From: Eugene Leitl (Eugene.Leitl@lrz.uni-muenchen.de)
Date: Mon Jan 06 1997 - 07:13:21 MST
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Message #7423
Date: Sun, 5 Jan 1997 02:38:59 -0800 (PST)
From: Doug Skrecky <oberon@vcn.bc.ca>
Subject: Can the Visser Technique be Improved?
In cryonet message 7415 Saul Kent revealed
that the cryoprotectant used in the Visser technique is
dimethylformamide. Apparently relatively low and nontoxic concentrations
of this are used in conjunction with fast freezing to cryopreserve rat
hearts without vitrification. A possible improvement in this technique
might be achieved by choosing a different cryoprotectant or perhaps
combination of cryoprotectants.
When one examines the effect of cryoprotectants on subzero
nonfreezing storage of rat hearts one finds an interesting correlation.
The order of merit is methanol > ethanol > ethylene glycol > propylene
glycol > glycerol. (1) It is interesting that this is also the same order
ranking for the membrane permeability of these various substances. (2)
Yet more effective cryoprotectants might be found in those with either a
still greater membrane permeability or a lower toxicity. The later factor
may be the reason polyethylene glycol is better at preserving function in
frozen hearts than glycerol and why glucose is in turn better than
polyethylene glycol at preserving unfrozen hearts at high subzero
temperatures. (3) (4)
The reason for the selection of dimethylformamide is obvious from the
following table of relative membrane permeabilities: (all solutes with
permeabilities greater than methanol are listed from reference 2)
NAME MEMBRANE PERMEABILITY
glycerol 1
methanol 19.6
dimethylformamide 20.5
dioxane 20.6
ethylene glycol
monomethyl ether 20.9
ethylene glycol
monoethyl ether 22.1
dimethylacetamide 25.4
monobutyrin 34.1
diethylacetamide 36.2
urea 41.1
triethylene glycol
diacetate 44.1
pyridine 62.8
Possible improvements in the cryoprotectant solution might accrue by
considering solutes with permeabilities higher than dimethylformamide,
possibily in combination with glucose.
(1) Cryobiology 30:366-375 1993
(2) The Journal of General Physiology 62: 714-736 1973
(3) Cryobiology 29: 87-94 1992
(4) Transplantation Proceedings 26(4) 2428-2430 1994
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