CELLULAR HETEROGENEITY IN THE MEMBRANA GRANULOSA OF DEVELOPING RAT FOLLICLES - ASSESSMENT BY FLOW-CYTOMETRY AND LECTIN-BINDING

The hormone-mediated maturation of ovarian follicles is apparently acc ompanied by position-specific differentiation of cells of the membrana granulosa. We have assessed the extent of this cellular heterogeneity by flow cytometry using a variety of fluorescein iso-thiocyanate-labe led lectins as probes. Follicular development was stimulated in immatu re rats by treatment with either diethylstilbestrol (DES) or equine CG (eCG). Lectin binding to monodispersed rat granulosa cells was then a nalyzed by flow cytometry. Our results demonstrate that there are two distinct populations of small (4-7 mu m) and large (9-12 mu m) granulo sa cells in follicles from DES- and eCG-treated animals. Both populati ons appear to be mitotically active and show specific lectin-binding c haracteristics. Six lectins (canavalia ensiforms, triticum vulgaris, m aclura pomifera, erythrina cristagalli, jacalin, and vicia villosa) bi nd equally to both small and large granulosa cells from the DES- and e CG-treated rats. In contrast, no binding to either cell population was detected with six other lectins (dolichos biflorus, griffonia simplic ifolia-II, lycopersicon esculentum, datura stramonium, solanum tuberos um, and ulex europaeus). Furthermore, four galactose-binding lectins ( bauhinia purpurea, glysine maximus, griffonia simplicifolia-I, and ara chis hypogaea) were found to identify specific subsets of granulosa ce lls. Three of these lectins (bauhinia purpurea, glysine maximus, and g riffonia simplicifolia-I) bind to only small granulosa cells from eith er DES- or eCG-treated immature rats. The fourth lectin (arachis hypog aea) identifies subpopulations of both small and large granulosa cells . Application of the four galactose-specific lectins to fixed sections of frozen ovaries demonstrated binding to the perioocyte and cumulus granulosa cells. We conclude that cellular heterogeneity exists within the follicular epithelium at various stages of follicular development with respect to cell size and expression of specific lectin-binding s ites.