Maintenance of the Cardiovascular Function in a Deeply Cooled Homeothermic Organism by Physiological Methods without External Rewarming.

Gradual cooling of homeothermic organisms is followed by slowing and arrest of breathing and heart contractions. During deep cooling, even relatively slight artificial ventilation decreases the lower temperature limit of life (by 4.5-5°C) and provides minimum oxygen supply to the heart and whole body. This allows us to restore cardiovascular function and prevent animal death after lethal cooling without eternal warming.

[Restoration of respiratory function of an organism after cold oppressions without warming].

In experiments on rats studied influence of intravenous introduction NazdTA on respiratory function of rats after the full termination of breath at deep hypothermia. It is shown that such method without warming application it is possible to lower on 1.5-2.0 degrees C a temperature threshold of approach full a cold paralysis of respiratory function of an organism. It confirms the hypothesis put forward earlier about an important role of infringement of balance of ions of calcium in cytoplasm of cells in development of a pathology of an organism at deep hypothermia. The received results can be used at resuscitation of victims accidental hypothermia.

[Resuscitation of vital activity in intensively cooled animals by physiological methods without rewarming].

White rat males (Wistar) were cooled in the water (9-10 °C) to the stop breathing (at rectal temperature 14.7 ± 0.5°, brain 16.0 ± 0.3 °C). After the removal of water animals were injected 0.5% solution disodium salt ethylenediaminetetraacetic acid (Na2EDTA) intravenously, which reduced the concentration of Ca2+ in the blood. Breathing rats were resumed after 4-8 min after administration Na2EDTA. Then one group of rats was cooled to the temperature of the body 12.2 ± 0.7 (brain 14.9 ± 0.3 °C). At such a low temperatures the breath of rats was long-term (2-3 hours) supported at the level of 12 ± 3 cycle/min. The second group of rats was not additionally cooled after the resumption of breathing. The same dose of Na2EDTA caused the rise of the respiratory rate to a higher level--29 ± 4 cycles/min at a temperature in the rectum and the brain 15-16°. Apparently, the process of accumulation of Ca2+ in the cytoplasm of the respiratory center's cells was slowed after the Na2EDTA injection to hypothermal rats in the bloodstream. It is assumed that the Na2EDTA injection under the hypothermia conditions was activated the cell's protection mechanisms from cold damage. The obtained results and the literature data allowed to conclude that nonhibernators are able to maintain basic life functions under significant reduction in the body temperature.

[Cold pathology and experimental therapy of deep hypothermia].

A complete cold paralysis of respiration and thermoregulation occurs in rats at the temperature in the brain 16.6 +/- 0.3 and in the rectum 15.2 +/- 0.3 degrees C. Under the conditions of room temperature 18-19 degrees C, the respiration never restores, and the animals die. This is believed [6] to be the result of calcium ion Ca2+ accumulation in the cells of respiration and thermoregulation centers. After the arrest of respiration the animals were injected with the solution of disodium salt of ethylenediaminetetraacetic acid (Na2EDTA), which binds calcium ions in the blood and facilitates their removal from the cell (explanations in the text). In 7-9 min after the injection the calcium content in the blood decreased and the respiration began to restore at the temperature of the cold paralysis. Thermoregulation was also restored. All the test animals survived. All the control animals, which were not injected with Na2EDTA, died.

Respiration restitution in rats following its termination in immersion hypothermia.

Rats were cooled in water until attaining profound hypothermia and respiratory arrest. After removal from water, 0.5% solution of Na2EDTA was administered intravenously. This led to a drop of blood [Ca2+] by 20-30% from the baseline and promoted recovery of respiration following its arrest lasting 10.3±1.4 min. By the 30th minute of Na2EDTA administration, respiration rate increased to 32.3±5.2 cycles per minute and respiration amplitude reached 68±4% of the baseline level. This effect was observed without special warming of the rats. It was concluded that the period during which the organism maintains viability in respiration arrest and disturbances in respiratory center are still reversible is prolonged under conditions of profound hypothermia.

Mechanism of suppression of physiological functions in hypothermia and means for their stimulation without body warming.

Cold-suppressed thermoregulatory reactions and respiration in rats in deep hypothermia (rectal body temperature (25-22 degrees C) were shown to be stimulated by injecting disodium ethylenediaminetetraacetic acid (EDTA) solution into the blood stream of cold rats at a dose of 16.5 mg/100 g (0.0045 mmol/100 g). EDTA binds Ca2+ ions in the blood, forming complexes. Increases in cold shivering and pulmonary respiration (by 5 min after the start of administration) coincided with a reduction in the blood Ca2+ concentration by 42-45% of normal. By 15 min after the start of the EDTA injection, the blood Ca2+ concentration returned to the normal level present in cold rats before EDTA treatment. This was accompanied by suppression of cold shivering and pulmonary respiration. Repeated injection of EDTA into the blood stream produced a new drop in blood Ca2+ and repeated stimulation of cold shivering and pulmonary respiration.

[Influence of EDTA injection in the blood of the rat on the process of self-warming after deep hypothermia].

With the help of thermometry and general calorimetry, changes in the heat exchange were determined in rats upon their getting out of artificial deep hypothermia (the temperature in the rectum was 19-20 degrees C) at the temperature in the calorimeter chamber of 20 degrees C. In initial study of self-warming (the temperature in the rectum was 21-22 degrees C), the rats were injected with 1 ml 0.5% ethylenediaminetetraacetic acid disodium salt (EDTA) with purpose of speeding up the recovery to normal state. The experiments showed that EDTA increased the total heat production and kept in on high level during 3 hous. During that time, the rectal temperature of the rats was 37.6 +/- 0.6 degrees C in the end of the experiments, but in the control that value was only 35.1 +/- 0.5 degrees C.

[Development of hypothermia in rats at increase of [Ca2+] in the blood].

In rats, data on influence of i. v. administration of calcium chloride on the level of [Ca2+] in the blood and on process of oppression ofthermoregulatory and respiratory functions in rats in hypothermia. 0.18 or 0.135 mmol Ca2+ on the 3rd minute from beginning of the administration increased [Ca2+] in the blood from 1.01 +/- 0.03 to 2.56 +/- 0.08 mM (or 2.27 +/- 0.06 mM). Then [Ca2+] was reduced gradually, in 20 minutes from administration, solution of CaCh [Ca2+] exceeded the initial level by 20-30 %. The increase of concentration of ionized calcium in the rat blood strengthened the cold oppression of breathing and cold shivering as compared with the control (administration of physiological solution). Arrest of breathing in rats after administration of CaCl2 solution occurred at higher rectal temperatures (21 +/- 0.03 degrees C) as compared with control experiments (18 +/- 0.4 degrees C), p < 0.05. It is suggested that increase of [Ca2+] in the blood strengthens effects of cold in the form of oppression of thermoregulatory and respiratory functions.

[Mechanisms of suppression of physiological function in hypothermia and a way of their restoration without body warming].

It was shown that in hypothermic rats (rectal temperature 25-22 degrees C) it was possible to stimulate responses that had been suppressed by cold (i. e. thermoregulation and breathing) with the aid of injecting a solution of ethylenediaminetetraacetic acid disodium salt (EDTA) in quantity 16.5 mg/100 g of body weight (0.0045 mmol/100 g) into the blood stream of the cooled animals. EDTA connects calcium ions in blood and forms complexes. It was shown that enhancement of cold shivering intensity and that of breathing (in 5 min after beginning the injection of EDTA) coincided with a 42-45 % reduction of [Ca2+] in the blood]. After 15 min following the beginning of injection of EDTA [Ca2+] into the blood stream, a return to the initial level was observed in cooled animals. Simultaneously we observed suppression of the cold shivering and breathing. The repeated injection of EDTA again caused similar fall of [Ca2+] in the blood and the following enhancement of cold shivering and breathing.

[Stimulation of physiological functions in cooled rats without rewarming after introducing ethylenediaminetetraacetate into lateral ventricles of the brain].

On cooling the animals to the rectum temperature from approximately 37 to 24-28 degrees C the decreases were shown to occur in the frequency and amplitude of respiration motions, in the intensity of muscle electrical activity (thermoregulation muscle tone and cold muscle shivering), in the frequency of heart contractions. In 3-8 min after introducing ethylenediaminetetraacetate into a lateral ventricle of the rat brain the frequency and the amplitude of respiration motions increased statistically reliable and so did the intensity of thermoregulation muscle tone and cold muscle shivering (judging from the total muscle electrical activity). The doses of EDTA, which caused this effect, were by a factor of 30 - 100 less than the doses, which caused a similar stimulation of the functions in cooled animals after introduction into the blood.

[Stimulation of thermoregulatory and respiratory functions with the help of EDTA and EGTA during deep hypothermia in rats]. / Stimuliatsiia termoreguliatornoi i dykhatel'noi funktsii s pomoshch'iu EDTA i EGTA glubokoi gipotermii u krys.

Influence of EDTA (C10H14N2Na2O8.2H2O) and EGTA (C14H24N2O10) on physiological functions homoiothermic organisms at deep hypothermia, was studied. White rats during cooling were in special sections without rigid fixing of head and limbs. In reply to intravenous introduction of EDTA and EGTA solutions, similar answers of the organisms were observed: raised breathing frequency and amplitude, intensity of electrical activity of muscles; these signs of activation of physiological functions lasted 8-10 minutes. Besides, of the 20th-30th minute after introduction of the second dose of preparations (at rectal temperature 17.1 +/- 0.5 degrees C), the secondary activation respiratory and thermoregulatory functions were registered. The termination of the cold shivering in experiments with introduction of EDTA and EGTA solutions occurred at lower temperatures in rectum and in a brain (16.7-17.3 degrees and 17.8-18.2 degrees C, resp.) than in control experiments (18.7 +/- 0.6 degrees C and 20.2 +/- 1.5 degrees C). The authors suppose that the activation of the thermoregulatory and respiratory functions is caused by a decrease in concentration of ions Ca2+ in the blood plasma.

[Development of deep hypothermia in rats with limited motor activity]. / Osobennosti razvitiia glubokoi gipotermii u krys pri razlichnoi stepeni ogranicheniia dvigatel'noi aktivnosti.

Changes of the main organism functions (breathing frequency, heart rate and shivering) were investigated under hypothermia in two groups of rats. Animals of the first group were fixed rigidly on the special platform with fixing of head and limbs, and those of the second one--the rats, were placed in a punched cylindrical chamber, inside which they could move freely forward and back. In 2.5-3.0 hours after anaesthesia the rats were placed in a refrigerator (-5 degrees C) until they stop breathing. Cessation of breathing of the first group rats occurred in 1.7 +/- 0.3 hours from the beginning of cooling at body temperature 17.3 +/- 0.6 degrees C and the brain temperature 15.7 +/- 0.5 degrees C. In the second group, a prolonged activation of the frequency of breathing, heart rate and intensity of electrical activity of muscles during 2.5-3.0 hours, was observed. Only in 4.5-5.0 hours, the breathing stopped at rectal temperature 12.3 +/- 1.1 degrees C and the brain temperature 12.9 +/- 0.9 degrees C. In these animals, the time of survival in the cold environment increased considerably and the temperature thresholds of the termination of breathing were lowered. Thus, the activation in the thermo-regulative muscle tone and in shivering muscles provides the most effective resistance against cooling of rats, reducing a surface of heat, dissipation and keeping the temperature of internal areas of body.

[Effect of Na2H2EDTA on the viability of rats' organism during hypothermia]. / Vliianie Na2H2EDTA na zhiznesposobnost' organizma krys pri gipotermii.

Effects of C-binding Na2H2EDTA (disodium salt of ethylene diamine tetraacetic acid) on the thermal control function of muscles and respiration in anesthetized rats of the Wistar strain cooled up to deep hypothermia were studied. Experiments were performed with the use of two cooling models, i.e. in water (10 +/- 0.2 degrees C) and air (-5 +/- 1 degrees C). Intravenous injection of 1 cm3 of the 0.6% Na2H2EDTA solution at 0.25 ml/min. was made first at the brain T = 24.0 +/- 0.4 degrees C and repeated at the brain T = 20.1 +/- 0.3 degrees C. Five to six minutes following injection the electrical myoactivity and respiration rate and amplitude grew up. The control experiments with injection of physiological solution were with null results. Injection of Na2H2EDTA reduced the thermal viability lower range as concluded from the drop of no-breathing temperature threshold and increase in tolerance period till respiration standstill by 10-15 min. It is supposed that physiological functions paralyzed by continuous cooling can be recovered by reducing active calcium in blood with intravenous Na2H2EDTA. This opens up brand new vistas for application of calcium-binding substances to assist overcooled organism.

[Blocking of cold paralysis of physiological functions]. / Blokada mekhanizmov kholodovogo paralicha fiziologicheskikh funktsii.

For more than 100-year history of studying hypothermia a rewarming of a cooled organism was always considered as the only method of restoring the cold-paralysed physiological functions and the vital activity of an overcooled organism of homoiothermal animals and humans. The paper gives the results of applying the method developed at our Laboratory, of restoring the cold-paralysed important physiological functions after a continuous cooling of rats, without rewarming the animals.

[Ca(2+) level in the animal blood and their cold resistance]. / Soderzhanie Ca(+) v krovi zhivotnykh i ikh ustoichivost' k kholodu.

Administration of small doses of the EDTA decreased by 15-20% the Ca2+ contentn in the blood plasma of rabbits and rats. The decrease coincided with an abrupt stimulation of the thermoregulation system of cooled animals. Restoration of the Ca2+ content in circulating blood coincided in time with repeated suppression of the system's functions. The findings corroborate the theory of a key role of the Ca2+ in sensitivity of the homoiothermal organism to cold and substantiates the method of restoring physiological functions in deep hypothermia without rewarming the body.

[Resuming of thermoregulation in rats during deep hypothermia with EDTA without rewarming the body]. / Vozobnovlenie funktsii termoreguliatsii u krys pri glubokoi gipotermii s pomoshch'iu EDTA bez otogrevaniia tela.

Administration of the EDTA, a substance binding calcium ions, helped to resume thermoregulation functions after their "cold paralysis" at the rectal temperature 17.1 +/- 0.2 degrees C and the brain temperature 19.2 +/- 0.2 degrees C, without warming up the animal. Enhancement of the muscle electrical activity following the EDTA administration was due both to increase in the number of bursts of shivering and to enhancement of their intensity. The findings suggest that, in hypothermia, improvement of conditions of the calcium ion transport from the cell cytosole into interstitial space helps to restoration of the cold muscle shivering as an important function of the thermoregulation.