X-Message-Number: 2778
From:
Date: Tue, 24 May 94 11:34:28 EDT
Subject: CRYONICS sheep heads
Following is a recent report of Ukrainian researchers on the sheep head work.
This covers perfusion and reperfusion of control heads. The freeze/thaw work
will probably not be finished until September. The IMMORTALIST will carry
photos and further information.
Subj: brief report of 1,2 stages
Date: 20 May 1994
Dear Mr. Ettinger,
We apologize for a delay in answering you. We hope that we do not
give you great inconveniences. Now we have a very strenuous work.
EFFECT OF PERFUSION ON TISSUE STRUCTURE OF SHEEP BRAIN
The weight of the control sheep head was 2.1 kg, and that of the
brain was 103 g. The head for perfusion was weighting 2.0 kg, and
the weight of the brain from it was 101 g. The cryoprotectant was
administered with the mean rate of 50 ml per 80 sec. The original
concentrations of glycerol were either 75% (v/v), or 80 % (w/w).
The concentration of glycerol in the last 20 ml of the fluid,
outflowing after perfusion, was 65% (w/w). The content of
glycerol in the brain fluid comprised 45% (w/w). reperfusion was
performed using 1 l of the Ringer's solution with the rate of 50
ml per 10 sec. One sheep head was used for analyzing glycerol
distribution in brain tissues. The weight of this head was 2.3
kg, and that of the glycerolized brain was 108 g. Perfusion
procedure was similar to that of the very first head. The
original concentration of glycerol was 82 (w/w). The
concentration of glycerol in the last 20 ml of perfusate
comprised 55% (w/w). The cortex of the large brain hemispheres
contained 57% of glycerol. A detailed description of the above
we'll submit You later on by air mail.
The particles of the grey and white substances were isolated from
the frontal lobes. At first we supposed to use hypothalamic
tissue, but substituted it for a hypophysial one afterwards,
since the cells of the hypothalamus are very diversified.
Staining of the nerve cells in the histologic slices was
conducted by a method of Nissel, and the hypophysial cells were
stained using methylene blue.
GREY SUBSTANCE
The cells of all six layers were nicely displayed at the
histological preparations of the grey substance. The
electron-microscopic shots demonstrate some alterations in the
native structure in 2 hours after the death of the brain. In
particular, the cysterns of the endoplasmic reticulum and Golgi
apparatus are expanded and vacuolized.
Perfusion of the brain did not result in the significant
pathology of the grey substance as evidenced by the histologic
slices, though a minor edema of the interneuronal space
(neuropile) was reported. We observed an increased vacuolization
of the neuronal bodies and fragmentation of the mitochondrial
crysts; however, they were difficult to be distinguished from the
post-mortem alterations. On the whole, the fine structures of the
neurones and their processes were well maintained following
reperfusion, and the capillary network was not damaged.
WHITE SUBSTANCE
We failed to identify any significant alterations in the white
substance on the histologic slices (at the depth of 1 cm from the
cortex) following reperfusion, though some blood vessels with the
exfoliating wall of the endothelium were found. The fine
structures of dendrites and axones, which comprise the bulk of
the white substance, were not significantly altered. A certain
layering of the myelin coat and shrinking of oligodendrites and
their nuclei were observed. The perinuclear space was slightly
expanded.
PITUITARY BODY
We studied the frontal lobe of the pituitary body. Histological
preparations displayed groups of the secretory cells among a
large amount of the big blood capillaries. After perfusion the
capillaries were expanding, and we observed the edema of the
tissue.
The fine structures of the secretory cells were altered
insignificantly, i.e. certain swelling and injuries of the
mitochondrial crysts were reported.
GENERAL CONCLUSION
Perfusion and reperfusion did not result in the significant
injuries in the structure and fine structure of the grey and
white brain substances. Some impairments were identified in the
circulatory system, and a moderate edema of the neuropile was
detected. The alterations in the vessels and the edema were more
pronounced in the pituitary body.
With best regards,
Sincerely,
Yu.Pichugin and G.Zhegunov
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