The effects of chemical fixation on the permeability of frog mesenteric capillaries

1. We have investigated the effects on microvascular permeability of two chemical fixatives, glutaraldehyde/formaldehyde mixture and osmium tetroxide, both of which are used widely in electron microscopy. 2. The permeability of single perfused frog mesenteric capillaries was assessed using the technique of Michel (1980) to estimate the hydraulic conductance (Lp) of the vessel walls and the effective osmotic pressure (sigma delta pi) which could be exerted across them by the neutral macromolecule, Ficoll 70. 3. In each experiment, Lp and sigma delta pi were estimated in the same capillary before and after chemical fixation with either an aldehyde mixture (n = 6) or 1% osmium tetroxide (n = 6). In the absence of serum proteins in the perfusate, the values of Lp recorded prior to fixation were high. The effect of both fixatives was to reduce Lp. With aldehydes the mean value (+/- S.E.M.) of Lp fell from 22.1 (+/- 6.3) x 10(-3) to 5.7 (+/- 2.0) x 10(-3) microns s-1 cmH2O-1. With osmium tetroxide the mean Lp fell from 29.1 (+/- 8.5) x 10(-3) to 8.6 (+/- 3.6) x 10(-3) microns s-1 cmH2O-1. 4. The perfusates contained the neutral macromolecule, Ficoll 70, at a concentration of 60 mg ml-1 which exerted an osmotic pressure of 35 cmH2O in a membrane osmometer. In the absence of plasma proteins in the perfusate, sigma delta pi was only a fraction of this perfusate oncotic pressure prior to fixation. After aldehyde treatment there was a small but insignificant decrease in sigma delta pi. Osmium tetroxide, however, increased sigma delta pi from a mean (+/- S.E.M.) of 6.3 (+/- 1.6) cmH2O before fixation to 16.4 (+/- 2.8) cmH2O after fixation, P less than 0.01 using a paired t test. 5. In three experiments, the reflection coefficient of the capillary wall to NaCl sigma NaCl, was measured using the technique of Curry, Mason & Michel (1976) before and after osmium fixation. Fixation raised sigma NaCl from a mean value (+/- S.E.M.) of 0.009 (+/- 0.001) to one of 0.017 (+/- 0.004). Although these changes may be interpreted as an increase in the contribution to the total Lp of the capillary wall of channels which are available only to water, the changes are too small for such a mechanism to account for the increase in reflection coefficient to Ficoll 70. 6. The reduction in Lp following chemical fixation could represent an increase in the hydraulic resistance of either the capillary wall itself, or in the tissues surrounding the vessel.(ABSTRACT TRUNCATED AT 400 WORDS)