Alteration of heat sensitivity by treatment with nonpermissive temperature or cycloheximide in temperature-sensitive CHO mutant tsH1 cell

1. 1.|The temperature-sensitive mutant CHO-tsH1 and wild type (CHO-SC) cells became thermal resistant when cells were treated for either 2 h at 39.5°C before heating at 43°C or 2 h with 10 μg/ml cycloheximide (CHM) before and during heating at 43°C.

2. 2.|There was a 2000-fold increase in survival after 2.5 h at 43°C by preincubation at 39.5°C in both cell types. There was also a 200- or 700-fold increase in survival after 2.5 h at 43°C by treatment with CHM in tsH1 or SC cell type respectively.

3. 3.|In contrast to the effects at 43°C, at 41.8°C these protective effects were not evident in tsH1 cells. In wild type, however, there was an 800- or 1800-fold increase in survival after 8 h at 41.8°C by preincubation at the temperature of 39.5°C or treatment with CHM, respectively.

4. 4.|Therefore, these results suggest that killing of tsH1 at low temperature hyperthermia (41.8°C) is probably due to denaturation of thermolabile leucyl-tRNA synthetase.

5. 5.|The denaturation of this enzyme may not be protected by inhibition of protein synthesis by preincubation at the nonpermissive temperature of 39.5°C or by CHM.

Mechanisms of heat protection by cycloheximide or puromycin: Involvement of polyamines?

1. 1.|When Chinese hamster ovary cells are treated with cycloheximide (10 μg/ml) or puromycin (100 μg/ml) for 2 h before and during heating at 43°C for 3 h, there is protection from hyperthermic killing; i.e. the plating efficiency increases 2000-fold from 3.7 × 10⁻⁵ to (6–9) × 10⁻².

2. 2.|The total intracellular levels of spermidine and spermine are not altered by the hyperthermic or drug treatments.

3. 3.|The small 30% decrease in intracellular putrescine observed after heating is not altered by drug treatment.

4. 4.|Heat protection by treatment with cycloheximide or puromycin cannot be attributed to changes in levels of total intracellular polyamines.

Employment of a turbidimetric assay system to measure heat-induced protein aggregation

1. 1.|We developed a turbidimetric assay system for quantitation of heat-induced protein aggregation which is presumably caused by protein denaturation.

2. 2.|Rhodanese in 6 M guanidinium chloride was employed in the assay system, because this protein recognizes hydrophobic sites on denatured proteins and aggregates.

3. 3.|Turbidity caused by protein-rhodanase aggregation was recorded at 320 nm by using a u.v./VIS spectrophotometer.

4. 4.|When heated, alcohol dehydrogenase (ADH) aggregates with rhodanese. The increase of ADH-rhodanese aggregation was correlated with the loss of enzymatic activity.

5. 5.|These results indicated that the aggregation was proportional to the extent of ADH denaturation which assumingly caused the loss of ADH activity during heating at 45.5°C.

6. 6.|Similar results were observed when cytosolic proteins from CHO cells were heated at 45.5°C. Heated cytosolic proteins promoted aggregation by complex formation with rhodanese. The aggregation increased with increasing heat dose.

7. 7.|Therefore, the rhodanese assay system can be employed usefully to quantitate the protein aggregation after heat stress.

Thermohaemolysis of human erythrocytes in isotonic NaCl/sucrose media during transient heating

1. 1.|The hyperhermia induced haemolysis of cells and resealed ghosts suspended in isotonic NaCl/sucrose media was studied upon transient heating.

2. 2.|At 61.5°C a process of temperature accelerated disturbance of membrane permeability barrier was initiated, wich was sensed by the consequent volume changes. Concomitantly with this process the thermohaemolysis appeared as a threshold colloid-osmotic lysis.

3. 3.|The initial temperature of this successive barrier disturbance was decreased linearly by ethanol. At 18% ethanol this barrier disturbance took place at 39°C while spectrin was denaturated at about 45°C. Apparently, the spectrin denaturation was not sufficient, nor was involved in, the initiation of this membrane disturbance.

4. 4.|The membrane of cells made ion permeable in the presence of 18% ethanol by heating to 39°C contained irreversible pores with a radius of about 0.45 nm.

5. 5.|This suggests a conformational change of a protein(s) in their formation, but not spectrin nor the anion channel.

6. 6.|Using specific amino reagents it was ascertained, that a superficial NH₃⁺ group dissociable at neutral pH impeded this thermo-induced pore formation.

7. 7.|Consistent results show that this formation of membrane pores initiated at 61.5°C may be included in the still unknown mechanism of thermohaemolysis.

Hyperthermia-induced changes in the morphology of CHO-K1 and their refractile inclusions

1. 1.|Chinese hamster ovary cells (CHO-K1) were heated at temperatures of 42°C and above.

2. 2.|Cells were cultured in microcapillaries to eliminate handling stress, and morphological changes were observed by light microscopy.

3. 3.|Increased incidence of membrane blebbing was noted between 1 and 2 h and few cells were viable after 2 h at 43°C.

4. 4.|Morphological changes, including the appearance of potocytotic blebs, were recorded by cinemicroscopy of microcapillary cultures on a heated microscope stage.

5. 5.|Lipid-rich refractile cell inclusions changed shape before blebbing occurred.

6. 6.|Cell retraction and rearrangement of organelles seen at 1 h at 43°C are the reverse of those seen in spreading post-trypsinized cells and suggest a thermal effect on the cytoskeleton.

The effect of incubation temperatuer on oxygen consumption and organ growth in domestic-fowl embryos

1. 1.|Oxygen consumption and organ growth were measured in domestic-fowl embryos incubated at different temperatures (36, 38 and 40°C).

2. 2.|Embryonic oxygen consumption was highest at an incubation temperature of 40°C and lowest at 36°C. These differences were ascribed largely to variations in embryo size at different incubation temperatures.

3. 3.|At incubation temperatuers of 40 and 38°C, there was a plateau in oxygen consumption late in incubation, but this was not apparent at 36°C.

4. 4.|At 36°C, some tissues (e.g. eyeballs) were “spared” the repression of growth that characterized the embryo as a whole, while other tissues (e.g. stomach) incurred a much greater growth reduction. Similarly, at 40°C, stomach growth exceeded that of the embryo as a whole, while the eyeballs were largely spared the enhanced growth.

5. 5.|A simple index of tissue age revealed that, in general, there were consensual changes in tissue maturity and growth at different temperatures but that there were some disparities between growth and maturity in individual organs.

Rapidly reversible enzyme inhibition in a temperature-sensitive mammalian cell mutant lacks thermotolerance

The temperature-sensitive (ts) Chinese hamster ovary (CHO) cell mutant tsH1 contains a thermolabile leucyl-tRNA synthetase. Upon incubation at the nonpermissive temperature of 39.5 degrees C, the enzyme became reversibly inhibited over a period of minutes, and the cells lost viability over a period of many hours. However, killing of tsH1 by acute heating at 45 degrees C was identical to that of wild-type (SC) cells. In addition, the heat-induced inhibition of protein synthesis was similar for both cell types, as measured after acute heating at 45 degrees C. Furthermore, both killing and inhibition of protein synthesis showed thermotolerance in both cell types. In contrast to the effects at 45 degrees C, at 39.5 degrees C, neither the inhibition of leucyl-tRNA synthetase activity nor the killing of tsH1 expressed thermotolerance. Also, treatment of tsH1 at 39.5 degrees C did not induce thermotolerance to killing at 45 degrees C. The inhibition of leucyl-tRNA synthetase activity in tsH1 at 39.5 degrees C was further distinguished from the 45 degrees C-induced inhibition of protein synthesis in SC cells by a much more rapid reversal of the inhibition of leucyl-tRNA synthetase activity. Also, the rate of reversal of the inhibition of protein synthesis by 45 degrees C in SC cells was decreased by increased heat dose. Such was not true for the 39.5 degrees C inhibition of leucyl-tRNA synthetase activity in tsH1. The data indicate that there exist two distinct types of thermal inhibition--one slowly reversible type which was observed during and after heating at 45 degrees C and both induced and expressed thermotolerance, and a second, rapidly reversible type, which was evident only during heating of tsH1 at 39.5 degrees C and neither induced nor expressed thermotolerance.     

Temperature effects on evoked potentials of hippocampal slices from noncold-acclimated, cold-acclimated and hibernating hamsters

1. 1.|Neural activity was recorded in hippocampal slices from noncold-acclimated, cold-acclimated and hibernating hamsters.

2. 2.|Action potentials from a population of hippocampal pyramidal neurons were evoked by stimulating an afferent fiber tract, the Schaffer collaterals. The temperature of the artificial cerebrospinal fluid bathing the slice was varied by controlling the temperature of a water chamber jacketing the recording chamber.

3. 3.|The temperature just below that at which a population spike could be evoked, T_t, was 15.8 ± 0.9°C (mean ± SEM) for noncold-acclimated hamsters, 13.9 ± 0.3°C for cold-acclimated hamsters and 12.3 ± 0.3°C for hibernating hamsters.

4. 4.|These thresholds for evoked activity were significantly different in noncold-acclimated, cold-acclimated and hibernating hamsters, and may reflect acclimation of hippocampal neurons to cold.

Development of thermoregulation in Hawaiian brown noddies (Anous stolidus pileatus)

1. 1.|Oxygen consumption ( ) and body temperture (T_b) of Hawaiian brown noddies (Anous stolidus pileatus [Aves: Laridae]) during late incubation and in the first 24 h after hatching were measured at ambient temperatures (T_a) between 28 and 38°C and between 15 and 43°C, respectively. Evaporative cooling by hatchings at T_a of 36–43°C was also measured.

2. 2.|Throughout the late incubation stages studied, and T_b both varied directly with T_a in an ectothermic pattern.

3. 3.|The hatchlings successfully regulated T_b at T_a between ca. 29 and 43°C.

4. 4.|The functional basis of the abrupt increase in thermoregulatory capacity with hatching is discussed.

Thermohaemolysis of human erythrocytes in sucrose containing isotonic media

1. 1.|The thermohaemolysis of human erythrocytes in NaCl/sucrose isotonic media can be best accounted for in terms of the colloid-osmotic theory of haemolysis.

2. 2.|The thermohaemolysis in NaCl saline was preceded by leakage of K⁺ and cell swelling. If the inner oncotic osmoactivity was balanced with external sucrose the cells progressively shrinked losing K⁺, but the haemolysis was strongly reduced.

3. 3.|Time dependence of the shrinking of cells and one-step resealed ghosts suspended in isotonic 60 mOsm NaCl/sucrose media was studied between 50 and 58°C.

4. 4.|After a lag period for cells only, this shrinking proceeded with apparently constant rate for cells and ghosts.

5. 5.|The rate constant of shrinking for cells and ghosts obeys the Arrhenius relation, giving the value of 250 ± 15 kJ/mol for the activation energy of shrinking in both cases. This is also the case for the activation energy of the membrane ion permeability constant.

6. 6.|These results are consistent with the thermal inactivation of membrane associated protein(s) acting as a trigger for the ion permeability barrier disturbance.

7. 7.|The mid-point temperature for these membrane events was about 61°C.

Effect of temperature on growth and bioenergetics of a tropical moth

1. 1.|Larval development and metamorphosis of Achaea junta were prolonged at 22°C, compared to 27, 32 and 35°C.

2. 2.|Overall rates of consumption, assimilation, production and metabolism of the larvae increased with temperature.

3. 3.|Efficiencies of assimilation and conversion of the digested food were significantly altered by life stage and temperature.

4. 4.|About 60% of the pupal energy was transferred to the imago at the tested temperatures.

Thermosensitivity during embryonic development of Lymnaea stagnalis (Mollusca)

1. 1.|The percentage of survival after 1 hr at 40.0°C is lowest at the larval trochophore stage and at hatching of the young snail.

2. 2.|Heat resistance depends on the stage of development.

3. 3.|From the early cleavage stage onwards a higher percentage of embryos can withstand high temperature after a previous heat treatment than without it.

4. 4.|The pattern of thermosensitivity is discussed in relation to the organizational level of the stage of development.

5. 5.|It is concluded that the developing Lymnaea is a suitable system to study heat resistance and thermotolerance at the level of cells, organs and organism.

Variations in the activity of fatty ACYL-CoA desaturases and electron-transport components in Tetrahymena microsomes with temperature and age of culture

1. 1.|Alterations in the fatty acid composition of microsomes were most marked in the exponential phase of both 39.5- or 15°C- grown Tetrahymena pyriformis NT-1.

2. 2.|Activities of palmitoyl-CoA and stearoyl-CoA desaturases were lower in 15°C cells than in 39.5°C cells, while the activity of oleoyl-CoA desaturase was higher in 15°C cells.

3. 3.|Activities of the terminal component of the desaturation system as well as all three desaturases (palmitoyl-CoA, stearoyl-CoA, oleoyl-CoA) were higher in the exponential phase than in the stationary phase for cells grown at both temperatures.

4. 4.|NAD(P)H-cytochrome c reductase activity and cytochrome b₅ content were reduced whereas NADH-ferricyanide reductase activity was increased in the stationary phase at both 39.5 and 15°C.

Hatching of freshwater sponge gemmules after low temperature exposure: Ephydatia mülleri (Porifera: Spongillidae)

1. 1.|Gemmules of Ephydatia mülleri can withstand exposure to temperatures down to −80°C for 63 days without loss of hatchability.

2. 2.|Hatching is slowed following exposure to temperatures below −27°C.

3. 3.|There is a slight but significant relationship between gemmule size and the time to hatch.

4. 4.|This species can withstand long-term exposure to winter air temperatures occurring within its known geographic range.

Exercise performance of normothermic and hyperthermic rats: Effect of warm rearing

1. 1.|Hypothalamic and rectal temperatures were recorded in 8 warm-reared (wr) and 12 control rats. Rats ran to exhaustion at a constant speed of 1.5 km h⁻¹ but at a variable ambient temperature adjusted to stabilize their hypothalamic temperature at 38.0°C (normothermia) or 41.0°C (hyperthermia). Blood lactate concentrations were determined before and after exercise.

2. 2.|Exercise caused exhaustion in normothermic control rats after 62.08 ± 5.43 min and in wr rats after 29.64 ± 2.09 min.

3. 3.|Hyperthermia shortened to one half (to 12.24 ± 1.36 min) and to one fourth (to 16.15 ± 1.20 min) the endurance time in wr and control rats, respectively.

4. 4.|There were no correlations between lactate concentraion and hyperthermia or endurance time.

5. 5.|In conclusion, in rats and other animals which have safe refuges, hyperthermia interferes with the ability to continue exercising.

The effect of temperature on caecal fermentation processes in a poikilothermic mammal, Heterocephalus glaber

1. 1.|The effect of temperature on caecal function was examined in the naked mole-rat Heterocephalus glaber, a poikilothermic mammal, which consumes a high proportion of fibre in its natural diet.

2. 2.|The temperature of optimal caecal function was determined from fermentation data measure at three specifically chosen temperatures (28, 33 and 40°C).

3. 3.|There was no significant difference between gas production at 33 and 40°C, however, gas production was significantly lower at 28°C.

4. 4.|The relative proportions of the gases produced were markedly different at 33 and 40°C (P ≤ 0.01). More methane and hydrogen were produced at 33°C than at 40°C.

5. 5.|These data suggest that microbial organisms within the caecum were active and functioning more effectively at 33°C (the preferred body temperature of the naked mole-rat) than at the other two temperatures.

Muscle temperature and muscle metabolism during short-term exercise in the goat

1. 1.|External heat exchangers acting on lower aortal blood temperature were used to dissociate hindleg muscle temperature (T_hlm) from general internal temperature (T_int) during short-term exercise of moderate intensity.

2. 2.|In series 1 39°C T_hlm was combined with 40.6°C T_int, and in series II 42°C T_hlm was combined with 39.8°C T_int.

3. 3.|At constant work rates, the 3°C difference in muscle temperature did not result in significantly different concentrations of muscle metabolites.

4. 4.|It is concluded that high local muscle temperature without general hyperthermia does not influence muscle metabolism during short-term moderate excercise.

The effect of acclimation temperature and the removal of peripheral temperature receptors on body temperature preference in the cockroach, Periplaneta americana

1. 1.|Body temperature preferences were compared between cockroaches acclimated to different ambient temperatures and between 25°C acclimated cockroaches and cockroaches deprived of their peripheral temperature receptors.

2. 2.|Acclimation to 35°C resulted in a significantly higher mean body temperature and low body temperature selected compared with 25°C acclimated cockroaches.

3. 3.|Cockroaches deprived of their peripheral temperature receptors showed a significantly higher mean high body temperature selected when compared to normal 25°C acclimated cockroaches.

4. 4.|It is concluded that cockroach temperature regulation is more precise than expected and that central temperature receptors are the primary sensing elements for cockroach thermoregulation.

Autonomic thermoregulatory effects of electrical stimulation of the hypothalamus in the sheep (Ovis aries)

1. 1.|The effects of electrical stimulation of the preoptic region, on autonomic thermoregulatory responses, were studied in conscious sheep at ambient temperatures of 5, 20, and 40°C.

2. 2.|Stimulation of the dorsal preoptic region elicited co-ordinated thermoregulatory responses characterized by increased respiratory frequency (RF), vasodilation of the ears and lowered body temperature. Stimulation inhibited shivering in cold environments.

3. 3.|The thermoregulatory responses were greater at 5°C in unshorn than in shorn sheep. Increased RF, induced at 20 and 40°C, persisted several minutes after stimulation ceased.

4. 4.|Intraventricular injection of noradrenaline reduced both normal and electrically-induced panting.

5. 5.|Sheep would press panels to electrically stimulate the preoptic region and this “self-stimulation” activated heat-loss mechanisms.

Effect of temperature on the electrical activity of the rat epididymis in vitro

1. 1.|Regional differences in the frequency of electrical activity in rat epididymis were maintained at all temperatures below 39°C.

2. 2.|The change in frequency per deg C increased with temperature and was highest in the temperature region of 34–39°C and the Arrhenius plots of the frequency were linear and parallel in all parts of the epididymis.

3. 3.|The Q₁₀ of the frequency varied between 2.2.–4.3.

4. 4.|The conduction velocity at the cauda epididymis was highest (2.8 mm/s) at 37°C. The Q₁₀ of the conduction velocity was 2.3 in the temperature region of 24–37°C.