The pressure response of (plant) cells to osmotic challenges depends on the reflection coefficient, sigma, of osmotically active solutes; it is less than predicted by the van't Hoff equation if sigma < 1. In Valonia utricularis, sigma is significantly reduced by internal (and, to a lesser extent, by external) unstirred layers, protecting the cytoplasm against vacuolar osmotic fluctuations. As shown by scanning and transmission electron microscopy, diffusion-restricted spaces are formed by innumerable small vacuoles that are interconnected with each other and with the central vacuole. They are embedded in networks of cytoplasmic strands connecting and encircling the organelles. Unstirred layers are also created in the central vacuole by an extensive network of acid mucopolysaccharide filaments (visualized by alcian blue staining). Mucopolysaccharides apparently also affect steady-state turgor by reducing the water activity. When the effective vacuolar osmotic pressure was adjusted to that of the bath by perfusion with an artificial vacuolar sap (AVS), an "offset turgor pressure" of 17 +/- 5 kPa was recorded. Consistent with the ultrastructural data, sigma values less than unity were calculated from the pressure response upon vacuolar addition of KCl or sucrose by perfusion (sigma(iKCl) = 0.63 +/- 0.13; sigma(isuc) = 0.58 +/- 0.17). Dilution of AVS yielded slightly higher sigma(iKCl) values (0.73 +/- 0.35). External addition to the artificial sea water (ASW) indicated that sigma(e) > sigma(i) for these osmotica. However, even in this case, sigma(esuc) (0.86 +/- 0.09) and sigma(ePEG) (0.58 +/- 0.08) were significantly less than sigma(eNaCl) (0.94 +/- 0.05) and sigma(eKCl) (0.91 +/- 0.13), presumably due to unstirred layers within the 4 micro m thick cell wall. Consistent with the low sigma values, a partial replacement of NaCl by osmotically equivalent amounts of sucrose (ASW(suc)), PEG and dextran, respectively, as well as replacement of Cl(-) by the large anion MES(-) induced an 'anomalous' hyposmotic turgor pressure response followed by the usual backregulation of pressure. After a 2-day preincubation in ASW(suc), significantly lower sigma(e) values were obtained both hyperosmotically (sigma(eNaCl) = 0.78 +/- 0.14; sigma(esuc) = 0.72 +/- 0.15) and hyposmotically (sigma(eNaCl) = 0.70 +/- 0.17; sigma(esuc) = 0.63 +/- 0.09), probably due to long-term effects on membrane structure to be elucidated yet. The freshwater alga Chara corallina lacked these apparently closely related structural and biophysical features of Valonia.
