Expression of T-Type Voltage-Gated Calcium Channel in the Cilia of Human Nasal Epithelial Cells

Abstract

<b><i>Introduction:</i></b> The mucociliary transport function of the airway epithelium is largely dependent on ciliary beating. The control signal of ciliary beating is thought to be intracellular Ca²⁺. We herein investigated the expression of T-type voltage-gated calcium channel (VGCC), a generator of intracellular Ca²⁺ oscillation, in the human nasal mucosa. <b><i>Methods:</i></b> The inferior turbinate was collected from patients with chronic hypertrophic rhinitis. The expression of T-type VGCC α1 subunits was examined by immunohistochemistry, transmission immunoelectron microscopy, Western blot, and real-time reverse transcription-polymerase chain reaction (RT-PCR). Participation of T-type VGCC in the ciliary beat regulation was examined by pharmacological inhibition tests using specific blockers of T-type VGCC in ex vivo measurements of the ciliary beat frequency (CBF) and ATP release and in intracellular Ca²⁺ imaging of isolated ciliated cells. <b><i>Results:</i></b> Immunohistochemical staining showed the expressions of T-type VGCC α1 subunits, Cav3.1 and Cav3.3, on the surface of the epithelial cells. At the ultrastructural level, immunoreactivity for Cav3.1 was localized on the surface of the cilia, and that for Cav3.3 was localized in the cilia and at the base of the cilia. The existence of Cav3.1 and Cav3.3 was confirmed at the protein level by Western blot and at the transcriptional level by real-time RT-PCR. Specific blockers of T-type VGCC, mibefradil and NNC 55-0396, significantly inhibited CBF. These blockers also inhibited a CBF increase induced by 8-bromo-cAMP/8-bromo-cGMP and significantly lowered the intracellular Ca²⁺ level of isolated ciliated cells in a time-dependent manner. On the other hand, the ATP release from the nasal mucosa was not changed by mibefradil or NNC 55-0396. <b><i>Conclusion:</i></b> These results indicate that T-type VGCC α₁ subunits, Cav3.1 and Cav3.3, exist at the cilia of the nasal epithelial cells and participate in the regulation of ciliary beating and that these channels act downstream of cAMP/cGMP.