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.     

Development of deep hypothermia in rats with limited motor activity

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.     

About a correlation between calcium ion concentration (Ca2+) in the blood and the state of physiological functions in animals in deep cooling

The changes in physiological functions of the organism (respiration, functions of the heart and vessels, thermoregulation) were studied. The concentration of Ca2+ ions in the blood of white rats was determined by the ion-selective electrodes at various stages of hypothermia. The aim of the study was to reveal changes in the blood concentration of ionized calcium in animals during their gradual cooling. In deep hypothermia (16 degrees C), calcium ion concentration in the blood increased by 30% against the norm which coincides with arrest of the cold shivering and lung ventilation. An increased content of Ca2+ in the blood is supposed to result in an increase in the content of these ions in the intercellular liquid and in the nervous cells, which is one of the reasons for the cold paralysis of the respiration and thermoregulation centers.     

Adaptive thermogenesis in Brandt's vole (Lasiopodomys brandti) during cold and warm acclimation

Brandt's voles (Lasiopodomys brandti) exposed to cold (5±1 °C) or warm (23±1 °C) showed some physiological and biochemical variations which might be important in adaptation to their environments. Cold acclimation induced increases in resting metabolic rate (RMR) and the serum triiodothyronine (T₃) level, the state-4 respiration of liver and muscle mitochondria were activated after 7 days when animals exposed to cold, and the activity of cytochrome c oxidase (COX) of liver and muscle mitochondria tended to rise with cold exposure. RMR and T₃ level decreased during warm acclimation. The state-4 respiration of liver mitochondria declined after 3 days and muscle after 7 days when animals exposed to warm, and the activities of COX of liver and muscle mitochondria tended to decrease with warm acclimation. The cold activation of liver and muscle mitochondrial respiration (regulated by T₃) was one of the cytological mechanisms of elevating RMR. Both state-4 respiration and COX activity of brown adipose tissue (BAT) mitochondria increased significantly during cold acclimation and decreased markedly after acclimated to warm. The uncoupling protein 1 (UCP1) contents in BAT increased after exposure to cold and decreased after warm acclimation. Nonshivering thermogenesis (NST) plays an important role in the process of thermoregulation under cold acclimation for Brandt's voles. Changes in thermogenesis is a important way to cold adaptation for Brandt's voles in natural environments.     

Stimulation of thermoregulatory and respiratory functions with the help of EDTA and EGTA during deep hypothermia in rats

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.     

Synchronized slow-amplitude modulations in the electromyograms of shivering muscles

The electromyograms (EMG) of shivering human subjects exposed to 0 degrees C air in an environmental chamber were analyzed to detect slow-amplitude modulations (SAMs, less than 1 Hz) in the EMGs of widely separated muscles and to study the relationship of these SAMs to respiration rate and skin temperature. Distinct amplitude modulations were observed in the raw EMGs during shivering. The peaks in EMG activity occurred simultaneously in the majority of the monitored muscles in all subjects. Pearson correlations between the average rectified EMGs of 93% of the muscles were significant (P less than 0.05). Visual analysis of the EMG and respiration signals indicated that the peaks in muscular activity occurred 6-12 times/min, whereas respiration ranged from 10 to 23 cycles/min. For all subjects respiration was at a higher frequency than amplitude modulation in the EMG. Comparison of EMG records with expiratory flow rate traces in shivering subjects indicated no one-to-one correlation between the occurrence of respiration and EMG amplitude modulations. Respiratory flow rate and average rectified EMG showed significant correlation in only 33% of the cases. In addition, skin temperature changes could not be correlated with the SAMS.     

Acute and delayed biochemical and physiological effects of exhaustive weightlifting exercise

The goal of the work was a study of the effect of exhaustive weightlifting exercise on prolonged changes in the physiological and biochemical variables characterizing the functional state of skeletal muscles. An exercise accentuated at muscles of the hip surface gave rise to a significant increase of the blood lactate concentration, which indicated that aerobic metabolism was a predominant mechanism of energy supply for muscle contraction. A reduction of the m. rectus femoris EMG amplitude and frequency, a decrease in the tone of tension, and an increase in the tone of relaxation were found immediately after exercise. One day later, the amplitude and the frequency of the EMG signal increased. On day 3, the activity of creatine kinase (CK, a marker of muscle injury) considerably increased, while the amplitude and frequency of EMG decreased. By the ninth day of recovery, all measured variables with the exception of CK were normalized. A significant negative correlation was found between the blood serum’s lactate concentration and m. rectus femoris EMG activity at the same time points. Blood serum CK activity and m. rectus femoris EMG and tone parameters were significantly correlated on the third postexercise day. The data demonstrate that exhaustive exercise-induced muscle injury resulted in phasic alterations in the electrical activity and the tone of the muscle apparently related to a decrease in pH because of lactate accumulation in the sarcoplasm and the cascade of reactions leading to muscle tissue damage.     

Metabolic changes associated with sustained 48-Hr shivering thermogenesis in the newborn pig

Metabolic changes associated with sustained 48-hr shivering thermogenesis were studied in piglets maintained at 34 (thermoneutrality) or 25°C (cold) between 6 and 54 hr of life. Despite their high shivering activity and elevated heat production, cold-exposed piglets exhibited a slightly lower rectal temperature than thermoneutral animals (-1.1°C; P < 0.01) at the end of the treatment. The enhancement of heat production and shivering activity were associated with a decrease in muscle glycogen (− 47%; P < 0.05) and total lipid content (− 23%; P < 0.05), a reduction of blood lactate levels (P < 0.05) and an enhancement of muscle cytochrome oxidase activity (+20%;P < 0.05), which suggests that muscle oxidative potential was increased by cold exposure. Potential for capturing lipids (lipoprotein lipase activity) was also higher in the redrhomboideus muscle (+ 71%; P < 0.01) and lower in adipose tissue (−58%; P < 0.01) of the cold-exposed piglets. Measurements performed at the mitochondrial level show no changes inrhomboideus muscle, but respiratory capacities (state IV and FCCP-stimulated respiration) and intermyofibrillar mitochondria oxidative and phosphorylative (creatine kinase activity) capacities were enhanced inlongissimus dorsi muscle (P < 0.05). These changes may contribute to provide muscles with nonlimiting amount of readily oxidable substrates and ATP necessary for shivering thermogenesis. A rise in plasma norepinephrine levels was also observed during the second day of cold exposure (P < 0.05).     

Electromyography of the opercularis muscle of Rana catesbeiana: An amphibian tonic muscle

The opercularis muscle of Rana catesbeiana originates on the suprascapular cartilage of the shoulder girdle and inserts on the otic opercular element. It is part of the levator scapulae musculature and lies dorsomedial to the levator scapulae superior and inferior muscles. Bipolar electrode recordings from all three muscles show electrical activity linked to cyclical firing of the posterior intermandibularis muscle, an important ventilatory muscle. The opercularis muscle shows low amplitude, erratic signals when animals are sumerged. Upon emergence of the snout region, the opercularis muscle shows rhythmic low amplitude activity at twice the rate of buccal pumping. Lung ventilation is synchronized with this rhythm and at ventilation the opercularis muscle shows higher amplitude activity. Upon submergence, opercularis activity again shows low level activity with no rhythmic pattern. Opercularis muscle activity has a major low frequency component (about 30 Hz) that probably corresponds to activity of tonic muscle fibers. Higher frequency signals (about 200–250 Hz) comparable to those of the levator scapulae muscles are also present and probably represent activity of phasic muscle fibers. Activity of the opercularis muscle is correlated with conditions in which aerial respiration is possible, and this pattern of activity supports an opercularis role in aerial hearing and/or detection of substrate vibrations. As far as we know, this is the first report of electromyographic analysis of a vertebrate tonic muscle.     

Hypoxia-induced changes in shivering and body temperature

Experiments were carried out on conscious cats to evaluate the general characteristics and modes of action of hypoxia on thermoregulation during cold stress. Intact and carotid-denervated (CD) conscious cats were exposed to ambient hypoxia (low inspired O2 fraction) or CO hypoxia in prevailing laboratory (23-25 degrees C) or cold (5-8 degrees C) environments. In the cold, both groups promptly decreased shivering and body temperature when exposed to either type of hypoxia. Small increases in CO2 concentration reinstituted shivering in both groups. At the same inspired concentration of O2, CD animals decreased shivering and body temperature more than intact cats. While this difference resulted, in part, from a lower alveolar PO2 in CD cats, a difference between intact and CD cats was apparent when the two groups were compared at the same alveolar PO2. During more prolonged hypoxia (45 min), shivering returned but did not reach normoxic levels, and body temperature tended to stabilize at a hypothermic value. Exposure to various levels of hypoxia produced graded suppression of shivering, with the result that the change in body temperature varied directly with inspired O2 concentration. Hypoxia appears to act on the central nervous system to suppress shivering and sinus nerve afferents appear to counteract this direct effect of hypoxia. In intact cats, this counteraction appears to be sufficient to maintain body temperature under hypoxic conditions at room temperature but not in the cold.     

Shivering thermogenesis in leg and breast muscles of galliform chicks and nestlings of the domestic pigeon.

We studied the ontogeny of shivering thermogenesis in breast and leg muscles of precocial galliforms (domestic fowl, grey partridge, and Japanese quail) and the altricial domestic pigeon using electromyography (EMG) and indirect calorimetry. Galliforms were able to increase heat production by shivering in leg muscles at the youngest age studied (1-2 d). Pectorals contributed to heat production from days 7-10 onward, but in the partridge and especially in the fowl, shivering by the pectorals was weaker than in the quail. In the pigeon, shivering began in pectorals and legs at 2 and 4 d of age, respectively, and pectorals had clearly the predominant role in thermogenesis. Despite the early beginning of electrical signs of shivering, significant thermogenesis did not appear in the pigeon before the age of 6 d. All galliforms shivered in bursts, like pigeons aged 2-4 d. From the age of 6 d onward, continuous shivering became predominant in the pigeon. In pectorals of 2-6-d-old pigeons, shivering did not increase linearly during decreasing ambient temperature, as in other muscles and species, but started abruptly, at full intensity. Furthermore, in 2-4-d-old pigeons, cooling induced movement activity in legs. The median frequency of shivering EMGs varied (1) with maturation of the muscle, (2) with size of the adult bird, and (3) between altricials and precocials.     

Postnatal changes in regional blood flow during cold-induced shivering in sow-reared piglets.

To determine whether newborn pigs are able to display adequate cardiovascular adjustments favouring shivering thermogenesis in skeletal muscles soon after birth, regional blood flow and fractional distribution of cardiac output were determined in 1-day-old (n = 6) and 5-day-old (n = 6) conscious piglets at thermal neutrality and during cold exposure, using coloured microspheres. Five-day-old piglets stayed with the sow before the experiment. The cold challenge was designed to induce a similar increase (approximately +90%) in heat production at both ages. Skeletal muscle blood flow increased with both age (p < 0.05) and cold exposure (p < 0.001), with the effect of cold being more pronounced in 5-day-old piglets than in 1-day-old piglets (+60%, p < 0.05). The difference between individual muscles increased with age, with fractional blood flow being 41% higher in rhomboideus than in longissimus thoracis muscle during cold exposure in 5-day-old piglets (p < 0.05). Cardiac output was similar at both ages and increased by 23% in the cold (p < 0.001). At 1 day of age, there was no redistribution of cardiac output among the internal organs during the cold challenge, while at 5 days of age, the increase in muscle fractional blood flow was associated with a reduction (p < 0.05) in the fraction of cardiac output reaching the skin (-24%), the small intestine (-21%), and the liver (-20%). In conclusion, these results suggest that there is a rapid postnatal improvement of cardiovascular adjustments favouring blood perfusion and probably heat production during cold-induced shivering in the most oxidative muscles studied. This cardiovascular response may play a role in the postnatal enhancement of thermoregulation in piglets.     

Effects of carbohydrate availability on sustained shivering II. Relating muscle recruitment to fuel selection

The purpose of this study was to quantify how shivering activity would be affected by large changes in fuel metabolism (see Haman F, Peronnet F, Kenny GP, Doucet E, Massicotte D, Lavoie C, and Weber J-M, J Appl Physiol 96: 000-000, 2004). Adult men were exposed to 10 degrees C for 2 h after a low-carbohydrate diet and exercise (Lo) and after high-carbohydrate diet without exercise (Hi). Using simultaneous metabolic and electromyographic (EMG) measurements, we quantified the effects of changes in fuel selection on the shivering activity of eight large muscles representing >90% of total shivering muscle mass. Contrary to expectation, drastic changes in fuel metabolism [carbohydrates 28 vs. 65% of total heat production (Hprod), lipids 53 vs. 23% Hprod, and proteins 19 vs. 12% Hprod for Lo and Hi, respectively] are achieved without altering the EMG signature of shivering muscles. Results show that total shivering activity and the specific contribution of each muscle to total shivering activity are not affected by large changes in fuel selection. In addition, we found that changes in burst shivering rate ( approximately 4 bursts/min), relative contribution of burst activity to total shivering ( approximately 10% of total shivering activity), and burst shivering intensity ( approximately 12% of maximal voluntary contraction) are the same between Lo and Hi. Spectral analysis of EMG signals also reveals that mean frequencies of the power spectrum remained the same under all conditions (whole body average of 78 +/- 5 Hz for Lo and 83 +/- 7 Hz for Hi). During low-intensity shivering, humans are therefore able to sustain the same thermogenic rate by oxidizing widely different fuel mixtures within the same muscle fibers.     

Effects of hypoxia and cold acclimation on thermoregulation in the rat.

The effects of hypoxia (inspired O2 fraction = 0.12) on thermoregulation and on the different sources of thermogenesis were studied in rats before and after periods of 1-4 wk of cold acclimation. Measurements of metabolic rate (VO2) and body temperature (Tb) were made at 5-min intervals, and shivering activity was recorded continuously in groups of rats subjected to three protocols. In protocol 1, rats were exposed to normoxia to an ambient temperature (Ta) of 5 degrees C for 2 h. In protocol 2, at Ta of 5 degrees C, rats were exposed for 30 min to normoxia, then for 45 min to hypoxia, and finally for 30 min to normoxia. In protocol 3, in the non-cold-acclimated (NCA) rats, Ta was decreased from 30 to 5 degrees C in steps of 5 degrees C and of 30-min duration while in cold-acclimated (CA) rats at 5 degrees C for 4-wk, Ta was increased from 5 to 30 degrees C in steps of 5 degrees C and of 30-min duration. Recordings were made in normoxia and in hypoxia on different days in the same animals. The results showed that 1) in NCA rats, cold exposure in normoxia induced increases in VO2 and shivering that were proportional to the decrease in Ta; 2) in CA rats in normoxia, for a given Ta, VO2 and Tb were higher than in NCA rats, whereas shivering was generally lower; and 3) in both NCA and CA rats, hypoxia induced a transient decrease in shivering and a sustained decrease in nonshivering thermogenesis associated with a marked decrease in Tb that was about the same in NCA and CA rats. We speculate that hypoxia acts on Tb control to produce a general inhibition of thermogenesis. Nonshivering thermogenesis is markedly sensitive to hypoxia, especially demonstrable in CA rats; a recovery or even an increase in shivering can compensate for the decrease in nonshivering thermogenesis.     

Inhibition of shivering during restraint hypothermia

Restraint hypothermia has often been described, but its cause has never been clarified. We hypothesized that it might be due to a suppression of shivering thermogenesis. Thus, we restrained conscious rats in an ambient temperature of 2 degrees C while measuring rectal (Tre) and tail skin temperatures, metabolic rate (MR), and shivering activity. When rats were cold exposed but not restrained, Tre fell 1.4 +/- 0.2 degrees C (SE) during the 1st h. When these same rats were restrained, Tre fell at a rate of 6.5 +/- 0.2 degrees C/h. MR averaged 15.7 +/- 1.4 W/kg for the unrestrained rats, but it averaged only 9.0 +/- 1.1 W/kg for the restrained rats. The restrained rats showed no signs of shivering. The animals were then subjected to a restraint adaptation regimen and then reexposed to cold. Restraint now produced a fall in Tre of only 2.6 +/- 0.7 degrees C/h. The animals shivered and generated an MR of 15.8 +/- 0.9 W/kg. Naive rats became hypothermic because restraint suppressed shivering activity. However, adapted rats continued to shiver and remained normothermic. We suggest that a stressful or threatening situation, such as restraint for a naive rat, inhibits shivering and leads to hypothermia in a cold environment. This would not occur in adapted rats because restraint is no longer stressful.     

Effects of the menstrual cycle on muscle recruitment and oxidative fuel selection during cold exposure

Differences in core temperature and body heat content, generally observed between the luteal and follicular phase of the menstrual cycle, have been reported to modulate the thermogenic activity of cold-exposed women. However, it is unclear how this change in whole body shivering activity will influence fuel selection. The goal of this study was to quantify the effects of the menstrual cycle on muscle recruitment and oxidative fuel selection during low-intensity shivering. Electromyographic activity of eight large muscles was monitored while carbohydrate, lipid, and protein utilization was simultaneously quantified in the follicular and luteal phases of the menstrual cycle in nonacclimatized women shivering at a low intensity. The onset (～25 min), intensity (～15% of maximal voluntary contraction), and pattern (～6 shivering bursts/min) of the shivering response did not differ between menstrual cycle phases, regardless of differences in core temperature and hormone levels. This resulted in lipids remaining the predominant substrate, contributing 75% of total heat production, independent of menstrual phase. We conclude that hormone fluctuations inherent in the menstrual cycle do not affect mechanisms of substrate utilization in the cold. Whether the large contribution of lipids to total heat production in fuel selection confers a survival advantage remains to be established.     

Electrical activity of the amygdala and the septum of rats on a high fat or a high protein diet

The authors studied biopotentials in the region of the amygdala and the septum of rats fed on a standard, high protein or high fat diet. During the first 3-6 days after changing from the standard to the high protein or high fat diet, a decrease in the amplitude of electrical activity was found in both the regions in question. After 3 days on the high fat or the high protein diet, an increase was found in the frequency of electrical activity in the amygdala or the septum, according to the type of diet. A study of the amplitude of electrical activity showed that the electrical potential in the septum was always lower than in the amygdala. The frequency spectrum analysis showed a marked change in the superimposed frequency curve only in animals fed on the high fat diet.     

Respiratory adjustments of the unanaesthetized chicken, Gallus domesticus, to elevated metabolism elicited by 2,4-dinitrophenol or cold exposure

The effects of pharmacologically elevated metabolism on ventilation, gaseous exchange and blood gases were studied in spontaneously breathing unanaesthetized decerebrate chickens using 2,4-dinitrophenol (DNP) injected intravenously in successive single doses of 2.5-5.0 mg/kg. These responses were compared with the cardiorespiratory adjustments to elevated metabolism evoked by shivering in conscious birds. Oxygen consumption increased with cumulative amounts of DNP, reaching 275 +/- 30% of control values at the maximum tolerated dose of 10-15 mg/kg. Increases in ventilation matched the changes in oxygen consumption via increases in both breathing frequency and tidal volume. Arterial blood gases and pH remained unchanged. Exposure to cold (Ta = 2 +/- 2 degrees C) caused oxygen consumption to increase to 185 +/- 21% of control values. Respiratory and cardiovascular adjustments were similar to those evoked by DNP and were comparable to those produced by low intensity treadmill exercise (cf. Gleeson and Brackenbury, 1984).     

Involvement of serotonergic pathways in postanaesthetic cold defence: dolasetron prevents shivering

Serotonin is regarded as an essential neurotransmitter in thermoregulation. We evaluated the influence of the serotonergic antagonist dolasetron on thermoregulation and postanaesthetic shivering in mildly hypothermic patients in a randomised placebo-controlled study. The incidence of shivering was 24% after dolasetron compared to 69% after placebo ( p<0.01). During the first 80 min postoperatively, core temperature remained lower in the treatment group ( p<0.05). After dolasetron, 20% of the patients experienced thermal discomfort compared to 54% after placebo ( p<0.05). Serotonin antagonism seems to lower the human thermal set-range thereby reducing metabolic cold defence and discomfort associated with postoperative hypothermia.     

Observations on the effect of salicylate in fever and the regulation of body temperature against cold.

Prostaglandins appear to be mediators, within the hypothalamus, of heat production and conservation during fever. We have investigated a possible role of prostaglandins in the nonfebrile rabbit during thermoregulation in the cold. Shorn rabbits were placed in an environment of 20 degrees C, and rectal and ear skin temperatures, shivering and respiratory rates were measured. A continuous intravenous infusion of leucocyte pyrogen was given to establish a constant fever of approximately 1 degree C, and after observation of a stable febrile temperature for 90 min, a single injection of 300 mg of sodium salicylate, followed by a 1.5 mg/min infusion was then given. After the salicylate infusion was begun, rectal temperature began to fall, and reached nonfebrile levels within 90 min. Shivering activity ceased, respiratory rates increased, and in two animals, ear skin temperature increased. When these same rabbits were placed in an environment of 10 degrees C, at a time they were not febrile, and an identical amount of salicylate was given, rectal and ear skin temperatures, shivering and respiratory rates did not change. These results indicate that prostagladins do not appear to be involved in heat production and conservation in the nonfebrile rabbit.