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Alpha-Adrenergic Agonists Stimulate Fluid Secretion in Lacrimal Gland Ducts.

ABSTRACT: Purpose:The role of adrenergic innervation in the regulation of lacrimal gland (LG) ductal fluid secretion is unknown. The Aim of the present study was to investigate the effect of adrenergic stimulation on fluid secretion in isolated LG duct segments and to study the underlying intracellular mechanisms.

Methods:Fluid secretion of isolated mouse LG ducts was measured using video-microscopy. Effect of various adrenergic agonists (norepinephrine, phenylephrine, and isoproterenol) on fluid secretion as well as inhibitory effects of specific antagonists on adrenergic agonist-stimulated secretory response were analyzed. Changes in intracellular Ca2+ level [Ca2+i] were investigated with microfluorometry.

Results:Both norepinephrine and phenylephrine initiated a rapid and robust fluid secretory response, whereas isoproterenol did not cause any secretion. Phenylephrine-induced secretion was completely blocked by ?1D-adrenergic receptor blocker BMY-7378. The endothelial nitric oxide synthase (eNOS) inhibitor L-NAME or guanylyl cyclase inhibitor ODQ reduced but not completely abolished the phenylephrine-induced fluid secretion, whereas co-administration of Ca2+-chelator BAPTA-AM resulted in a complete blockade. Phenylephrine stimulation induced a small, but statistically significant elevation in [\(Ca_i^{2 + }\)].

Conclusions:Our results prove the direct role of ?1-adrenergic stimulation on LG ductal fluid secretion. Lack of isoproterenol-induced fluid secretory response suggests the absence of ?-receptor mediated pathway in mouse LG ducts. Complete blockade of phenylephrine-induced fluid secretion by BMY-7378 and predominant inhibition of the secretory response either by L-NAME or ODQ suggest that ?-adrenergic agonists use the NO/cGMP pathway through ?1D receptor. Ca2+ signaling independent from NO/cGMP pathway may also play an at least partial role in ?-adrenergic induced ductal fluid secretion.

SUBMITTER: Szarka D

PROVIDER: S-EPMC7718821 | biostudies-literature | 2020 Dec

REPOSITORIES: biostudies-literature

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Alpha-Adrenergic Agonists Stimulate Fluid Secretion in Lacrimal Gland Ducts.

Szarka Dóra D Elekes Gréta G Berczeli Orsolya O Vizvári Eszter E Szalay László L Ding Chuanqing C Tálosi László L Tóth-Molnár Edit E

Investigative ophthalmology & visual science 20201201 14

<h4>Purpose</h4>The role of adrenergic innervation in the regulation of lacrimal gland (LG) ductal fluid secretion is unknown. The Aim of the present study was to investigate the effect of adrenergic stimulation on fluid secretion in isolated LG duct segments and to study the underlying intracellular mechanisms.<h4>Methods</h4>Fluid secretion of isolated mouse LG ducts was measured using video-microscopy. Effect of various adrenergic agonists (norepinephrine, phenylephrine, and isoproterenol) on ...[more]

PMID: 33259608

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Project description:BackgroundAqueous-deficient dry eye disease (ADDED) resulting from dysfunction of the lacrimal gland (LG) is currently incurable. Although LG stem/progenitor cell-based therapy is considered to be a promising strategy for ADDED patients, the lack of a reliable serum-free culture method to obtain enough lacrimal gland stem cells (LGSCs) and the basic standard of LGSC transplantation are obstacles for further research.MethodsAdult mouse LGSCs were cultured in Matrigel-based 3D culture under serum-free culture condition, which contained EGF, FGF10, Wnt3A, and Y-27632. LGSCs were continuously passaged over 40 times every 7 days, and the morphology and cell numbers were recorded. LGSCs were induced to differentiate to ductal cells by reducing Matrigel rigidity, while fetal bovine serum was used for the induction of acinar cells. RT-PCR or qRT-PCR analysis, RNA-sequence analysis, H&E staining, and immunofluorescence were used for characterization and examining the differentiation of LGSCs. LGSCs were allotransplanted into diseased LGs to examine the ability of repairing the damage. The condition of eye orbits was recorded using a camera, the tear production was measured using phenol red-impregnated cotton threads, and the engraftments of LGSCs were examined by immunohistochemistry.ResultsWe established an efficient 3D serum-free culture for adult mouse LGSCs, in which LGSCs could be continuously passaged for long-term expansion. LGSCs cultured from both the healthy and ADDED mouse LGs expressed stem/progenitor cell markers Krt14, Krt5, P63, and nestin, had the potential to differentiate into acinar or ductal-like cells in vitro and could engraft into diseased LGs and relieve symptoms of ADDED after orthotopic injection of LGSCs.ConclusionWe successfully established an efficient serum-free culture for adult mouse LGSCs aiming at LG repair for the first time. Our approach provides an excellent theoretical and technical reference for future clinical research for ADDED stem cell therapy.

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Alpha-Adrenergic Agonists Stimulate Fluid Secretion in Lacrimal Gland Ducts.

Publications

Alpha-Adrenergic Agonists Stimulate Fluid Secretion in Lacrimal Gland Ducts.

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