Biography: Peter Gouras, M.D., Ph.D. h.c., is a Professor of Opthamology at Columbia University. He is a pioneer in the transplantation of retinal pigment epithelial cells and successfully treated a rat model of retinitis pigmentosa by transplanting these cells. Translating his research from the bench to the bedside, he also performed the first human retinal cell transplant in 1994. His current research employs electron microscopy to study the effects of aging, genetics, and calorie restriction on a primate model of macular degeneration. He also serves on the scientific advisory board of the Cryonics Institute.

Andy McKenzie: Through your research, you’ve worked on the chemical fixation of a number of different tissues. As I understand it, this has chiefly been on the the retina, where you have done groundbreaking work, but you’ve also fully preserved rhesus monkey brains for subsequent study. In you experience, what have been one or two of the major challenges that are involved with scaling from smaller tissues to larger ones?

Peter Gouras: For the brain it is essential to perfuse the organ as rapidly as possible after death, minutes rather than hours post mortem. I have always perfused monkey heads with their brains by first perfusing with cool saline, about 500ml, through the ascending aorta, clamping off the descending aorta and checking the venous outflow in a cut jugular vein for the loss of erythrocyte redness. Then I follow this with a buffered solution of glutaraldehyde (4%), about 500 ml. This usually produces some muscular contractions due to aldehyde irritation of some neural circuits. These slight motor movements provide me with a clue that the perfusate has gone through the brain. Good perfusion can always be checked by examining the tissue for any erythrocytes in vessels during histological examination usually best by electron microscopy. For mouse brain you must enter the circulation through the heart and much less solution is needed. For the retina it is not necessary to perfuse. One must only inject about 5-10 ml of the fixative into the vitreous cavity. The fixative rapidly diffuse into the retina and choroid and provides as good fixation as with perfusion.

Andy McKenzie: What do you think are the key one or two obstacles to be overcome if brain preservation were to be performed as an emergency elective procedure in a more formal medical setting?

Peter Gouras: I have tried to suggest that there has never been any perfusion of a human brain as far as I can tell. I have suggested here at Columbia that the donor organ team perfuse the brain of a donor who gives up other organs for transplantation. This could be added to his death request as an organ donor. We have a large facility in the Pathology Department that obtains human brains but only after long (10 hours) post mortem times. The brain is then dissected and put in a solution of 10% formaldehyde. The fixative reaches the brain interior by slow diffusion. I have examined the visual cortex of one of their human brains by electron microscopy and the status of the tissue is poor. One can perform some immunohistochemistry examined by light microscopy but one cannot do any circuit tracing, which is unfortunate. It would be a major advantage to have the donor teams cooperate by performing brain perfusion after the other transplantable organs are removed. One would have to eliminate any possible toxicity from aldehyde in the atmosphere but then you can really approach the fascinating project of tracing with great precision all the connections in a human brain.

Andy McKenzie: How do you think that you could measure, mitigate, or get around (eg, via a different perfusion agent) the problem of aldehyde toxicity?

Peter Gouras: I don’t think glutaraldehyde perfusion is a problem with proper ventilation. An option could be to perfuse the brain with a cold solution to reduce the brain temperature for transport and then do the fixation under a chemical hood.

Andy McKenzie: What do you think are the most pressing research problems in brain preservation?

Peter Gouras: 1. The unpredictable delay between clinical death and any attempt to preserve the brain.

2. The failure to develop a successful way so far to deep freeze the brain and rewarm it without producing damage.

3. Rejuvenating a senescent brain.

Andy McKenzie: Thank you so much for the fascinating answers, Dr. Gouras!