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Imada Toshihiro, Kai Jin, Yusuke Izuta, Shigeru Nakamura, Takahiro Adachi, Kazuo Tsubota; In vivo dynamics of Ca2+-dependent myoepithelial contraction in lacrimal gland. Invest. Ophthalmol. Vis. Sci. 2017;58(8):2253.
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© ARVO (1962-2015); The Authors (2016-present)
Lacrimal gland (LG) myoepithelial cells (MECs) are distributed surrounding the acinar cells. It has been reported that intracellular Ca2+ ([Ca2+]i)-dependent myoepithelial contraction plays a role in squeezing the acinar cells and expelling tear components within isolated LG alone. In this study, we verified the relationship between [Ca2+]i and myoepithelial contraction in vivo Ca2+ dynamics imaging together with the YC3.60 transgenic mice (a mouse lines expressing Ca2+ sensor yellow cameleon 3.60, which is established based on the Cre/loxP system under control of a CAG promoter).
Eight-week-old female YC3.60 transgenic mice were used. Mice were placed in a prone position, and their LG were exposed by skin incision on the right temporal side of the head under anesthesia. In vivo FRET (fluorescence resonance energy transfer) ratio imaging of LG was performed by two-photon microscopy in order to visualize the Ca2+ dynamics. The LG was excited at 840 nm, and the fluorescence emission of CFP and YFP were detected at 460-500 nm and 520-560 nm, respectively. The images were acquired at approximately 1 frame/sec. To calculate the FRET ratio, the fluorescence images of each emission wavelength were analyzed by the software FluoView FV1200MPE. The area surrounded by MECs is measured by Image J software. The changes in FRET ratio and myoepithelial contraction after intravenous injection of 1 mg/kg pilocarpine, cholinergic tear secretion stimulant, were evaluated.
YC3.60 probe was specifically expressed in the LG MECs, not in acinar cells, of YC3.60 transgenic mice. Pilocarpine injection increased the FRET ratio in the MECs, and their surrounded area was immediately decreased after increase in FRET ratio. The area surrounded by MECs decreased to approximately 80% of that of before pilocarpine injection.
Our study demonstrated that [Ca2+]i-dependent myoepithelial contraction can be visualized in living animal. It suggested that YC-3.60 transgenic mouse are useful for elucidation the physiological role of MECs in tear secretion in living animal.
This is an abstract that was submitted for the 2017 ARVO Annual Meeting, held in Baltimore, MD, May 7-11, 2017.
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