Primary culture of rat taste bud cells that retain molecular markers for taste buds and permit functional expression of foreign genes

Taste buds are constituted of several kinds of cells which have distinct ch aracteristics and play different roles. In this study, we have established an in vitro culture system by optimizing the method for isolating the cells and by selecting culture media and reagents effective for cell viability a nd adhesion. As a result, the taste bud cells were adhesive and viable for over 3 days when cultured onto Matrigel-coated dishes in medium based on ke ratinocyte. growth medium. The cells retained molecular markers for both th e cytoskeleton and intracellular signaling such as cytokeratin 8 and phosph olipase C beta2. In addition, three intracellular signaling molecules, gust ducin, phospholipase C beta2, and inositol 1,4,5-trisphosphate receptor typ e 3, are expressed in the same correlation as those hi vivo, although the r atio of signaling molecule-positive cells vs. total cells was somewhat lowe r in the culture than in vivo. Next, we tried several methods to introduce foreign genes into the cells, and obtained a greater than 90% efficiency of introduction using an adenovirus vector. Finally, we show that an exogenou sly expressed myc-tagged alpha 1A-adrenoceptor sorts into the plasma membra ne, and transduces a ligand-dependent signal resulting in intracellular [Ca 2+] increase in about half of the infected cells. These results suggest that taste bud cells after 3 days of culture retain c haracteristic molecular markers, and may prove useful for describing the mo lecular and physiological features of taste bud cells, and that these cells can be further manipulated by adenovirus-mediated gene introduction. (C) 2 001 IBRO. Published by Elsevier Science Ltd. All rights reserved.