FIXATION INDUCES DIFFERENTIAL POLARIZED TRANSLOCATIONS OF ORGANELLES IN HYPHAE OF SAPROLEGNIA-FERAX

Saprolegnia hyphal tips were examined during fixation, and living or freeze-substituted tips were quantitatively compared with those fixed in commonly employed formulations of paraformaldehyde and glutaraldehyde. Treating hyphae with fixatives induced extensive longitudinal translocations of the cytoplasm and organelles, usually beginning with contractions toward the tip. These translocations were minimal in the extreme apex (approximately 10-mu-m) and more extensive subapically. Hypertonic media or hypotonic buffers seldom or never induced translocations, respectively; in contrast, hypotonic buffers containing detergents or the Ca2+-ionophore, A23187, frequently induced contractions. All fixations caused net nucleus movement away from the tip, with the amount of displacement depending on the pre-fixation distance from the tip. Similarly, all fixations moved the most-apical of saltatory vesicles away from the tip, but the total number in the apex increased or decreased depending on the fixative used. The patterns of these results suggest that nucleus and vesicle distribution controls may be related (with respect to most-apical organelles) but also at least partially independent (with respect to organelle populations in hyphal tips). Hyphal diameter was reduced by some, but not all fixations; this variability did not correlate with displacements of either organelle, nor with fixative osmotic pressure. Evidently fixative-induced changes are more complex and systematic in highly polarized tip-growing cells than previously reported in other, less polarized, cell types. These results also suggest that hyphae contain multiple and complex organelle distribution and hyphal diameter control systems which can be readily altered, often subtly, by fixation protocols commonly and uncritically employed in immunocytochemical and ultrastructural analyses, and that fixation can cause serious cellular reorganization.