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Edit Toth-Molnar, Dora Szarka, Orsolya Berczeli, Eszter Vizvari, Zoltan Rakonczay, Peter Hegyi, Chuanqing Ding; Intracellular pathways of α1 adrenergic stimulation-evoked fluid secretion in isolated lacrimal gland ducts in mice. Invest. Ophthalmol. Vis. Sci. 2018;59(9):4923.
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© ARVO (1962-2015); The Authors (2016-present)
Recently we found that α1 adrenergic agonist phenylephrine (PHE) resulted in a robust fluid secretion in isolated lacrimal gland (LG) ducts in mice. Earlier reports demonstrated the involvement of the NO/cGMP pathway in the PHE-induced protein secretion of LG acinar cells from rat, but the intracellular pathways used by α1 adrenergic agonists to stimulate ductal fluid secretion is unknown. Therefore the aim of the present study was to investigate the intracellular mechanisms underlying α1 adrenergic-stimulated LG ductal fluid secretion in mouse.
LG duct segments were isolated from mouse LG as we previously described. Ends of the ducts seal forming a closed intraluminal space during incubation. Fluid secretion as a swelling response can be quantified by video-microscopy. Effects of endothelial nitric oxide synthase (eNOS) inhibitor L-NAME, neuronal nitric oxide synthase (nNOS) inhibitor S-methyl-L-thiocitrulline, guanylate cyclase (GC) inhibitor ODQ and α1D-adrenergic receptor inhibitor BMY-7378 were investigated in the PHE-induced ductal fluid secretion. Changes of intracellular Ca2+ concentration ([Ca2+]i) after PHE stimulation was measured with microfluorometry using FURA 2AM. Data were expressed as the x-fold change of the PHE-induced luminal volume (LV) increase, which was standardized to 1.0. Data were presented as means ± SEM.
PHE (10 µM) stimulation resulted in a significant LV change (1.64±0.28-fold increase, p=0.004). PHE-evoked ductal fluid secretion was dose-dependently reduced by L-NAME (max. inhibition at 10 µM: 73.06±10.5% decrease in LV, p=0.006) while inhibition of nNOS had no effect on the fluid secretory response (p=0.21). ODQ decreased PHE-induced LV increase (max. inhibition at 1 µM: 72.1± 12.1 % decrease in LV, p=0.005). BMY-7378 dose-dependently reduced ductal fluid secretion (max. inhibition at 1 µM: 80.06±12.7%. p=0.007). PHE stimulation resulted in a small, but statistically significant increase in [Ca2+]i (7.51± 1.07%, p=0.001) in isolated LG ducts.
LV increase induced by PHE was reduced by α1D adrenergic receptor blockage, or by inhibition of either eNOS or GC. These data suggest that α1-adrenergic agonists use the NO/cGMP pathway through α1D receptor stimulation to increase fluid secretion in isolated mouse LG ducts. The functional relevance of the PHE-induced small elevation in [Ca2+]i needs further investigation.
This is an abstract that was submitted for the 2018 ARVO Annual Meeting, held in Honolulu, Hawaii, April 29 - May 3, 2018.
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