T. Shimizu et al., Ca2+-sensing receptor-mediated regulation of volume-sensitive Cl- channelsin human epithelial cells, J PHYSL LON, 528(3), 2000, pp. 457-472
1. Since extracellular Ca2+ or Mg2+ has been reported to modulate swelling-
activated Cl- currents, we examined the expression of the G protein-coupled
Ca2+-sensing receptor (CaR) and its involvement in the regulation of volum
e-sensitive Cl- channels in a human epithelial cell line (Intestine 407).
2. Reverse transcriptase-polymerase chain reaction and immunoblotting analy
sis showed that Intestine 407 cells express CaR mRNA and protein.
3. The swelling-activated whole-cell Cl- current was voltage-independently
augmented by extracellular Ca2+ or Mg2+. In addition, Ca2+ or Mg2+ voltage-
dependently accelerated the inactivation kinetics of the Cl- current.
4. Neomycin, spermine and La3+ augmented volume-sensitive Cl- currents. How
ever, these CaR agonists failed to affect depolarization-induced inactivati
on.
5. Intracellular application of GTP gammaS, but not GDP betaS, increased th
e amplitude of the swelling-induced Cl- current without affecting the basal
current. The upregulating effect of Ca2+ on the Cl- current amplitude was
abolished by either GTP gammaS or GDP betaS. In contrast, GTP gammaS and GD
P betaS failed to affect the inactivation kinetics of the Cl- current and t
he accelerating effect of Ca2+ thereon.
6. The Cl- current amplitude was enlarged by stimulation with forskolin, di
butyryl cAMP and IBMX. During the cAMP stimulation, extracellular Ca2+ fail
ed to increase the Cl- current but did accelerate depolarization-induced in
activation.
7. It is concluded that stimulation of the CaR induces upregulation of volu
me-sensitive Cl- channels via a G: protein-mediated increase in intracellul
ar cAMP in the human epithelial cell. However, the accelerating effect of e
xtracellular divalent cations on the inactivation kinetics of the Cl- curre
nt is induced by a mechanism independent of the CaR and cAMP.