Interaction of rate of latent heat removal during freezing and rates of subsequent cooling and warming on survival of the blood protozoan, Babesia rodhaini

Babesia rodhaini parasites in murine blood containing 1.5 m DMSO were frozen at two rates, as judged by the duration of the “freezing plateau”, then cooled to ?196 °C and rewarmed at two rates to detect interactions between the duration of the plateau and rates of subsequent cooling and rewarming. Infectivity tests showed that fast and slow freezing (plateau times of about 1 sec and 30 sec, respectively) had similar effects on parasite survival when cooling was at 130 °C/min and warming was at 800 °C/min. However, when either the cooling rate was increased to 3500 °C/min or the warming rate was decreased to 2.3 °C/min, fast freezing decreased parasite survival more than did slow freezing. It is suggested that fast freezing accentuated the damaging effects of fast cooling and slow warming by increasing intracellular ice formation.     

Effects of microwave exposure and temperature on survival of mice infected with Streptococcus pneumoniae

Female CD-1 mice were injected with an LD₅₀ dose of Streptococcus pneumoniae and then exposed to 2.45 GHz (CW) microwave radiation at an incident power density of 10 mW/cm² (SAR = 6.8 W/kg), 4 h/d for 5 d at ambient temperatures of 19 °C, 22 °C, 25 °C, 28 °C, 31 °C, 34 °C, 37 °C and 40 °C. Four groups of 25 animals were exposed at each temperature with an equal number of animals concurrently sham-exposed. Survival was observed for a 10-d period after infection. Survival of the sham-exposed animals increased as ambient temperature increased from 19 °C–34 °C. At ambient temperatures at or above 37 °C the heat induced in the body exceeded the thermoregulatory capacity of the animals and deaths from hyperthermia occurred. Survival of the microwave-exposed animals was significantly greater than the shams (～20%) at each ambient temperature below 34 °C. Based on an analysis of the data it appears that the hyperthermia induced by microwave exposure may be more effective in increasing survival in infected mice than hyperthermia produced by conventional methods (ie, high ambient temperature). Microwave radiation may be beneficial to infected animals at low and moderate ambient temperatures, but it is detrimental when combined with high ambient temperatures.     

The effect of low- and high-power microwave irradiation on in vitro grown <Emphasis Type="Italic">Sequoia</Emphasis> plants and their recovery after cryostorage

Two distinct microwave power levels and techniques have been studied in two cases: low-power microwave (LPM) irradiation on in vitro Sequoia plants and high-power microwave (HPM) exposure on recovery rates of cryostored (?196°C) Sequoia shoot apices. Experimental variants for LPM exposure included: (a) in vitro plants grown in regular conditions (at 24 ± 1°C during a 16-h light photoperiod with a light intensity of 39.06 μEm^?2 s^?1 photosynthetically active radiation), (b) in vitro plants grown in the anechoic chamber with controlled environment without microwave irradiation, and (c) in vitro plants grown in the anechoic chamber with LPM irradiation for various times (5, 15, 30, 40 days). In comparison to control plants, significant differences in shoot multiplication and growth parameters (length of shoots and roots) were observed after 40 days of LPM exposure. An opposite effect was achieved regarding the content of total soluble proteins, which decreased with increasing exposure time to LPM. HPM irradiation was tested as a novel rewarming method following storage in liquid nitrogen. To our knowledge, this is the first report using this type of rewarming method. Although, shoot tips subjected to HPM exposure showed 28% recovery following cryostorage compared to 44% for shoot tips rewarmed in liquid medium at 22 ± 1 °C, we consider that the method represent a basis and can be further improved. The results lead to the overall conclusion that LPM had a stimulating effect on growth and multiplication of in vitro Sequoia plants, while the HPM used for rewarming of cryopreserved apices was not effective to achieve high rates of regrowth after liquid nitrogen exposure.     

Characteristics of microwave evoked body movements in mice

Microwave evoked body movements were studied in mice. A resonant cavity was used to provide head and neck exposure of the mouse to pulsed and gated continuous wave (CW) 1.25 GHz microwaves. No difference in response to pulsed and gated CW stimuli of equal average power was found. The incidence of the microwave evoked body movements increased proportionally with specific absorption (dose) when the whole-body average specific absorption rate was at a constant level (7300 W/kg). Under a constant average specific absorption rate, the response incidence reached a plateau at 0.9 kJ/kg. For doses higher than 0.9 kJ/kg, response incidence was proportional to the specific absorption rate and reached a plateau at 900 W/kg. Body movements could be evoked by a single microwave pulse. The lowest whole-body specific absorption (SA) tested was 0.18 kJ/kg, and the corresponding brain SA was 0.29 kJ/kg. Bulk heating potentials of these SAs were less than 0.1 °C. For doses higher than 0.9 kJ/kg, the response incidence was also proportional to subcutaneous temperature increment and subcutaneous heating rate. The extrapolated absolute thresholds (0% incidence) were 1.21 °C temperature increment and 0.24 °C/s heating rate. Due to high subcutaneous heating rates, these microwaves must be perceived by the mouse as an intense thermal sensation but not a pain sensation because the temperature increment was well below the threshold for thermal pain. Results of the present study should be considered in promulgation of personnel protection guideline against high peak power but low average power microwaves. © 1994 Wiley-Liss, Inc.     

Effect of cooling and rewarming rates on glycerolated human erythrocytes.

Human erythrocytes suspended in a sodium-free buffered salt solution containing glycerol in 1 m concentration (1 part of packed cells to 4 parts buffered salt solution) were frozen by slow, moderately rapid, or very rapid cooling to various subzero C temperatures. The frozen specimens, after a 5-min storage period at a given temperature, were thawed at low, moderately high, or very high rates. The hemolysis in the frozen and thawed samples was measured by a colorimetric determination of the hemoglobin released from the damaged cells. At ?10 °C, the highest freezing temperature employed, nearly 100% recovery of intact erythrocytes was obtained irrespective of the cooling and rewarming conditions. The extent of the hemolysis after exposure to lower freezing temperatures depended upon the cooling and rewarming conditions. Moderately rapid and very rapid freezing to, and thawing from temperatures below ?40 °C permitted significantly higher recoveries of intact cells than the other freezing/ thawing combinations. In the temperature range ?15 to ?30 °C the combination slow cooling and slow rewarming afforded maximum protection. Very rapid freezing/ slow thawing was the most damaging combination throughout the entire freezing range. The results were interpreted in part by a conventional two-factor analysis, lower cooling rates allowing concentrated salts to determine hemolysis, higher cooling rates destroying the cells by intracellular freezing. Apparent anomalies were explained in terms of a generalized “thermal/osmotic” shock according to which the erythrocytes were subject to greater hemolysis the higher the rates of cooling and/or warming.     

The effect of cooling and warming rate on cortical cell function of glycerolized rabbit kidneys

Experiments previously reported (I. A. Jacobsen, D. E. Pegg, H. Starklint, J. Chemnitz, C. J. Hunt, P. Barfort, and M. P. Diaper, Cryobiology19, 668, 1982) suggested that rabbit kidneys permeated with 2 M glycerol are least damaged during freezing and thawing if they are cooled very slowly (1 °C/ hr). Using similar techniques of glycerolization, cooling, storage at ?80 °C, rewarming, and deglycerolization, active cell function in cortical tissue slices prepared from such kidneys has now been studied. Oxygen uptake, tissue $\text{K}{}^{\mathrm{+}}\text{Na}{}^{\mathrm{+}}$ ratio after incubation, and slice/medium PAH ratio after incubation were measured. Kidneys cooled at 3.1 °C/min and warmed at 4.2 °C/min gave poor results in the previous studies and the lowest levels of cell function in the present experiments. Kidneys cooled at 1 °C/hr exhibited degrees of slice function that were dependent on warming rate: warming at 1 °C/min was better than warming at either 1 °C/hr or c.20 °C/min. These results refine the previously drawn conclusions, (loc cit) and indicate optimal cooling and warming rates for rabbit kidneys containing 2 M glycerol, in the region of 1 °C/hr cooling and 1 °C/min warming. These rates are much lower than have hitherto been used by others for any system. Some implications of these findings are discussed.     

The effect of cooling and warming rates on the survival of cryopreserved L-cells

A tissue culture assay has been used to measure the survival of murine lymphoma cells (L-cells) after freezing and thawing in the presence of 2 M glycerol or 1.6 M dimethyl sulfoxide. The effect of variations in cooling rate (0.1 to 10.0 °C/min) and warming rate (0.3 to 200 °C/min) were studied. It was found that survival exhibited a peak at the “conventional” combination of slow cooling and rapid warming (~1 and 200 °C/ min, respectively). It was also shown, however, that a second peak of similar magnitude occurred when the cells were cooled and rewarmed at 0.2-0.3 °C/min. These results are interpreted on the basis of current theories of freezing injury, stressing the importance of damage produced by the recrystallization of intracellular ice and by solute loading. The ultraslow rates of cooling and rewarming which produced the second survival peak are practicable for whole organs, and their potential importance for organ cryopreservation is apparent.     

Strategies of freeze avoidance in larvae of the goldenrod gall moth,Epiblema scudderiana: Laboratory investigations of temperature cues in the regulation of cold hardiness

Laboratory manipulations of ambient temperature were used to investigate the role of temperature in triggering or modulating cold-hardiness adaptations, supercooling-point depression and cryoprotectant accumulation, in larvae of the goldenrod gall moth, Epiblema scudderiana (Clemens), a freeze-intolerant species. Low temperature strongly facilitated cryoprotectant synthesis; larvae subjected to a 1°C per day decrease in temperature showed a major increase in the rate of glycerol synthesis when temperature fell below 5°C with highest rates of synthesis, greater than 90 μmol g⁻¹ d⁻¹, at temperatures between 0 and −10°C. Conversely, abrupt rewarming of larvae from −18 to 23°C in mid-November stimulated a rapid loss of glycerol (from a starting level of 1763 ± 278 μmol/g wet weight) with a half time of only 1.5 days. Supercooling-point depression was not keyed to ambient temperature but appeared to be an endogenous event occurring over the same time interval in laboratory animals held at warm or cold temperatures, as well as in outdoor animals. Rewarming of cold-adapted larvae in November resulted in only a small rise in supercooling point (and did not break diapause) but rewarming in February resulted in a 19°C increase in supercooling point in 4 days, followed rapidly by pupation.     

Survival of tissue culture cells frozen by a two-step procedure to -196 degrees C. II. Warming rate and concentration of dimethyl sulphoxide.

Chinese hamster tissue culture cells in dimethyl sulphoxide (5%) required a lower holding temperature (?35 °C) for optimal survival on slow warming from ?196 °C using a two-step cooling schedule, compared with that required (?25 °C) when warming was rapid. A lower concentration of dimethyl sulphoxide (1%) did not affect the “protection” against damage on cooling from the holding temperature to ?196 °C and thawing. The results suggest that protective agents allow cells to be cooled initially to the holding temperature and minimize damage at the holding temperature. Damage following subsequent cooling and thawing may thus occur mainly as dilution shock on rewarming. This can be minimized by allowing the cells to shrink at the holding temperature.     

Thin-layer liquid crystal thermometry of cells in vitro during hyperthermal microwave irradiation

A nonperturbing technique of thin-layer liquid crystal thermometry was developed to quantitate heating of Chinese hamster ovary cells and the bacterium Serratia marcescens when exposed to 2450-MHz microwave fields at 0.2–0.5 W/cm². Cells suspended in culture medium were injected into 5-cm glass microcapillary tubes coated on the inside with a thin layer of liquid crystal. The tubes were sealed and placed parallel to the electric field in a watertight waveguide exposure chamber where they were heated by circulating temperature-controlled water. Even at high circulation rates, liquid crystal color changes indicated local microwave capillary tube heating of 0.1–0.25 °C. Precision of measurement was 0.02 °C. Observations during microwave heating were significantly different from observations without microwaves at the 1% level, and heating increased as circulating water flow was reduced from 300 ml/s to 100 ml/s. The results of a cell survival assay following hyperthermal treatment were in good agreement with expectations based on the observations of microwave heating using liquid crystals.     

Fluorescence depolarization studies of red cell membrane fluidity. The effect of exposure to 1.0-GHz microwave radiation

The internal viscosity of human red blood cell membranes was investigated during exposure to continuous wave 1.0-GHz microwave radiation using fluorescence measurements of a lipid seeking molecular probe, diphenylhexatriene. Samples were exposed in a Crawford cell arranged so that fluorescence was measured during microwave exposure; specific absorption rates calculated from electrical measurements were approximately 0.6, 2 and 15 W/kg. Measurements were obtained at selected temperatures between 15 °C and 40 °C and as a function of the duration of exposure at 23 °C. Arrhenius-type plots of the temperature profile data were linear and showed no difference between exposed and control samples. The exposure duration data also showed no difference between exposed and control samples except for a small effect of elevated temperature at the highest exposure. The activation energy for motion of the fluorescent probe in its environment within the membrane lipid was not affected by the application of the microwave energy and no evidence for a lipid phase transition was found. These results indicate that the increased cation efflux from red cells, observed by others at certain transition temperatures during microwave exposure, was more likely to have been caused by alteration of the membrane bound protein than by changes in the lipid constituents of the red cell membrane.     

Temperature-dependent hysteresis in somatosensory and auditory evoked potentials

Fourteen adult patients undergoing open heart surgery under induced hypothermia had median nerve, short-latency somatosensory evoked potentials (SSEPs) recorded during cooling (from 36°C to 19°C) and subsequent rewarming. Similar data on another group of patients who had brain-stem auditory evoked potentials (BAEPs) were also analyzed. Hypothermia produced increased latencies of the major SSEP and BAEP components and the latencies returned to normal with subsequent warming. The temperature-latency relationship during the cooling phase was significantly different from that during the warming phase. For SSEP components the temperature-latency relationship was linear during cooling and curvilinear during warming, whereas for BAEP it was curvilinear both during cooling and warming. Furthermore, the regression curves were different during the two phases of temperature manipulation, particularly for temperatures below 30°C both for SSEP and BAEP components. At the onset of warming there was an initial exaggerated warming response on the evoked potential (EP) latencies and amplitude of the EP components. The temperature-latency regression curves were uniformly less steep during the warming phase compared to those during cooling. These findings suggest the existence of hysteresis in the relationship between temperature and EP latencies. The latencies at a given temperature below 30°C depend on whether that temperature is reached during cooling or during warming.     

Influence of various cooling and warming temperatures on survival after thawing of cryopreserved Plasmodium berghei

Heavy concentrations of viable P. berghei in the natural milieu [20% ($\text{v}\text{v}$) parasitized red blood cells, or 20% ($\text{w}\text{v}$) homogenate of splenic tissue in which malarial cells sequestered wer suspended in a serum-free, protective medium. Various rates of cooling are designated as low (1.3 °C/min) and intermediate (4 °C/ min) on exposure in cold gas evolving from liquid nitrogen refrigerant to ?70 °C, and this followed by direct immersion in the low temperature refrigerant (?196 °C). Cooling designated high was accomplished by abrupt immersion of the sealed vials with the live malaria-bearing tissue in the liquid nitrogen refrigerant. Rates of warming and thawing were designated low (after slow rewarming of frozen tissue in air at 25.5 °C) and high (after rapid rewarming and thawing in a water bath at 40 °C). Strip chart recordings were made of the complete cooling and freezing wave patterns of the suspending medium to ?70 ° C. The functional survivals of the freeze-thaw P. berghei malaria were measured by a special infectivity titration method.None of the cooling and freezing treatments adversely influenced the parasite survivals. Our data showed the optimum cooling velocity that maximally protected this highly lethal P. berghei strain within the host erythrocyte matrix was 1.3 ° C/min to ?70 to ?196 ° C. The functional survivals of two RBC stabilates with P. berghei, after retrieval from 25 days storage in the liquid nitrogen refrigerant, excelled by more than 100-fold the infectivity titer found by viability assay in the pool of the 0-days nonfrozen infected RBC.The precise factors favoring the maximal survivals of the freeze-thaw P. berghei are unclear. Several factors, singly or in combination, may have played key roles in protecting the living P. berghei from the freeze-thaw damage. These factors are: The composition of the suspending medium fortified by additions of bicarbonate, glucose, lactalbumin hydrolysate and yeastolate; the presence of naturally occurring peptide-containing materials surrounding the parasites in the host red cell milieu; and the protective glycerol agent. Any of these constituents singly or combined possess potential for reducing freeze-thaw injury to the parasites to produce maximal survivals.     

Efficacy of forced-air and inhalation rewarming by using a human model for severe hypothermia

Goheen, M. S. L., M. B. Ducharme, G. P. Kenny, C. E. Johnston, John Frim, Gerald K. Bristow, and Gordon G. Giesbrecht. Efficacy of forced-air and inhalation rewarming by using a humanmodel for severe hypothermia. J. Appl.Physiol. 83(5): 1635-1640, 1997.We recentlydeveloped a nonshivering human model for severe hypothermia by usingmeperidine to inhibit shivering in mildly hypothermic subjects. Thisthermal model was used to evaluate warming techniques. On threeoccasions, eight subjects were immersed for ~25 min in 9°C water.Meperidine (1.5 mg/kg) was injected before the subjects exited thewater. Subjects were then removed, insulated, and rewarmed in anambient temperature of 20°C with either1) spontaneous rewarming (control),2) inhalation rewarming withsaturated air at ~43°C, or 3)forced-air warming. Additional meperidine (to a maximumcumulative dose of 2.5 mg/kg) was given to maintain shiveringinhibition. The core temperature afterdrop was 30-40% less duringforced-air warming (0.9°C) than during control (1.4°C) andinhalation rewarming (1.2°C) (P < 0.05). Rewarming rate was 6- to 10-fold greater during forced-airwarming (2.40°C/h) than during control (0.41°C/h) andinhalation rewarming (0.23°C/h) (P < 0.05). In nonshivering hypothermic subjects, forced-air warming provided a rewarming advantage, but inhalation rewarming did not.

     

At the tipping point: Differential influences of warming and deoxygenation on the survival,emergence, and respiration of cosmopolitan clams

Although warming and low dissolved oxygen (DO) levels are co‐occurring significant climatic stressors in the ocean, the combined effects of these stressors on marine benthic animals have not been well established. Here, we tested the effects of elevated temperatures and low dissolved oxygen levels on the survival, emerging behavior from sediment, and the respiration of juvenile cosmopolitan Manila clams (Venerupis philippinarum) by exposing them to two temperatures (20 and 23.5°C) and DO levels (3.5 and 6–7 mg/L). Although within previously described tolerable ranges of temperature and DO, this 3.5°C increase in temperature combined with a 50% decrease in DO had a devastating effect on the survival of clams (85% mortality after 8 days). The mortality of clams under normoxia at 23.5°C appeared to be higher than under the low DO condition at 20°C. On the other hand, more clams emerged from sediment under the low DO condition at 20°C than under any other conditions. Oxygen consumption rates were not significantly affected by different conditions. Our results suggest temperature elevation combined with low oxygen additively increases stress on Manila clams and that warming is at least as stressful as low DO in terms of mortality. However, low DO poses another threat as it may induce emergence from sediment, and, thus increase predation risk. This is the first evidence that a combination of warming and deoxygenation stressors should reduce population survival of clams much more so than changes in a single stressor.     

Survival of frozen-thawed human red cells as a function of cooling and warming velocities

Human red cells were equilibrated for 30 min at 20 °C in buffered saline containing 2 m glycerol and then frozen to ?196 °C at 0.27, 1.7, 59, 180, 480, 600, and 1300 °C/ min and warmed at 0.47, 1, 26, 160, and 550 °C/min. Cells frozen at 600 and 1300 °C/min responded in the classical fashion for cells containing intracellular ice; i.e., survivals were low when warming was slow (<10%), but increased progressively with increasing warming rate. The sensitivity to slow warming presumably reflects the recrystallization of intracellular ice. Cells frozen at 59 and 180 °C/ min yielded high survivals at all warming rates. This response is also consistent with the findings for other cells cooled just slowly enough to preclude intracellular ice. Cells frozen very slowly at 0.27 and 1.7 °C/ min, however, responded differently; survivals were considerably higher when warming was slow (0.47 or 1 °C/min) than when it was 26, 160, or 550 °C/min. This response is analogous to that observed recently by others in mouse embryos and in higher plant tissue-culture cells and to that observed for many years in higher plants. It also confirms previous observations of Meryman in human red cells. It may reflect osmotic shock from rapid dilution but, if so, the basis of the osmotic shock is uncertain.     

Effect of freezing and freeze-drying on the viability and storage of Lilium longiflorum L. and Zea mays L. pollen

The freeze-preservation of pollen is dependent on the interaction of several factors such as freezing rate, thawing rate, freeze-drying temperature and duration, storage temperature and environment and rehydration rates. Changes in any of these variables affects the others directly or indirectly.Rapid freezing of pollen at rates of approximately 200 °C/min maintains the highest degree of viable pollen in combination with rapid thawing rates of 218 °C/min. Rapid cooling and slow rewarming resulted in a substantial loss of pollen viability. This might indicate that intracellular ice crystals formed during rapid cooling perhaps grow into larger ice masses during slow rewarming or storage at temperatures above ?50 °C.The germinability of pollen freeze-dried at temperatures below ?50 °C was also prolonged over that of the controls. Germination values for unfrozen pollen stored for 30 days at 0–5 °C averaged 50% for lily and 20% for corn. Freeze-dried pollen stored for 30 days at the same temperature yielded considerably higher viability percentages for both lily and corn pollen. Drying time is an important factor, perhaps indicating that residual moisture is critical. Freeze-dried pollen can be stored at higher temperatures than frozen and control pollen. Freeze-dried material stored for five months at 0–5 °C, upon slow rehydration yielded intact grains which has average germination percentages of 25 for lily and 15 for corn. The same pollen upon rapid rehydration showed rupturing of 20–40% of the cells and practically no germination.     

Differential cerebral hypothermia

Differential cerebral hypothermia was induced in these experiments by isolating the cerebral circulation in the halothane-anesthetized goat. The brain was perfused through isolated cerebral branches of the internal maxillary artery using a height-adjusted reservoir system which provided a constant inflow pressure. Cerebral blood flow (CBF) and cerebral O₂ metabolic rate (CMRO₂) were measured continuously as brain temperatures were decreased from 38 to 28, 18 and 8 °C and during rewarming. Arterial blood gases were maintained constant. During hypothermia CBF decreased at brain temperatures of 28 °C and did decrease further at 18 or 8 °C. CMRO₂ decreased linearly from 38 to 8 °C and was 7% control levels at 8 °C. CBF and CMRO₂ returned to control levels upon rewarming. Cerebral lactate metabolism did not change significantly during hypothermia or rewarming. Evoked cortical potentials were abolished at 8 °C but recovered upon rewarming. These results indicate that if adequate brain perfusion is maintained during hypothermia and rewarming, recovery of CBF, metabolism, and brain neural activity can be obtained.     

Studies on microwave and blood-brain barrier interaction

This investigation was aimed at correlating changes of blood-brain-barrier permeability with the quantity and distribution of absorbed microwave energy inside the brain of adult Wistar rats anesthetized by sodium pentobarbital. Through use of thermographic methods and a direct-contact applicator at the animal's head, the pattern of absorbed microwave energy was determined. Indwelling catheters were placed in the femoral vein and in the left external carotid artery. Evans blue and sodium fluorescein in isotonic saline were used as visual indicators of barrier permeation. Exposure to pulsed 2,450-MHz radiation for 20 min at average power densities of 0.5, 1, 5, 20, 145 or 1,000 mW/cm², which resulted in average specific absorption rates (SARs) of 0.04, 0.08, 0.4, 1.6, 11.5 or 80.0 mW/g in the brain, did not produce staining, except in the pineal body, the pituitary gland, and the choroid plexus — regions that normally are highly permeable. Except for these regions, staining was also absent in the brains of sham-exposed animals. The rectal temperature, as monitored by a copper-constantan thermocouple, showed a maximum increase of less than 0.75°C from a mean pre-exposure temperature of 36.6°C. The highest brain temperature recorded in a similar group of animals using a thickfilm carbon thermistor was less than 41.0°C.     

Cardiovascular effects of levosimendan during rewarming from hypothermia in rat

BackgroundPrevious research aimed at ameliorating hypothermia-induced cardiac dysfunction has shown that inotropic drugs, that stimulate the cAMP, – PKA pathway via the sarcolemmal β-receptor, have a decreased inotropic effect during hypothermia. We therefore wanted to test whether levosimendan, a calcium sensitizer and dose-dependent phosphodiesterase 3 (PDE3) inhibitor, is able to elevate stroke volume during rewarming from experimental hypothermia.MethodsA rat model designed for circulatory studies during experimental hypothermia (4 h at 15 °C) and rewarming was used. The following three groups were included: (1) A normothermic group receiving levosimendan, (2) a hypothermic group receiving levosimendan the last hour of stable hypothermia and during rewarming, and (3) a hypothermic placebo control group. Hemodynamic variables were monitored using a Millar conductance catheter in the left ventricle (LV), and a pressure transducer connected to the left femoral artery. In order to investigate the level of PKA stimulation by PDE3 inhibition, myocardial Ser23/24-cTnI phosphorylation was measured using Western-blot.ResultsAfter rewarming, stroke volume (SV), cardiac output (CO) and preload recruitable stroke work (PRSW) were restored to within pre-hypothermic values in the levosimendan-treated animals. Compared to the placebo group after rewarming, SV, CO, PRSW, as well as levels of Ser23/24-cTnI phosphorylation, were significantly higher in the levosimendan-treated animals.ConclusionThe present data shows that levosimendan ameliorates hypothermia-induced systolic dysfunction by elevating SV during rewarming from 15 °C. Inotropic treatment during rewarming from hypothermia in the present rat model is therefore better achieved through calcium sensitizing and PDE3 inhibition, than β-receptor stimulation.