Murine endolymphatic sac development in tissue culture: An in vitro model for sac function

Abstract

Numerous studies have attempted to elucidate the function of the mammalian endolymphatic sac (ELS). All of these studies have been performed on in vivo specimens and are thus influenced by humoral and tissue factors extraneous to the sac. In contrast, an in vitro model would provide an opportunity to study ELS cells in a carefully controlled environment. This report presents our experience with tissue culturing the murine endolymphatic sac removed from 16 and 18 gestational day fetuses. Light (LM) and transmission electron microscopical (TEM) evaluations of the developing endolymphatic sac were performed over periods of one, four, and seven days in tissue culture. In order to confirm growth and maturation, three-dimensional reconstructions from serial sections of the cultured ELS were made and compared with published accounts of in vivo murine ELS development for equivalent periods of time. Both whole and dissected otocysts were grown in tissue culture and compared with one another. Two different tissue culture medias were investigated, each with and without the addition of collagenase, used to soften the dense fibrous capsule of the otocyst and thus facilitate dissection and histological preparation. The impact of collagenase and the tissue culture medias on endolymphatic sac growth were studied. Results demonstrated that murine ELS cells were able to differentiate and mature in tissue culture, as confirmed by LM, TEM, and three-dimensional reconstructions. After an initial delay, in vitro maturation of cells in tissue culture paralleled normal in vivo growth and in some specimens appeared to show accelerated maturation. This in vitro model should prove useful in efforts to define ELS function and in providing a technique for tissue culturing human ELS from normal and diseased ears.