The perinuclear centriole-containing centrosome is not the major microtubule organizing center in Sertoli cells

Microtubules are abundant in Sertoli cells and are predominantly arranged parallel to the long axis of the cell. In addition, the centrioles occur in a perinuclear position in the basal one third of the cell. In this study we investigate the importance of the perinuclear centriole-containing centrosome as a microtubule organizing center (MTOC) in Sertoli cells. In all experiments, rat testes were perfusion fixed and then processed for electron and/or fluorescence microscopy. For fluorescence microscopy, fragments or dissected pieces of seminiferous epithelium were labeled for tubulin and actin. The three-dimensional pattern of microtubules in Sertoli cells was determined using data collected with a confocal microscope and analyzed using the NIH-Image program (written by Wayne Rasband at the NIH). The detailed arrangement of microtubules around, and the number of microtubule ends associated with, the centrosome were determined from composite projections constructed from serial thin sections. The nucleating potential of the perinuclear centrosome was determined by perfusing testes for 6 h with 10 micrograms/ml nocodazole and then for up to 3 h with control buffer prior to fixation and analysis with confocal and standard fluorescence microscopy. Microtubules are not organized around a focal perinuclear site and few microtubule ends are associated with the centrosome. Moreover, in cells recovering from nocodazole treatment, microtubules first appear in apical (peripheral) processes. Our data indicate that the centriole-containing perinuclear centrosome is not a significant MTOC in Sertoli cells. Rather, microtubules are nucleated in peripheral regions and project basally. Based on the observations that microtubules appear to "cuff" the nucleus, intermediate filaments are concentrated around the nucleus, microtubules project into the perinuclear intermediate filament network, and microtubules and intermediate filaments are often coaligned, we suggest that microtubules are anchored into position at the base of the cell via linkages with the intermediate filament network. Our nucleation-anchorage model of microtubule organization in Sertoli cells may be applicable to other epithelial systems.