A method for differentiating nonunique estimates of membrane transport properties: Mature mouse oocytes exposed to glycerol

Measurement of the osmotic response of a cell in the presence of cryoprotec tant facilitates the determination of permeability coefficients which, in t urn, can be used to design cryopreservation protocols which minimize osmoti c stress. One problem encountered in determining permeability coefficients, using the Kedem-Katchalsky (K-K) model of membrane permeability, is that s everal combinations of the three passive coupled transport coefficients, na mely, hydraulic permeability (L-p, mu m min(-1) atm(-1)), solute permeabili ty (P (gly), mu m s(-1)), and the reflection coefficient (sigma), can give a similar fit to the measured data. A method for determining the "correct" set of coefficients is suggested. The osmotic response of 10 metaphase II m ouse oocytes was measured on perfusion with 1.5 mol L-1 glycerol at 24 degr ees C. For 8 of 10 oocytes perfused, two combinations of L-p, P (gly), and sigma gave a predicted response which closely matched the measured osmotic response, depending upon the initial estimates supplied to the software for these parameters. For the remaining two oocytes, similar values for the pe rmeability coefficients were generated regardless of the initial estimates. To determine the correct set of parameters, the K-K equations were used to predict experimental conditions for which volumetric histories would be di stinctly different for the two sets of "best-fit parameters," and then addi tional experimental data were compared to these predictions. Thus a further three oocytes were perfused with 0.2 or 0.5 mol L-1 glycerol in the absenc e of nonpermeating solute. In the presence of both 0.2 and 0.5 mol L-1 glyc erol, L-p = 2.11 +/- 0.69, P (gly) = 0.0016 +/- 0.0015, and sigma = 0.44 +/ - 0.11 yielded a very poor fit to the measured response while L-p = 0.98 +/ - 0.70, P (gly) = 0.0031 +/- 0.0021, and sigma = 0.91 +/- 0.15 yielded a cl ose fit to the measured response. Thus the latter combination of coefficien ts was taken to be correct. (C) 1999 Academic Press.