Figure 5. Fluorescence images of Sertoli cells stained with quinacrine.

Quinacrine stains individual secretory vesicles in the cell cytoplasm. Sertoli cells in culture were incubated with 3 µM quinacrine for 30 min, washed and photographed under fluorescence illumination immediately (A and C) and at 1 min intervals for 10 min of incubation in the absence or presence of rT₃, respectively (B and D). Incubation of cells with 10^-17 M rT₃ caused fusion of quinacrine-loaded vesicles to the plasma membrane and release of the fluorescent content into the surrounding medium, as seen by the loss of fluorescence from most vesicles located at the cell periphery. This effect was observed after 4 min incubation with rT₃. Also, Sertoli cells were incubated for 10 min with 500 nM of RGD peptide or 1 µM of flunarizine prior to incubation with quinacrine, washed and incubated with 3 µM quinacrine for 30 min. Quinacrine-loaded Sertoli cell cultures, pre-treated with RGD or flunarizine for 10 min, were incubated in the absence or presence of 10^-17 M rT₃ and photographed under fluorescence illumination immediately (E, G, I and K) and at 1-min intervals for 10 min of incubation in the absence or presence of rT₃ (F, H, J and L). Incubation of cells in the presence of 500 nM RGD peptide or 1 µM flunarizine prevented the fusion of quinacrine-loaded vesicles to the plasma membrane and release of the fluorescent content. (A) Control, 0 min. (B) Control, 4 min. (C) rT₃, 0 min. (D) rT₃, 4 min. (E) RGD, 0 min. (F) RGD, 4 min. (G) rT₃ + RGD, 0 min. (H) rT₃ + RGD, 4 min. (I) Flunarizine, 0 min. (F) Flunarizine, 4 min. (G) rT₃ + Flunarizine, 0 min. (H) rT₃ + Flunarizine, 4 min. Experiments were performed 3 times with similar results. Bar = 10 µm.