junctions are populated by claudin‐dependent pores that define paracellular ion conductance and charge selectivity. Consistent with this claudin‐2 expression increases paracellular Na + permeability of MDCK I monolayers. However, available measures cannot resolve function of individual pores. To overcome this limitation, we have used patch clamp techniques to define local tight junction function. Local tight junction recordings reveal transient 63±8 pS openings (0.004±0.001 Hz). Claudin‐2 expression reduces global TER by a 71±1% and increases the frequency of individual openings to 0.023±0.007 Hz without changing their size (57±3 pS). Ion substitution analyses demonstrate that the openings are selective for cations (P Na /P Cl = 4.9 ± 0.9) with radii <2.8Å. Global analyses suggest that flux across claudin‐2 I66C pores is blocked by cysteine derivatization. Consistent with this, while claudin‐2 I66C expression had effects similar to claudin‐2 expression, openings in MDCK I expressing claudin‐2 I66C were acutely terminated upon cysteine derivatization. These data show that claudin‐2 creates local, transient tight junction flux events that define global epithelial barrier function. We suggest that opening and closing of paracellular claudin pores forms the molecular basis of tight junction size and charge selectivity.