Planarian cell culture: a comparative review of methods and an improved protocol for primary cultures of neoblasts

To develop an improved method for preparing and cultivating planarian cells , several protocols published previously were compared with each other. Cel ls, and in particular neoblasts, proved remarkably resistant to hyposmotic conditions. However, survival periods depended critically on the content in nutrients and on osmotic conditions. Starting from an optimized method to disintegrate planarian tissues and prepare purified neoblast fractions, dif ferent media and additives were tried. Hyposmotic media and layers of extra cellular matrix components enhanced the adhesion of neoblasts and favoured the formation of transient processes. Proteins in the medium supported long -term survival of neoblasts that retained a spherical shape. Eventually, an isosmotic medium was devised that supported the survival of neoblasts with a viability of 46% on day 31 of primary cultures. With light microscopical techniques, no signs of differentiation were observed in these cultures. M itoses were detected until the second day of cultivation. In contrast, cult ures of total cells still displayed mitoses after 7 days of cultivation. So me guidelines are proposed for future research directed towards establishin g permanent neoblast lines.