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Thawing of rabbit kidneys from −79 °C with 2450 MHz radiation
ABSTRACTS,
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betwoe 80 and 90 mm Hg. Following contra1 perfusion, kidneys were perfused with 1.41 M [3H]DMS0 (0.88 &i/ml). The total voIume of venous and of ureteral effluents were collected at 2-min intervals for 30 min. The kidneys were then perfused with DMSO-free perfusate for 30 min and similar 2-min samples coIlected. In other experiments kidneys were perfused far either 5, 10, 15, or 30 min with [‘HIDMSO. Samples ( 100 mg) were immediately taken from cortex and medulla. Tissue samples were also taken from kidneys following the 30-min washout perfusion. Results indicate that tissue accumulation of DMSO in the medulla slightly preceded that in the cortex. The equilibration of DMSO is 60% complete within 15 min and 100% complete within 30 min; washout can be completed within 30 min. This work was supported by Grant No. AM 17816-01 from NIB. 70. Thawing of Rabbit Kidneys from -79°C with 2450 MHz Radiation. C. P. BURNS,’ E. C. I~URDETT, A. M. KAROW. (Engineering Experiment Station, Georgia Institute of Technology, Atlanta, GA 30332 and hfedical College of Georgia, Augusta, Georgia 30902). This study was done to develop instrumentation that will thaw rabbit kidneys at several different uniform rates. Kidneys (9-12 g) were perfused for 30 min at 37°C on a nonrecirculating circuit at a constant pressure of 6&80 mm Hg. The oxygenated perfusate, rich in K+ and Mg2* (1. SUT~. one of several Res. 14, 7 ( 1973) ), contained dimethyl sulfoxide (DMSO) concentrations: 0, 0.7, 1.4 or 2.1 M. Kidneys were then placed in a -80°C freezer; 30°C freezer; 30”C/min cooling rate, Two microwave radiators were studied. A dielectric-loaded (fused silica) waveguide probe permitted thawing rates approaching 2O”C/min at 1000 W; but ‘hot spots” developed on the kidney surface because a uniform-interface between the organ and radiator could not be achieved. A vertically mounted, sIightly flared horn radiator permitted placing the kidneys in direct contact with SiOaTiOa dielectric-loading material to achieve 9O”C/min thaw from -79°C with 400 W of radiated power. Other thaw rates can be obtained by changing the power level. Each kidney was perfused postthaw for 30 min at 37°C. The most uniform thawing was achieved with kidneys which had been perfused with 0.7 or 1.4 M DMSO; temperahne differentials between renal poles < 3°C. Postthaw perfusate flow rates were greatly diminished when 0 or 2.1 XI DMSO had been used. The viability of thawed
ANNUAL
577
M,EETING
kidneys will be tested by transplantation studies. This work was supported by NIH AM 17816-01.
in future Grant
No.
71. Posthypetionic Osmotic Shock and Myocardial Cryoinjury. G. M. FAHY AND A. M. &ROW, JR. (Departments of Pharmacology and Surgery, Medical College of Georgia, Augusta, Georgia 30902). Previous work in this Iaboratory (Cryobiology 12, 130 (1975)) suggests that cardiac cryoinjury takes pIace during thawing as a postthypertonic osmotic effect similar to that believed to kil1 frozen red cells ( CryobioZogy 9, 9, I.6 ( 1972); B&him. Biophys. Acta 10, 414 ( 1953)). Freezing injury by this mechanism couId potentially be prevented or reversed by the following methods: Replacement of extracellular permeable solute with extracellular impermeant solute; removal of all extracellular solute other than osmotically required cryoprotectant; postthaw exposure of cells to hypertonic impermeant solute to counter intracellular hypertonicity and allow cells to release excess solute while volume-protected. Rat hearts were perfused and frozen to -17°C in the presence of 2.1 &I dimethyl sulfoxide (DMSO) with previously described apparatus (Cryobiology 8, 280, 350 ( 1971) ), To prevent cryoinjury, NaCl was replaced in the usual balanced salt solution (BSS) with mannitol on an equiosmolar basis (noncorrected) prior to freezing, or hearts were perfused with simpIe buffered DMSO. To reverse cryoinjury, postthaw perfusate contained 300 mosbr mannitol for the first 20 min of postthaw perfusion, or hearts were perfused for 6 min with DMSO-BSS postthaw, in case DMSO itseIf should have been imbibed by heart cells during freezing. None of the described approaches resulted in better recovery than that recorded previously (Cryobiology 9, 38, ( 1972) ). Perfusion with buffered DMSO was lethal in itself after 4-5 min despite presumed lack of near-complete pcnetration. The results do not support an osmotic shock mechanism for myocardium. This work was partially supported by NIH Grant No. GM 08472-12. 72. A Study of the Eflects of Perfusion of Clycero~ in Rut Hearts. L. J. MENZ AND A. E. Surgery LaboraFISCHER.* (Experimental tories, Department of Surgery, Saint Louis University, 1402 South Grand BouIevard, Saint Louis, Missouri 63104). Isolated rat hearts were weighed and cannulated at the root of the aorta in preparation for a
IZTH
betwoe 80 and 90 mm Hg. Following contra1 perfusion, kidneys were perfused with 1.41 M [3H]DMS0 (0.88 &i/ml). The total voIume of venous and of ureteral effluents were collected at 2-min intervals for 30 min. The kidneys were then perfused with DMSO-free perfusate for 30 min and similar 2-min samples coIlected. In other experiments kidneys were perfused far either 5, 10, 15, or 30 min with [‘HIDMSO. Samples ( 100 mg) were immediately taken from cortex and medulla. Tissue samples were also taken from kidneys following the 30-min washout perfusion. Results indicate that tissue accumulation of DMSO in the medulla slightly preceded that in the cortex. The equilibration of DMSO is 60% complete within 15 min and 100% complete within 30 min; washout can be completed within 30 min. This work was supported by Grant No. AM 17816-01 from NIB. 70. Thawing of Rabbit Kidneys from -79°C with 2450 MHz Radiation. C. P. BURNS,’ E. C. I~URDETT, A. M. KAROW. (Engineering Experiment Station, Georgia Institute of Technology, Atlanta, GA 30332 and hfedical College of Georgia, Augusta, Georgia 30902). This study was done to develop instrumentation that will thaw rabbit kidneys at several different uniform rates. Kidneys (9-12 g) were perfused for 30 min at 37°C on a nonrecirculating circuit at a constant pressure of 6&80 mm Hg. The oxygenated perfusate, rich in K+ and Mg2* (1. SUT~. one of several Res. 14, 7 ( 1973) ), contained dimethyl sulfoxide (DMSO) concentrations: 0, 0.7, 1.4 or 2.1 M. Kidneys were then placed in a -80°C freezer; 30°C freezer; 30”C/min cooling rate, Two microwave radiators were studied. A dielectric-loaded (fused silica) waveguide probe permitted thawing rates approaching 2O”C/min at 1000 W; but ‘hot spots” developed on the kidney surface because a uniform-interface between the organ and radiator could not be achieved. A vertically mounted, sIightly flared horn radiator permitted placing the kidneys in direct contact with SiOaTiOa dielectric-loading material to achieve 9O”C/min thaw from -79°C with 400 W of radiated power. Other thaw rates can be obtained by changing the power level. Each kidney was perfused postthaw for 30 min at 37°C. The most uniform thawing was achieved with kidneys which had been perfused with 0.7 or 1.4 M DMSO; temperahne differentials between renal poles < 3°C. Postthaw perfusate flow rates were greatly diminished when 0 or 2.1 XI DMSO had been used. The viability of thawed
ANNUAL
577
M,EETING
kidneys will be tested by transplantation studies. This work was supported by NIH AM 17816-01.
in future Grant
No.
71. Posthypetionic Osmotic Shock and Myocardial Cryoinjury. G. M. FAHY AND A. M. &ROW, JR. (Departments of Pharmacology and Surgery, Medical College of Georgia, Augusta, Georgia 30902). Previous work in this Iaboratory (Cryobiology 12, 130 (1975)) suggests that cardiac cryoinjury takes pIace during thawing as a postthypertonic osmotic effect similar to that believed to kil1 frozen red cells ( CryobioZogy 9, 9, I.6 ( 1972); B&him. Biophys. Acta 10, 414 ( 1953)). Freezing injury by this mechanism couId potentially be prevented or reversed by the following methods: Replacement of extracellular permeable solute with extracellular impermeant solute; removal of all extracellular solute other than osmotically required cryoprotectant; postthaw exposure of cells to hypertonic impermeant solute to counter intracellular hypertonicity and allow cells to release excess solute while volume-protected. Rat hearts were perfused and frozen to -17°C in the presence of 2.1 &I dimethyl sulfoxide (DMSO) with previously described apparatus (Cryobiology 8, 280, 350 ( 1971) ), To prevent cryoinjury, NaCl was replaced in the usual balanced salt solution (BSS) with mannitol on an equiosmolar basis (noncorrected) prior to freezing, or hearts were perfused with simpIe buffered DMSO. To reverse cryoinjury, postthaw perfusate contained 300 mosbr mannitol for the first 20 min of postthaw perfusion, or hearts were perfused for 6 min with DMSO-BSS postthaw, in case DMSO itseIf should have been imbibed by heart cells during freezing. None of the described approaches resulted in better recovery than that recorded previously (Cryobiology 9, 38, ( 1972) ). Perfusion with buffered DMSO was lethal in itself after 4-5 min despite presumed lack of near-complete pcnetration. The results do not support an osmotic shock mechanism for myocardium. This work was partially supported by NIH Grant No. GM 08472-12. 72. A Study of the Eflects of Perfusion of Clycero~ in Rut Hearts. L. J. MENZ AND A. E. Surgery LaboraFISCHER.* (Experimental tories, Department of Surgery, Saint Louis University, 1402 South Grand BouIevard, Saint Louis, Missouri 63104). Isolated rat hearts were weighed and cannulated at the root of the aorta in preparation for a