In vitro Bovine Ciliary Body/Epithelium in a Small Continuously Perfused Ussing Type Chamber

Cell Structure and Function

Kwok-Hei Mok¹  Stuart Hodson²  Michel Millodot¹  Kam-Len Lee³  Wing-Tak Siu¹  Chi-Ho To¹  Siu-Kee Tse¹ 

[1] Laboratory of Ocular Physiology and Biochemistry, The Department of Optometry and Radiography, The Hong Kong Polytechnic University;Department of Optometry and Vision Sciences, University of Wales Cardiff;Department of Applied Biology and Chemical Technology, The Hong Kong Polytechnic University

关键词: ascorbate transport;    ciliary body;    ciliary epithelium;    ouabain;    Ussing-Zerahn chamber;   

DOI  :  10.1247/csf.23.247

学科分类：分子生物学，细胞生物学和基因

来源: Japan Society for Cell Biology

【 摘 要 】

References(14)Cited-By(2)Our goal is to assess the viability of an in vitro preparation of bovine ciliary body /epithelium (CBE) in a small volume Ussing-type chamber. A new small volume Ussing-type chamber with continuous perfusion was developed for bovine CBE. The trans-CBE electrical parameters were monitored and the electrical responses of the CBE to ouabain (1 and 0.01 mM) were recorded. The trans-CBE fluxes of [14C]-L-ascorbate and [3H]-L-glucose were also studied. The bovine CBE preparation was stable inside the chamber in terms of its potential difference (PD), short circuit current (SCC) and trans-CBE resistance. They were - 0.51 ± 0.05 mV (aqueous side negative), - 5.43±0.04 μAcm-2 and 94±2 Ω.cm2 (mean±s.c.m., n=35), respectively. The preparation hyperpolarised when 0.01 mM ouabain was administered to the aqueous side, depolarised when ouabain was applied to the stromal side. [3H]-L-glucose diffusion was about 74 nEq h-1cm-2 in either direction (n = 12). Taking the area magnification factor of the CBE into consideration, the diffusional L-glucose flux across the bovine CBE was comparable to other tight epithelia. A significant net ascorbate flux (0.26±0.05 nEq h-1cm-2, n=4, p<0.01) was found in the stroma to aqueous direction. We have developed a viable in vitro bovine CBE preparation which was (1) electrically stable, (2) responsive to ouabain, (3) tight to L-glucose diffusion, and (4) capable of actively secreting ascorbate. A net trans-CBE chloride transport (0.81±0.30 μEq h-1cm-2, n = 12, p = 0.01) from stromal to aqueous side was found in the present in vitro model under short-circuited conditions.

【 授权许可】

Unknown