Role of intracellular Ca2+ in modulation of tight junction resistance in A6 cells.
The role of intracellular Ca2+ in the development and maintenance of epithelial tight junctional integrity is poorly understood. We assessed tight junctional resistance (Rj) in confluent monolayers of A6 cells that were treated with mucosal amiloride such that the transepithelial resistance (Rt) reflects Rj. Solution Ca2+ concentration [Ca2+] was reduced by ethylene glycol-bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid (EGTA) addition to the bathing solutions. Reduction of mucosal [Ca2+] to 1 microM or reduction of serosal Ca2+ to 100 microM did not significantly alter Rt. However, a further decrease of serosal Ca2+ to 40 microM caused the resistance to fall to < 12% of the control value. Following restoration of serosal [Ca2+], Rt increased to a new steady-state value within approximately 15 min. The magnitude of recovery of Rt was inversely correlated with the length of time the epithelium was exposed to low serosal [Ca2+]. To further test the effects of asymmetric Ca2+ removal, the serosal [Ca2+] was chelated using EGTA to reduce Rt. When the Ca2+ ionophore A-23187 was subsequently added to the mucosal solution, Rt increased from 20% to 60% of the control level. In addition, cells were loaded with the fluorescent Ca2+ indicator, Calcium Green, and the temporal relationship between changes in Rt and intracellular Ca2+ was determined. Following removal of serosal Ca2+, cell Ca2+ decreased, followed by a decrease in Rt. In contrast, returning Ca2+ to the serosal bathing solution resulted in a parallel increase of both Rt and cell [Ca2+]. These data strongly suggest that changes in intracellular [Ca2+] play an important role in the regulation of Rj.
Jovov
B
B
Department of Physiology and Biophysics, University of Texas Medical Branch, Galveston 77555.
Lewis
S A
SA
Crowe
W E
WE
Berg
J R
JR
Wills
N K
NK
D000584:Amiloride / Q000494:pharmacology*; D000818:Animals; D000001:Calcimycin / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D002460:Cell Line; D004533:Egtazic Acid / Q000494:pharmacology; D004848:Epithelium / Q000187:drug effects / Q000502:physiology; D007365:Intercellular Junctions / Q000187:drug effects / Q000502:physiology*; D007668:Kidney; D007700:Kinetics; D008433:Mathematics; D009092:Mucous Membrane / Q000187:drug effects / Q000502:physiology; D013997:Time Factors
D000001:Calcimycin; D004533:Egtazic Acid; D000584:Amiloride; D002118:Calcium