G protein-coupled estrogen receptor promotes acrosome reaction via regulation of Ca2+ signaling in mouse sperm†.

G protein-coupled estrogen receptor (GPER), a seven-transmembrane G protein-coupled receptor, mediates the rapid pre-genomic signaling actions of estrogen and derivatives thereof. The expression of GPER is extensive in mammal male reproductive system. However, the functional role of GPER in mouse sperm has not yet been well recognized. This study revealed that GPER was expressed at the acrosome and the mid-flagellum of the mouse sperm. The endogenous GPER ligand 17ß-estradiol and the selective GPER agonist G1 increased intracellular Ca2+ concentration ([Ca2+]i) in mouse sperm, which could be abolished by G15, an antagonist of GPER. In addition, the G1-stimulated Ca2+ response was attenuated by interference with the phospholipase C (PLC) signaling pathways or by blocking the cation channel of sperm (CatSper). Chlortetracycline staining assay showed that the activation of GPER increased the incidence of acrosome-reacted sperm. Conclusively, GPER was located at the acrosome and mid-flagellum of the mouse sperm. Activation of GPER triggered the elevation of [Ca2+]i through PLC-dependent Ca2+ mobilization and CatSper-mediated Ca2+ influx, which promoted the acrosome reaction of mouse sperm.

Increased intracellular Cl- concentration promotes ongoing inflammation in airway epithelium.

Airway epithelial cells harbor the capacity of active Cl- transepithelial transport and play critical roles in modulating innate immunity. However, whether intracellular Cl- accumulation contributes to relentless airway inflammation remains largely unclear. This study showed that, in airway epithelial cells, intracellular Cl- concentration ([Cl-]i) was increased after Pseudomonas aeruginosa lipopolysaccharide (LPS) stimulation via nuclear factor-κB (NF-κB)-phosphodiesterase 4D (PDE4D)-cAMP signaling pathways. Clamping [Cl-]i at high levels or prolonged treatment with LPS augmented serum- and glucocorticoid-inducible protein kinase 1 (SGK1) phosphorylation and subsequently triggered NF-κB activation in airway epithelial cells, whereas inhibition of SGK1 abrogated airway inflammation in vitro and in vivo. Furthermore, Cl--SGK1 signaling pathway was pronouncedly activated in patients with bronchiectasis, a chronic airway inflammatory disease. Conversely, hydrogen sulfide (H2S), a sulfhydryl-containing gasotransmitter, confers anti-inflammatory effects through decreasing [Cl-]i via activation of cystic fibrosis transmembrane conductance regulator (CFTR). Our study confirms that intracellular Cl- is a crucial mediator of sustained airway inflammation. Medications that abrogate excessively increased intracellular Cl- may offer novel targets for the management of airway inflammatory diseases.

Cellular Mechanism Underlying Hydrogen Sulfide Mediated Epithelial K+ Secretion in Rat Epididymis.

As a novel gasotransmitter, hydrogen sulfide (H2S) elicits various physiological actions including smooth muscle relaxation and promotion of transepithelial ion transport. However, the pro-secretory function of H2S in the male reproductive system remains largely unclear. The aim of this study is to elucidate the possible roles of H2S in modulating rat epididymal intraluminal ionic microenvironment essential for sperm storage. The results revealed that endogenous H2S-generating enzymes cystathionine ß-synthetase (CBS) and cystathionine γ-lyase (CSE) were both expressed in rat epididymis. CBS located predominantly in epithelial cells whilst CSE expressed primarily in smooth muscle cells. The relative expression level of CBS and CSE escalated from caput to cauda regions of epididymis, which was paralleled to the progressively increasing production of endogenous H2S. The effect of H2S on epididymal epithelial ion transportation was investigated using short-circuit current (I SC), measurement of intracellular ion concentration and in vivo rat epididymal microperfusion. Our data showed that H2S induced transepithelial K+ secretion via adenosine triphosphate-sensitive K+ (KATP) channel and large conductance Ca2+-activated K+ (BKCa) channel. Transient receptor potential vanilloid 4 (TRPV4) channel-mediated Ca2+ influx was implicated in the activation of BKCa channel. In vivo studies further demonstrated that H2S promoted K+ secretion in rat epididymal epithelium. Inhibition of endogenous H2S synthesis caused a significant decrease in K+ concentration of cauda epididymal intraluminal fluid. Moreover, our data demonstrated that high extracellular K+ concentration actively depressed the motility of cauda epididymal sperm in a pH-independent manner. Collectively, the present study demonstrated that H2S was vital to the formation of high K+ concentration in epididymal intraluminal fluid by promoting the transepithelial K+ secretion, which might contribute to the maintenance of the cauda epididymal sperm in quiescent dormant state before ejaculation.

Functional expression of G protein-coupled receptor 30 in immature rat epididymal epithelium.

The aim of this study is to investigate the functional role of G protein-coupled receptor 30 (GPR30) in the epididymis. We found that GPR30 is expressed in the epithelium of the immature rat epididymis and is involved in chloride secretion into the caudal epididymis lumen. The short-circuit current (Isc) experiments showed that in primary cultured caudal epididymis epithelium, activation of GPR30 by its specific agonist G1 induced a mono-phasic current increase, and G15, the specific antagonist of GPR30, could completely inhibit the current induced by G1. The G1-induced Isc was largely blocked by application of the non-specific chloride channel inhibitor diphenylamine-dicarboxylic acid (DPC), or by the cystic fibrosis transmembrane conductance regulator (CFTR) inhibitor CFTRinh-172 , suggesting that the current was mainly mediated through CFTR. In addition, after stimulating GPR30 by G1, the intracellular concentration of cAMP in the epithelium was significantly increased, indicating that the cAMP signal pathway is involved and could be responsible for the CFTR activation. Finally, to further investigate the function of GPR30 in vivo, G15 was administrated into rats subcutaneously. The osmotic pressure of the micro perfusion solution from epididymis was measured and the sperms were collected. Results showed that there was an osmotic pressure increase of the perfusion solution from G15 treated rats. When the GPR30 was inhibited by G15 endogenously, the motility of sperms decreased. Our data demonstrated that GPR30 is involved in the formation of caudal epididymis fluid micro-environment thus affecting sperm motility.