Mitochondrial respiration in muscle and liver from cold-acclimated hypothyroid rats.

The effects of long-term cold exposure on muscle and liver mitochondrial oxygen consumption in hypothyroid and normal rats were examined. Thyroid ablation was performed after 8-wk acclimation to 4 degrees C. Hypothyroid and normal controls remained in the cold for an additional 8 wk. At the end of 16-wk cold exposure, all hypothyroid rats were alive and normothermic and had normal body weight. At ambient temperature (24 degrees C), thyroid ablation induced a 65% fall in muscle mitochondrial oxygen consumption, which was reversed by thyroxine but not by norepinephrine administration. After cold acclimation was reached, suppression of thyroid function reduced muscle mitochondrial respiration by 30%, but the hypothyroid values remained about threefold higher than those in hypothyroid muscle in the warm. Blockade of beta- and alpha1-adrenergic receptors in both hypothyroid and normal rats produced hypothermia in vivo and a fall in muscle, liver, and brown adipose tissue mitochondria respiration in vitro. In normal rats, cold acclimation enhanced muscle respiration by 35%, in liver 18%, and in brown adipose tissue 450% over values in the warm. The results demonstrate that thyroid hormones, in the presence of norepinephrine, are major determinants of thermogenic activity in muscle and liver of cold-acclimated rats. After thyroid ablation, cold-induced nonshivering thermogenesis replaced 3,5,3'-triiodothyronine-induced thermogenesis, and normal body temperature was maintained.     

Role of hyperthyroidism in increased thermogenesis in the cold-acclimated Syrian hamster

The oxygen consumption of cold-acclimated Syrian hamsters (measured in pentobarbital-anaesthetized animals) was 59% greater than that of warm-acclimated hamsters. Upon return of the cold-acclimated hamsters to 24 degrees C, the elevated metabolic rate declined slowly, with half-life of approximately 1 day. The increase correlated well with our previous finding of a four-fold increase in serum 3,5,3'-triiodothyronine concentration in cold-acclimated hamsters and the slow decline in concentration of this hormone during deacclimation. Daily administration of T3 (for 7 days) to warm-acclimated hamsters at 24 degrees C resulted in a similar increase in oxygen consumption that persisted 1 day after the last injection and had disappeared by 7 days after the last injection. We conclude that the high concentration of serum triiodothyronine in the cold-acclimated hamster exerts a thermogenic effect that probably contributes to thermoregulatory thermogenesis in the cold.     

The effects of magnesium on state 3 respiration of liver mitochondria from control and cold-acclimated rats and hamsters.

1. Increasing the Mg2+ concentration results in a depression of succinoxidase-linked state 3 respiration of liver mitochondria from both control and cold-acclimated rats and hamsters. 2. It appears that in the cold-acclimated hamster, liver mitochondrial respiration is more sensitive to changes in Mg2+ levels than that of the rat.     

Diaphragmatic plate electrode stimulation of the hamster diaphragm

We developed a new technique of diaphragmatic stimulation by apposing plate electrodes directly against the diaphragm (DPS) in adult Golden Syrian hamsters. The electrophysiological and the mechanical responses to DPS were compared with those with phrenic nerve stimulation. In four animals, evaluation of the electromyogram before and after curare demonstrated that plate electrode stimulation occurred via the phrenic nerve filaments. In four animals, similar transdiaphragmatic pressure was produced at maximal current with DPS and phrenic nerve stimulation. Using DPS increasing current beyond a certain level resulted in recruitment of muscles besides the diaphragm. In six animals, an external abdominal pressure of 15 cmH2O produced maximal transdiaphragmatic pressure, suggesting that the diaphragm was contracting near optimal position with this external abdominal pressure. In another four animals the twitch and pressure-frequency characteristics with the use of DPS were found to be reproducible over a 2-h period. We conclude that DPS is an effective method of diaphragmatic stimulation and should prove to be a valuable technique to study the diaphragm in long-term studies of small rodents.     

Ouabain-sensitive and -insensitive ATPase activities in the kidney and the liver of spontaneously hypertensive rats.

Ouabain-sensitive and-insensitive ATPase activities were measured in the kidneys and the kidneys and the liver of male and female rats of the Lyon hypertensive (LHS) and Lyon normotensive strain (LNS). At the age of 48 +/- 2 weeks, hypertensive rats exhibited a significantly higher blood pressure and body weight than age-matched normotensive rats. Ouabian-insensitive ATPase activity was decreased in the kidney, but not in the liver of hypertensive rats compared to normotensive rats. Ouabian-sensitive ATPase activity was non significantly decreased in the kidney of hypertensive rats.     

Measurement of intracellular pH in hamster diaphragm by absorption spectrophotometry

The regulation of intracellular pH (pHi) is important in controlling muscle contraction. In these experiments, a spectrophotometric method of determining pHi was developed, and the method was then used to study muscle pHi regulation during CO2-induced changes in extracellular pH (pHb). Studies were performed in vitro on 27 diaphragm muscle strips obtained from adult hamsters. pHi was measured from the ratio of the absorbances of the acid (lambda = 530 nm) and alkaline (lambda = 460 nm) forms of a vital dye, neutral red, using the unstained diaphragm spectrum as a reference blank. A standard neutral red calibration curve constructed from eight diaphragm muscle homogenates indicated that the absorbance ratio was highly linear, with pH over the range 6.00-8.00. In intact muscle strips gassed with 95% O2-5% CO2, pHb was 7.45 +/- 0.03 (SE) and pHi was 7.00 +/- 0.01 (SE). When the muscle was aerated with CO2 concentrations from 3 to 30%, pHb and pHi changed rapidly and reached a steady state in 10-15 min. However, when pHb ranged from 6.80-7.80, pHi changed little from the value observed when pHb was 7.40. When pHb was less than 6.80 or greater than 7.80, changes in pHi and pHb were quantitatively similar. The results suggest that, in the isolated diaphragm, overall pHi is stable and effectively buffered over a wide range of CO2-induced changes in buffer solution pH.     

Increased respiration in skeletal muscle mitochondria from cold-acclimated ducklings: uncoupling effects of free fatty acids

Intermyofibrillar mitochondria from skeletal muscle (m. gastrocnemius) and liver mitochondria were isolated from cold-acclimated (4 degrees C) or control (30 degrees C) 4-week old ducklings. The respiratory rate of isolated mitochondria, with Na-succinate as substrate, was followed polarographically at 25 degrees C in order to determine the basal respiratory rate, the rate of respiration in the presence of free fatty acids (FFA) (Na-palmitate), and the fully uncoupled rate, after addition of FCCP. The basal respiration (which in liver mitochondria was unaffected by acclimation to cold) was higher (+53%) in intermyofibrillar mitochondria from cold-acclimated ducklings than from controls, and the maximal FCCP-stimulated respiration was also increased (+98%) by acclimation to cold. FFA-stimulated respiration increased as a function of FFA concentration in both types of mitochondria. The increase in respiration due to FFA was about double in intermyofibrillar mitochondria from cold-acclimated ducklings than that of controls, but in liver mitochondria there was no increase due to cold. The membrane potential was estimated by the dye safranine in the absence or in the presence of FFA in the incubation medium. There were no significant differences in the basal membrane potential in the two groups and the addition of FFA led to the same depolarization in both groups. The significance of these alterations for acclimation to cold is discussed.     

Effect of low chloride on relaxation in hamster diaphragm muscle

With muscle fatigue the chloride (Cl-) conductance of the sarcolemmal membrane decreases. The role of lowered Cl- conductance in the prolongation of relaxation seen with fatigue was studied in isolated hamster diaphragm strips. The muscles were studied in either a Krebs solution or a low Cl- solution in which half of the NaCl was replaced by Na-gluconate. Short tetanic contractions were produced by a 160-ms train of 0.2-ms pulses at 60 Hz from which tension (T) and the time constant of relaxation were measured. Resting membrane potential (Em) was measured using KCl-filled microelectrodes with resistances of 15-20 M omega. Mild fatigue (20% fall in tension) was induced by 24-25 tetanic contractions at the rate of 2/s. There was no difference in Em or T in the two solutions, either initially or with fatigue. The time constant of relaxation was greater in low Cl- solution, both initially (22 +/- 3 vs. 18 +/- 5 ms, mean +/- SD, P less than 0.05) and with fatigue (51 +/- 18 vs. 26 +/- 7 ms, P less than 0.005). Lowering of sarcolemmal membrane Cl- conductance appears to play a role in the slowing of relaxation of hamster diaphragm muscle seen with fatigue.     

Increased compliance in diaphragm muscle of the cardiomyopathic Syrian hamster

We investigatedthe hypothesis that diaphragm compliance was abnormal incardiomyopathic Syrian hamsters (CSH), an experimental model ofmyopathy. The passive elastic properties of isolated diaphragm muscleswere analyzed at both the muscle and sarcomere levels. We used thefollowing passive exponential relationship between stress () andstrain ():  = (E_o/)(e  1), where E_o is the initialelastic modulus and  is the stiffness constant. Immunocytochemistryprocedures were used to analyze the distribution of two key elasticcomponents of muscle, extracellular collagen and intracellular titinelastic components, as well as the extracellular matrix glycoproteinlaminin. Muscle and sarcomere values of  were nearly twofold lowerin CSH (8.7 ± 1.9 and 8.3 ± 1.4, respectively) than in controlanimals (19.7 ± 1.7 and 16.8 ± 2.1, respectively)(P < 0.01 for each). Compared withcontrols, E_o was higher in CSH.Sarcomere slack length was significantly longer in CSH than in controlanimals (2.1 ± 0.1 vs. 1.9 ± 0.1 µm,P < 0.05). The surface area ofcollagen I was significantly larger in CSH (17.4 ± 1.8%) than incontrol animals (12.4 ± 0.7%, P < 0.05). There was no change in the distribution of titin or lamininlabelings between the groups. These results demonstrate increaseddiaphragm compliance in cardiomyopathic hamsters. The increase in CSHdiaphragm compliance was observed despite an increase in the surfacearea of collagen and was not associated with an abnormal distributionof titin or laminin.

     

冷暴露对中缅树鼩肝脏、膈肌和心肌线粒体呼吸的影响

在4℃急性冷暴露(1 h,4 h,8 h,24 h)和持续冷暴露 (7 d,14 d,28 d)条件下,测定中缅树鼩膈肌、心肌和肝脏的线粒体状态Ⅲ呼吸、状态Ⅳ呼吸、呼吸控制率(RCR)、线粒体蛋白含量以及肝脏线粒体P/O值的变化.结果表明:肝脏线粒体状态Ⅲ、状态Ⅳ呼吸随着低温处理时间的延长,呼吸速率均极显著增加,在28 d后分别增加了132.9%、124.4%(P<0.01),RCR与对照比较,在8 h和7 d组分别显著增加了35.8%和48.4%(P<0.05),线粒体蛋白含量也极显著增加,在28 d后增加了104.7%(P<0.01),P/O值在整个低温处理过程中呈下降趋势,在28 d后降低了40.2%,达到极显著水平(P<0.01);膈肌线粒体状态Ⅲ呼吸在整个低温处理期间没有显著变化,状态Ⅳ呼吸在28 d达到极显著增加(P<0.01,64.9%),RCR在28 d后显著降低(P<0.05, 42.1%),线粒体蛋白只有4 h组有极显著增加(P<0.01,45.2%);心肌的状态Ⅲ呼吸在8 h组有着极显著的增加(P<0.01, 54.7%),状态Ⅳ呼吸随着低温处理时间的增加而显著增加,28 d后增加了94.7%(P<0.01),RCR在28 d后降低37.8%(P<0.01),线粒体蛋白表现出先下降再上升的趋势,8 h组下降37.8%(P<0.01),28 d增加25.2%(P<0.05).说明中缅树鼩在冷胁迫的条件下肝脏线粒体呼吸能力显著增强,主要表现为状态Ⅳ呼吸即质子漏产热的显著增强,膈肌和心肌的线粒体呼吸也具有一定的适应性变化,补偿了冷胁迫条件下中缅树鼩增加的能量需求,是中缅树鼩在冷胁迫中重要的适应对策.     

Changes in mixed-function oxidase system in the perfused liver of the cold-acclimated rat

Changes in the hepatic cytochrome P-450-dependent drug-metabolizing system were studied in perfused livers obtained from cold-acclimated male Wistar rats after 30 days of cold exposure (4C) when using hexobarbital as a substrate. In fasted animals the cold-acclimated rats showed higher levels of hexobarbital metabolic rates compared to control rats, but there was no significant difference in fed animals. The maximum rates of hexobarbital metabolism produced by xylitol perfusion were also significantly higher in the perfused liver of cold-acclimated rats. It was concluded that the function of the cytochrome P-450 system for hexobarbital in cold-acclimated rats changed due to both an increase in the activity of the cytochrome P-450 system and to changes in regulation of the cytochrome P-450 system by the supply of reducing equivalents.