Ca²⁺-sensing receptor-mediated regulation of volume-sensitive Cl^- channels in human epithelial cells

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 volume-sensitive Cl^- channels in a human epithelial cell line (Intestine 407). 2. Reverse transcriptase-polymerase chain reaction and immunoblotting analysis 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. However, these CaR agonists failed to affect depolarization-induced inactivation. 5. Intracellular application of GTPγS, but not GDPβS, increased the 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 GTPγS or GDPβS. In contrast, GTPγS and GDPβS failed to affect the inactivation kinetics of the Cl^- current and the accelerating effect of Ca²⁺ thereon. 6. The Cl^- current amplitude was enlarged by stimulation with forskolin, dibutyryl cAMP and IBMX. During the cAMP stimulation, extracellular Ca²⁺ failed to increase the Cl^- current but did accelerate depolarization-induced inactivation. 7. It is concluded that stimulation of the CaR induces upregulation of volume-sensitive Cl^- channels via a G protein-mediated increase in intracellular cAMP in the human epithelial cell. However, the accelerating effect of extracellular divalent cations on the inactivation kinetics of the Cl^- current is induced by a mechanism independent of the CaR and cAMP.