X-Message-Number: 15693
Date: Mon, 19 Feb 2001 13:02:03 -0800
From: Hugh Hixon <>
Subject: Terminal cryoprotectant concentration/Re: Message #15685

In Message #15685, David Pascal writes in part:

"I've answered this already, but it's still worth examining, in that it's
an excellent example of why CI people have not bothered to answer it.  It
is first of all, factually incorrect.  'On CryoNet it has been
established that CI uses a more concentrated solution of glycerol than
has been used by any other organization.'  By whom?  Jeff's 'mystery
correspondents'?  If (again) a person can expend the effort to type
www.alcor.org (a mighty effort, I admit), one will notice that Alcor's
web site states in a description of their procedures:  "After the
surgical access is completed, a cold (about 5oC, just above freezing)
perfusate is recirculated through the patient while the cryoprotective
agent (glycerol) is gradually increased from a 4% glycerol solution to
75%..."  This used to be standard for all patients; I would imagine it's
now standard for whole-body patients, though I've heard Alcor has yet
another CPA (secret, of course, but that's OK - all secrets are fine,
provided they're not CI's)."

------------------------------
Alcor apologises for a poorly written description.  To be correct and
somewhat more elaborate;

Alcor's perfusion circuit incorporates a mixing reservoir, which provides
perfusate to the arterial pump.  75% glycerol MHP-2 is added to the mixing
reservoir as venous return from the patient is removed from the circulation
in equal volume. The result is a cryoprotectant ramp from 4%v/v glycerol to
a cutoff point of about 55%v/v glycerol (~7.5M).  There are pauses and
changes in the ramp rate to attempt to balance toxicity and time against
cryoprotection.  The concentrations of the components of MHP-2 (Alcor's
organ preservation solution) is the same in both the 4% and the 75% glycerol
perfusates to maintain electrolyte and substrate balances, and to prevent
edema.  The object of the ramp is to prevent the severe osmotic shrinkage of
cells that occurs when tissues are suddenly exposed to high concentrations
of osmotically active active agents such as glycerol (incidentally, this is
how disinfectants such as alcohol and brine work, by sudden severe
dehydration of cells).  At the ramp rates that Alcor uses, the glycerol and
water concentrations inside and outside the cells remain fairly close as the
glycerol/water ratio is increased. That is, glycerol replaces water in the
cells at near-equilibrium conditions.  To simply hammer the tissues with
7.5M glycerol until the arterial and venous concentrations matched has
always been considered a very bad idea.

Because the brain is uniquely protected by the blood-brain barrier (BBB),
cryoprotection of the brain is somewhat more complicated; the brain is
progressively dehydated through the BBB until in a glycerol concentration
range of about 1.8 - 2.2 M, cell shrinkage of the capillary lining
physically tears apart the tight junctions between the cells and allows
entry of the glycerol, after which cryoprotection of the brain proceeds as
described above.

This picture is complicated by the breakdown of the osmotic barrier of the
capillary bed for verious reasons, particularly including long intervals
between death and perfusion.  Once the differential permeability of the
capillary bed is lost, the result is severe tissue edema, and the brain
swelling and pump pressure causes the brain to be extruded through the
various openings in the brain case.

Hugh Hixon

Rate This Message: http://www.cryonet.org/cgi-bin/rate.cgi?msg=15693