Hydrogen sulfide stimulates CFTR in Xenopus oocytes by activation of the cAMP/PKA signalling axis

Mike Althaus,Marcel Nier,Alexander Perniss,Kathrin Preiss

doi:10.1038/s41598-017-03742-5

Abstract

Hydrogen sulfide (H2S) has been recognized as a signalling molecule which affects the activity of ion channels and transporters in epithelial cells. The cystic fibrosis transmembrane conductance regulator (CFTR) is an epithelial anion channel and a key regulator of electrolyte and fluid homeostasis. In this study, we investigated the regulation of CFTR by H2S. Human CFTR was heterologously expressed in Xenopus oocytes and its activity was electrophysiologically measured by microelectrode recordings. The H2S-forming sulphur salt Na2S as well as the slow-releasing H2S-liberating compound GYY4137 increased transmembrane currents of CFTR-expressing oocytes. Na2S had no effect on native, non-injected oocytes. The effect of Na2S was blocked by the CFTR inhibitor CFTR_inh172, the adenylyl cyclase inhibitor MDL 12330A, and the protein kinase A antagonist cAMPS-Rp. Na2S potentiated CFTR stimulation by forskolin, but not that by IBMX. Na2S enhanced CFTR stimulation by membrane-permeable 8Br-cAMP under inhibition of adenylyl cyclase-mediated cAMP production by MDL 12330A. These data indicate that H2S activates CFTR in Xenopus oocytes by inhibiting phosphodiesterase activity and subsequent stimulation of CFTR by cAMP-dependent protein kinase A. In epithelia, an increased CFTR activity may correspond to a pro-secretory response to H2S which may be endogenously produced by the epithelium or H2S-generating microflora.

Highlights

Hydrogen sulfide (H2S) has been recognized as a signalling molecule which affects the activity of ion channels and transporters in epithelial cells
Human CFTR was heterologously expressed in Xenopus laevis oocytes and channel activity was measured electrophysiologically by two-electrode voltage-clamp (TEVC) microelectrode recordings
Since we were not able to stimulate CFTR activity by increasing the forskolin concentration to 30 μM, we considered the employed concentration of 5 μM as maximally effective, an observation which is consistent with a reported EC50 value of ~0.07 μM for forskolin in airway epithelia[24]

Summary

Introduction

Hydrogen sulfide (H2S) has been recognized as a signalling molecule which affects the activity of ion channels and transporters in epithelial cells. The cystic fibrosis transmembrane conductance regulator (CFTR) is an epithelial anion channel and a key regulator of electrolyte and fluid homeostasis. Na2S enhanced CFTR stimulation by membranepermeable 8Br-cAMP under inhibition of adenylyl cyclase-mediated cAMP production by MDL 12330A These data indicate that H2S activates CFTR in Xenopus oocytes by inhibiting phosphodiesterase activity and subsequent stimulation of CFTR by cAMP-dependent protein kinase A. The cystic fibrosis transmembrane conductance regulator (CFTR) is a chloride and bicarbonate conducting anion channel which is found in many vertebrate epithelia and essential for the epithelial regulation of electrolyte and fluid homeostasis. Multiple cellular signalling cascades regulate the activity or membrane abundance of CFTR8, allowing for a precise regulation of chloride flux and, eventually, electrolyte and fluid homeostasis. In addition to a high H2S metabolising capacity in epithelial cells[17], we recently hypothesised that epithelia use their electrolyte and liquid secreting machinery as a defence strategy in order to flush potentially harmful sources for H2S from the epithelial surfaces[15]

Methods

Results

Conclusion