Investigations of thermotropic phase changes in peripheral nerve of frog and rat. A spin label study

The temperature-dependent fluidity of myelin of frog and rat peripheral nerve (Nervus ischiadicus) was studied using the spin label technique. In frog nerve a phase change was detected at 38 °C. In rat nerve no sharp phase change could be established, and the lipid-depleted frog and rat nerve also showed no transition. From the spectral data, it was concluded that in frog and rat nerve the lipid-protein interactions are different, i.e. species dependent. Ca²⁺-depletion of frog nerve caused a loss of transition, while rat nerve remained unaffected. Thus it was indicated that, in frog nerve, Ca²⁺ is involved in the phase change. In the total lipid extract of frog nerve a phase change centered at 32 °C occurred, while the total lipid extract of rat nerve again showed no transition. It is suggested that a connexion exists between our results and investigations on the temperature dependence of an axonal conduction block of nerve.     

Effects of thermal acclimation on nervous conduction and muscle contraction in the frog Rana temporaria

The effects of season and acclimation temperature on the latency of the leg withdrawal reflex and three of its components have been studied: conduction velocity in the sciatic nerve, spinal conduction time, and contraction time of gastrocnemius muscle. The latency of the leg withdrawal reflex was markedly shortened by cold acclimation: the reaction times were at 6 degrees C 1.54 s in 4 degrees C acclimated and 3.97 s in 24 degrees C acclimated winter frogs. Also, the temperature dependence of the reflex latency was reduced by cold acclimation. Thus, frogs acclimated to cold responded to external stimuli in cold more rapidly than warm-acclimated ones. This cold adaptation of the reflex could not be explained by changes in its studied components. These made up only one-tenth of the reflex response time, and either did not show significant cold acclimation (muscle contraction and spinal conduction times in summer) or showed inverse acclimation, especially when measured at high temperatures (i.e. conduction velocities were reduced by acclimation to cold). Thus, the cold acclimation of the reflex response probably resides in the sensory component of the response. The inverse temperature adaptation response of conduction velocities may reflect a reduced ion permeability across cellular membranes in cold which decreases metabolic energy expenditure during inactive periods.     

Effect of long-term thermal acclimation on the electrical avoidance conditioning of fish

The learning behaviour and memory formation of ectotherms, especially of fish, depend significantly on the acclimation temperature. Although temperature is known to affect different physiological factors in the nervous system, the exact manner in which memory and learning are affected by these alterations is not clear. Large variations in the acclimation time before learning takes place, are striking. With regard to long-lasting compensatory changes in the polarity of membrane-bound neuronal gangliosides (1) and in the bio-electrical activity of the CNS (post-synaptic potential amplitudes (2)) following thermal acclimation it was of interest to investigate the time course of acclimation on the learning ability of fish subjected to a new environmental temperature.     

Seasonal variation in conduction velocity of action potentials in squid giant axons

To determine whether the electrical properties of the squid giant axon are seasonally acclimated, action potentials, recorded at different temperatures, were compared between giant axons isolated from Loligo pealei caught in May, from relatively cold waters (approximately 10 degrees-12 degrees C), and in August, from relatively warm waters (approximately 20 degrees C). Parameters relating to the duration of the action potential (e.g., maximum rate of rise, maximum rate of fall, and duration at half-peak) did not change seasonally. The relationship between conduction velocity and temperature remained constant between seasons as well, in spite of the fact that May axons were significantly larger than August axons. When normalized to the fiber diameter, mean May conduction velocities were 83% of the August values at all temperatures tested, and analysis of the rise time of the action potential foot suggested that a change in the axoplasmic resistivity was responsible for this difference. Direct measurements of axoplasmic resistance further supported this hypothesis. Thus seasonal changes in the giant axon's size and resistivity are not consistent with compensatory thermal acclimation, but instead serve to maintain a constant relationship between conduction velocity and temperature.     

Acute and chronic effects of temperature, and of nutritional state, on ion homeostasis and energy metabolism in teleost hepatocytes

Short- and long-term effects of temperature on ion flux and energy turnover were studied in hepatocytes from thermally acclimated trout and roach. In trout hepatocytes K⁺ efflux was insensitive towards acute exposure to low temperature but was downregulated during cold acclimation of the fish so as to balance the uncompensated decreased K⁺(Rb⁺) uptake of the cells. In contrast, both K⁺(Rb⁺) uptake and K⁺ efflux of roach hepatocytes were temperature sensitive in the short term. These acute effects, however, were offset during cold acclimation by a near perfect compensation of both fluxes leading to re-establishment of ion flux homeostasis at the original level. Our findings, based on a new method permitting the simultaneous monitoring of K⁺ efflux and uptake in the same cell population, provide experimental verification of two of the three possible strategies, recently discussed by Cossins et al. (1995), by which the ionic steady state of fish cells may adjust to acute and chronic temperature change. By comparing hepatocytes from two groups of trout, one kept on a maintenance diet (ration I), the other fed ad libitum (ration II), we discovered striking effects of nutritional state on the absolute levels as well as on the temperature relationships of K⁺ uptake and protein synthetic activity. Both of these functions in the hepatocytes increased in the ration II fed as compared to the ration I fed trouts, but the increase of protein synthetic activity was greater and more uniform at the three experimental temperatures than that of K⁺ uptake. Moreover, protein synthetic activity proved to be considerably more temperature sensitive than K⁺ uptake and, in contrast to the latter, showed a compensatory response after cold acclimation. Accepted: 16 December 1996     

Prostaglandin E1 modifies nerve conduction and interferes with local anaesthetic action.

Prostaglandin E1 (PGE1) altered both the amplitude and conduction velocity of the compound action potential in frog sciatic nerve. Concentrations up to 1 ng/ml increased both amplitude and conduction velocity but at higher concentrations both effects were reversed. Procaine, chloroquine, indomethacin and SC19220 all reduced action potential amplitude and conduction velocity. These local anaesthetic type actions could be partially or completely prevented by PGE1.     

Prostaglandin E1 modifies nerve conduction and interferes with local anaesthetic action

Prostaglandin E1 (PGE1) altered both the amplitude and conduction velocity of the compound action potential in frog sciatic nerve. Concentrations up to 1 ng/ml increased both amplitude and conduction velocity but at higher concentrations both effects were reversed. Procaine, chloroquine, indomethacin and SC19220 all reduced action potential amplitude and conduction velocity. These local anaesthetic type actions could be partially or completely prevented by PGE1.     

Characterization of γ-glutamyltranspeptidase in the liver of the frog: 2. Response to season,temperature and thyroid hormone in Rana pipiens

The impact of season and temperature on frog liver γ-glutamyltranspeptidase was assessed by measuring the activity of this enzyme in plasma membranes isolated from the livers of Rana pipiens obtained as summer and winter frogs; subjected to short-term (3 weeks) temperature acclimation; and subjected to multiple-temperature shifts. Plasma levels of T₃ were determined. γ-Glutamyltranspeptidase was found to be 2·2-fold higher in the summer frog relative to the winter frog; decreased by 44 percent in the summer frog by cold acclimation and increased by 1·7-fold in the winter frog by warm acclimation; and increased by 1·9-fold in the summer frog and 2·8-fold in the winter frog subjected to multiple-temperature shifts. Plasma T₃ levels were found to be 42-fold higher in the summer frog relative to the winter frog; decreased by 42 percent by cold acclimation and increased by 2·9-fold by warm acclimation; and decreased by 39 percent and 38 percent in the summer and winter frogs subjected to multiple temperature shifts. T₃ replacement during the last phase of the multiple-temperature shift protocol, restored the plasma T₃ levels to 75 percent of the control levels and prevented the increase evoked by the multiple-temperature shifts in γ-glutamyl-transpeptidase activity. Indeed, enzyme activity in the T₃ replaced state was 19 percent lower than in the control state. The involvement of thyroid hormone as a negative regulator of enzyme activity is discussed.     

Energy reserves during food deprivation and compensatory growth in juvenile roach: the importance of season and temperature

The effect of 21 days of starvation, followed by a period of compensatory growth during refeeding, was studied in juvenile roach Rutilus rutilus during winter and summer, at 4, 20 and 27° C acclimation temperature and at a constant photoperiod (12L : 12D). Although light conditions were the same during summer and winter experiments and fish were acclimated to the same temperatures, there were significant differences in a range of variables between summer and winter. Generally winter fish were better prepared to face starvation than summer fish, especially when acclimated at a realistic cold season water temperature of 4° C. In winter, the cold acclimated fish had a two to three‐fold larger relative liver size with an approximately double fractional lipid content, in comparison to summer animals at the same temperature. Their white muscle protein and glycogen concentration, but not their lipid content, were significantly higher. Season, independent of photoperiod or reproductive cycle, was therefore an important factor that determined the physiological status of the animal, and should generally be taken into account when fish are acclimated to different temperature regimes. There were no significant differences between seasons with respect to growth. Juvenile roach showed compensatory growth at all three acclimation temperatures with maximal rates of compensatory growth at 27° C. The replenishment of body energy stores, which were utilized during the starvation period, was responsible for the observed mass gain at 4° C. The contribution of the different energy resources (protein, glycogen and lipid) was dependent on acclimation temperature. In 20 and 27° C acclimated roach, the energetic needs during food deprivation were met by metabolizing white muscle energy stores. While the concentration of white muscle glycogen had decreased after the fasting period, the concentrations of white muscle lipid and protein remained more or less constant. The mobilization of protein and fat was revealed by the reduced size of the muscle after fasting, which was reflected in a decrease in condition factor. At 20° C, liver lipids and glycogen were mobilized, which caused a decrease both in the relative liver size and in the concentration of these substrates. Liver size was also decreased after fasting in the 4° C acclimated fish, but the substrate concentrations remained stable. This experimental group additionally utilized white muscle glycogen during food deprivation. Almost all measured variables were back at the control level within 7 days of refeeding.     

Interactions of season and temperature acclimation in the control of metabolism in Amphibia

1. 1.|Studies concerning the seasonal variation and the temperature acclimation of metabolism and their control in Amphibia are reviewed.

2. 2.|Both season and temperature acclimation affect the activities of the central and autonomic nervous systems.

3. 3.|These changes are mediated especially by alterations in the activity of the thyroid and through the autonomic nerves.

4. 4.|The fact that common control mechanisms are involved may explain some of the often observed metabolic interactions of season and temperature acclimation.

Influence of frequency and temperature on the mechanisms of nerve conduction block induced by high-frequency biphasic electrical current

The influences of stimulation frequency and temperature on mechanisms of nerve conduction block induced by high-frequency biphasic electrical current were investigated using a lumped circuit model of the myelinated axon based on Schwarz and Eikhof (SE) equations. The simulation analysis showed that a temperature-frequency relationship was determined by the axonal membrane dynamics (i.e. how fast the ion channels can open or close.). At a certain temperature, the axonal conduction block always occurred when the period of biphasic stimulation was smaller than the action potential duration (APD). When the temperature decreased from 37 to 15 degrees C, the membrane dynamics slowed down resulting in an APD increase from 0.4 to 2.4 ms accompanied by a decrease in the minimal blocking frequency from 4 to 0.5 kHz. The simulation results also indicated that as the stimulation frequency increased the mechanism of conduction block changed from a cathodal/anodal block to a block dependent upon continuous activation of potassium channels. Understanding the interaction between the minimal blocking frequency and temperature could promote a better understanding of the mechanisms of high frequency induced axonal conduction block and the clinical application of this method for blocking nerve conduction.     

The possible functional role of neuronal gangliosides in temperature adaptation of vertebrates

Comparative studies on brain gangliosides of more than 60 vertebrate species show correlations between concentration and the level of evolutionary organization: poikilothermic lower vertebrates (fish, amphibs, reptiles) contain about 110 to 700 μg ganglioside bound NeuAc/g. fresh wt., homeothermic birds and mammals, on the other side, 500 to 1000 μg. The composition of brain gangliosides in poikilotherms is much more complex and variable (more multisialogangliosides) as compared with homeotherms (domination of less polar fractions). There are distinct correlations between brain ganglioside composition and state of thermal adaptation: Fishes being adapted to habitates with extreme temperatures (antarctic icefish — tropic fish) are characterized by quite opposite ganglioside patterns (domination of high versus less polar fractions). During seasonal acclimatization and experimental acclimation of fish to cold or during hibernation and early postnatal development of mammals poly-sialylations of brain gangliosides occur. With regard to this the individual brain structures react differently.

The results are taken for evidence that variations in the composion of synaptic membrane-bound gangliosides may induce long-term alterations in viscosity and permeability of the neuronal membrane by which the neuronal transmission might be kept on a constant level during the process of temperature adaptation.     

Changes in cell membrane fluidity affect the sodium transport across frog skin and its sensitivity to amiloride

1. 1-5 mM n-hexanol added to the outer (mucosal) medium of isolated skin of the frog Rana temporaria increases the short circuit current (Isc) across it. 2. This effect shows a saturable dependency on the outer sodium concentration, also when NaCl is replaced by Na2SO4. 3. n-Hexanol at a concentration of 1 mM, and cold acclimation of the frogs, which increases the fluidity of epidermal cell membranes, do not affect the sensitivity of Isc to the inhibiting effect of amiloride. 4. n-Hexanol at a concentration (5 mM) which causes a fluidization of cell membrane preparations from isolated frog epidermis also increases the sensitivity of Isc to amiloride. 5. The effects of low concentrations of n-hexanol and of cold acclimation probably depend on an increase of the permeability of apical membranes of epidermal cells to sodium caused by membrane fluidization. At higher concentrations of n-hexanol, a further disordering of the membrane structure occurs with a better access of amiloride to its action sites.     

The evolution of endothermy

A theory to account for the evolution of heat production for endothermy in the vertebrates is presented. It is argued that thermoregulatory responses to cold, thyroxine, and the Na⁺ pump are related functionally and phylogenetically.Fish regulate their body temperature behaviorally. For example, if the ambient temperature is too cold, they exhibit appetitive behavior and swim to an area where it is warmer. The incidental heat produced by the increased activity is lost through the gills due to the pattern of circulation. In fish the increased requirement for oxygen during increased activity demands an increased transfer of oxygen and of ions and water across the gill membranes. Thus, any increase in oxygen demand causes an obligatory stimulation of the Na⁺ pump. The evolution of endothermy (that is, of non-shivering thermogenesis) from behavioral thermoregulation of fish can be envisioned as a bypassing of the behavoiral response of fish and a direct stimulation of the Na⁺ pump to produce heat. The attraction of this argument is the ubiquity of Na⁺ transport across membranes.It is also argued that thyroxine was selected for as one type of control. Thyroxine could be selected as a control mechanism because its main function in fish is ion regulation. In addition, thyroxine effects the general level of spontaneous activity, increases chill resistance, alters the ability to sense salinity, and also alters the behavioral response of fish to changes in temperature. The argument is further supported by recent observations which indicate that a major fraction of the thyroxine induced elevated metabolic rate in mammals is due to stimulation of the Na⁺ pump. The above suppositions may also explain why the internal temperature sense of mammals is so sensitive to Na⁺, and may in fact suggest one possible feedback signal in homeotherms.     

The role of antioxidant defense in freezing tolerance of resurrection plant Haberlea rhodopensis

Haberlea rhodopensis Friv. is unique with its ability to survive two extreme environmental stresses—desiccation to air-dry state and subzero temperatures. In contrast to desiccation tolerance, the mechanisms of freezing tolerance of resurrection plants are scarcely investigated. In the present study, the role of antioxidant defense in the acquisition of cold acclimation and freezing tolerance in this resurrection plant was investigated comparing the results of two sets of experiments—short term freezing stress after cold acclimation in controlled conditions and long term freezing stress as a part of seasonal temperature fluctuations in an outdoor ex situ experiment. Significant enhancement in flavonoids and anthocyanin content was observed only as a result of freezing-induced desiccation. The total amount of polyphenols increased upon cold acclimation and it was similar to the control in post freezing stress and freezing-induced desiccation. The main role of phenylethanoid glucoside, myconoside and hispidulin 8-C-(2-O-syringoyl-b-glucopyranoside) in cold acclimation and freezing tolerance was elucidated. The treatments under controlled conditions in a growth chamber showed enhancement in antioxidant enzymes activity upon cold acclimation but it declined after subsequent exposure to −10 °C. Although it varied under ex situ conditions, the activity of antioxidant enzymes was high, indicating their important role in overcoming oxidative stress under all treatments. In addition, the activity of specific isoenzymes was upregulated as compared to the control plants, which could be more useful for stress counteraction compared to changes in the total enzyme activity, due to the action of these isoforms in the specific cellular compartments.Supplementary informationThe online version contains supplementary material available at 10.1007/s12298-021-00998-0.     

Dynamic Phosphoproteome Profiling of Zebrafish Embryonic Fibroblasts during Cold Acclimation

Temperature affects almost all aspects of the fish life. To cope with low temperature, fish have evolved the ability of cold acclimation for survival. However, intracellular signaling events underlying cold acclimation in fish remain largely unknown. Here, the formation of cold acclimation in zebrafish embryonic fibroblasts (ZF4) is monitored and the phosphorylation events during the process are investigated through a large‐scale quantitative phosphoproteomic approach. In total, 11 474 phosphorylation sites are identified on 4066 proteins and quantified 5772 phosphosites on 2519 proteins. Serine, threonine, and tyrosine (Ser/Thr/Tyr) phosphorylation accounted for 85.5%, 13.3%, and 1.2% of total phosphosites, respectively. Among all phosphosites, 702 phosphosites on 510 proteins show differential regulation during cold acclimation of ZF4 cells. These phosphosites are divided into six clusters according to their dynamic changes during cold exposure. Kinase–substrate prediction reveals that mitogen‐activated protein kinase (MAPK) among the kinase groups is predominantly responsible for phosphorylation of these phosphosites. The differentially regulated phosphoproteins are functionally associated with various cellular processes such as regulation of actin cytoskeleton and MAPK signaling pathway. These data enrich the database of protein phosphorylation sites in zebrafish and provide key clues for the elucidation of intracellular signaling networks during cold acclimation of fish.     

Thermal acclimation,mitochondrial capacities and organ metabolic profiles in a reptile (<Emphasis Type="Italic">Alligator mississippiensis</Emphasis>)

Reptiles thermoregulate behaviourally, but change their preferred temperature and the optimal temperature for performance seasonally. We evaluated whether the digestive and locomotor systems of the alligator show parallel metabolic adjustments during thermal acclimation. To this end, we allowed juvenile alligators to grow under thermal conditions typical of winter and summer, providing them with seasonally appropriate basking opportunities. Although mean body temperatures of alligators in these groups differed by approximately 10°C, their growth and final anatomic status was equivalent. While hepatic mitochondria isolated from cold-acclimated alligators had higher oxidative capacities at 30°C than those from warm-acclimated alligators, the capacities did not differ at 20°C. Cold acclimation decreased maximal oxidative capacities of muscle mitochondria. For mitochondria from both organs and acclimation groups, palmitate increased oligomycin-inhibited respiration. GDP addition reduced palmitate-uncoupled rates more in liver mitochondria from warm- than cold-acclimated alligators. In muscle mitochondria, carboxyatractyloside significantly reduced palmitate-uncoupled rates. This effect was not changed by thermal acclimation. The aerobic capacity of liver, skeletal muscle and duodenum, as estimated by activities of cytochrome c oxidase (COX), increased with cold acclimation. At acclimation temperatures, the activities of COX and citrate synthase (CS) in these organs were equivalent. By measuring COX and CS in isolated mitochondria and tissue extracts, we estimated that cold acclimation did not change the mitochondrial content in liver, but increased that of muscle. The thermal compensation of growth rates and of the aerobic capacity of the locomotor and digestive systems suggests that alligators optimised metabolic processes for the seasonally altered, preferred body temperature. The precision of this compensatory response exceeds that typically shown by aquatic ectotherms whose body temperatures are at the mercy of their habitat.