β₁-Adrenergic Receptors but not β₂-Adrenergic or Vasopressin Receptors Regulate K⁺ Secretion in Vestibular Dark Cells of the Inner Ear

Abstract.

Receptors were identified pharmacologically in functional studies where K⁺ secretion was monitored as transepithelial current (I _sc ). Further, receptors were identified as transcripts by cloning and sequencing of reverse-transcriptase polymerase chain reaction (RT-PCR) products. I _sc under control conditions was 796 ± 15 μA/cm² (n= 329) in gerbilline VDC and 900 ± 75 μA/cm² (n= 6) in murine VDC. Forskolin (10⁻⁵ m) but not 1,9-dideoxy-forskolin increased I _sc by a factor of 1.42 ± 0.05 (n= 7). 10⁻⁹ m Arg⁸-vasopressin and 10⁻⁹ m desmopressin had no significant effect in gerbilline and murine VDC. Isoproterenol, norepinephrine, epinephrine and prenalterol stimulated I _sc maximally by a factor of 1.38 ± 0.04 (n= 7), 1.59 ± 0.06 (n= 6), 1.64 ± 0.03 (n= 8) and 1.37 ± 0.03 (n= 6), respectively. The EC ₅₀ values were (1.4 ± 0.7) × 10⁻⁸ m (n= 36), (2.5 ± 1.0) × 10⁻⁸ m (n= 31), (1.7 ± 0.7) × 10⁻⁷ m (n= 36) and (5 ± 4) × 10⁻⁷ m (n= 32), respectively. Propanolol inhibited isoproterenol-induced stimulation of I _sc . Atenolol, ICI118551 and CGP20712A inhibited isoproterenol-induced stimulation of I _sc with a pK_DB of 5.0 × 10⁻⁸ m (pK _DB = 7.30 ± 0.07, n= 38), 4.4 × 10⁻⁸ m (pK _DB = 7.36 ± 0.14, n= 37) and 6.8 × 10⁻¹² m (pK _DB = 11.17 ± 0.12, n= 37), respectively. RT-PCR of total RNA isolated from microdissected vestibular labyrinth tissue using specific primers revealed products of the predicted sizes for β₁- and β₂-adrenergic receptors but not for β₃-adrenergic receptors. Sequence analysis of the amplified cDNA fragments from gerbilline tissues revealed a 96.4%, 91.5% and 89.6% identity compared to rat β₁-, β₂- and β₃-adrenergic receptors, respectively. These results demonstrate that K⁺ secretion in VDC is under the control of β₁- but not β₂- or β₃-adrenergic receptors or vasopressin-receptors.