β1- and β2-adrenergic stimulation-induced electrogenic transport by human endolymphatic sac epithelium and its clinical implications

Abstract

The endolymphatic sac (ES) is a cystic structure of the inner ear connected to the cochlea and vestibule, which plays a role in regulating ion homeostasis in inner ear fluid. Disruption of ion homeostasis can cause inner ear disorders with hearing loss and dizziness, such as Meniere's disease. Herein, we found, for the first time, functional evidence for the involvement of β₁- and β₂-adrenergic receptors in apical electrogenic ion transport by human ES epithelium by using electrophysiological/pharmacological and molecular biological methods, which were dependent on K⁺ and Cl^- ion transport. The apical electrogenic transport was absent or very weak in ES epithelia of patients with Meniere's disease. These results suggested that adrenergic stimulation via β₁- and β₂-adrenergic receptors in the human ES was involved in regulation of inner ear fluid ion homeostasis and impairment of this response could be a pathological mechanism of Meniere's disease.

Figure 1. Schematic figure of inner ear structures, and ion composition of the endolymph in each compartment.

The black circular areas in the vestibule and cochlear duct contain sensory epithelia.

Figure 2. Full length gel images of transcript expression of β₁- and β₂-adrenergic receptors.

(a) Receptor transcript expression in a positive control (human heart). (b) Receptor transcript expression in the human endolymphatic sac.

Figure 3. Isoproterenol (10 μM)-induced electrogenic transport by the human endolymphatic sac epithelium.

(a) Representative figure of isoproterenol-induced cation absorption/anion secretion current (type A). (b) Representative figure of isoproterenol-induced cation secretion/anion absorption current (type B). (c) Representative figure showing no effect of isoproterenol in the electrogenic transport (type C). Tissue viability was assessed by the detection of the current change after Ba²⁺ (1 mM) application. (d) Mean amount of current change induced by isoproterenol in type A, type B, and type C. IPN, isoproterenol.

Figure 4. Inhibition of isoproterenol-induced electrogenic transport by Ba²⁺ (1 mM) in human endolymphatic sac epithelium.

(a) Representative figure showing the effect of Ba²⁺ in type A current. (b) Representative figure showing the effect of Ba²⁺ in type B current. (c) Mean inhibitory effect of Ba²⁺ for isoproterenol-induced trans-epithelial current. IPN, isoproterenol. *p < 0.05.

Figure 5. Changes in electrogenic transport induced by DIDS (100 μM) in the human endolymphatic sac epithelium.

(a) Representative figure of DIDS-induced cation absorption/anion secretion current. (b) Representative figure of DIDS-induced cation secretion/anion absorption current. (c) Representative figure showing no effect of DIDS on electrogenic transport. DIDS, 4,4′-diisothiocyano-2,2′-stilbenedisulfonic acid.

Figure 6. Inhibition of isoproterenol-induced electrogenic transport by DIDS (100 μM) in the human endolymphatic sac epithelium.

(a) Representative figure showing the effect of DIDS on type A current. (b) Representative figure showing the effect of DIDS on type B current. (c) Mean inhibitory effect of DIDS for isoproterenol-induced trans-epithelial current. IPN, isoproterenol; DIDS, 4,4′-diisothiocyano-2,2′-stilbenedisulfonic acid; *p < 0.05.

Figure 7. Effect of selective β-adrenergic receptor blockers on isoproterenol-induced electrogenic transport.

(a and b). Representative figures of the inhibitory effects of β₁-adrenergic receptor blocker (CGP20712A) on type A and B isoproterenol-induced electrogenic transport. (c). Representative figure showing the inhibitory effect of β₂-adrenergic receptor blocker (ICI18551) on type A isoproterenol-induced electrogenic transport. (d) Mean inhibitory effects of CGP20712A and ICI18551 on isoproterenol-induced electrogenic transport. *p < 0.05.

Figure 8. Effect of isoproterenol on electrogenic transport by human endolymphatic sac epithelia inpatients with Meniere’s disease.

(a) Representative figure showing no effect of isoproterenol on electrogenic transport. (b) Representative figure of isoproterenol-induced small type B current. (c) Differences in the presence of isoproterenol-induced electrogenic transport between disease-free controls and patients with Meniere’s disease. IPN, isoproterenol; numbers in the bar graph represent the number of samples; *p < 0.05.

Figure 9. Protein expression of β₁- and β₂-adrenergic receptors in the human endolymphatic sac epithelium of disease-free controls and patients with Meniere’s disease.

AR, adrenergic receptors; Cont, disease-free controls; MD, Meniere’s disease; β1, β₁-adrenergic receptor; β2, β₂-adrenergic receptor. Scale bar in the left uppermost figure indicates 50 μm.