Acute toxicity and radioprotective effectiveness of gammaphos on intramuscular administration to mice

In experiments on mice it was shown that acute toxicity of gammaphos (WR-2721) was 790 mg/kg and 862 mg-kg after intraperitoneal and intramuscular administration, respectively. Gammaphos in the dose of 100 mg/kg, injected intramuscularly, increased the radioresistance of mice in nearly the same way as cystamine, in the dose of 150 mg/kg, did. The increase in the dose of gammaphos up to 200 or 300 mg/kg, injected intramuscularly, enhanced the radioprotective effect. No change was observed in the radioprotective efficiency of gammaphos after intramuscular injection as compared to intraperitoneal administration of the protective agent in the same dose.     

Chemical protection against genetic effect of radiation in male mice

A study was made of the protective effect of some radioprotective agents against dominant lethal mutations (DLM) in postspermatogonial stages and reciprocal translocations (RT) in spermatogonia induced by gamma-radiation. Among the radioprotective agents used, cystaphos, a combination of cystamine and 5-MOT and a mixture of 6 components proved to be most effective against DLM, and cystaphos, gammaphos and cystamine combined with 5-MOT proved effective against RT. The degree of radioprotective efficacy was relatively low. The efficacy of cystamine in protecting against RT was higher with exposure of gonocytes of 18.5-day embryos than spermatogonia of pubertal animals. The degree of the radioprotective effect varied depending on the stage of spermatogenesis, and, in all cases, it was lower than that observed in studies of protection against lethal effects of ionizing radiation.     

Antimutagenic activity of dextran gammaphos derivatives]

In experiments with V-79 Chinese hamster cell culture the influence of dextran gammaphos derivatives on the mutagenic effects of gamma-radiation was studied by the number of cells with micronuclei and fragmented nuclei. Products of interaction between gammaphos and dialdehyde dextran were shown to have a higher antimutagenic activity than gammaphos.     

Delta opioid peptide [D-Ala2,D-Leu5]enkephalin promotes cell survival

By studying the hibernation in ground squirrels, a protein factor termed hibernation induction trigger (HIT) was found to induce hibernation in summer-active ground squirrels. Further purification of HIT yielded an 88-kD peptide that is enriched in winter hibernator. Partial sequence of the 88-kD protein indicates that it may be related to the inhibitor of metalloproteinase. Delta opioid [D-Ala(2),D-Leu(5)]enkephalin (DADLE) also induced hibernation. HIT and DADLE were found to prolong survival of peripheral organs preserved en bloc or as a single preparation. These organs include the lung, the heart, liver and kidney. DADLE also promotes survival of neurons in the central nervous system. Methamphetamine (METH) is known to cause destruction of dopaminergic (DA) terminals in the brain. DADLE blocked and reversed the DA terminal damage induced by METH. DADLE acted against this effect of METH at least in part by attenuating the mRNA expressions of a tumor necrosis factor p53 and an immediate early gene c-fos. DADLE also blocked the neuronal damage induced by ischemia-reperfusion following a transient middle cerebral artery occlusion. In PC12 cells, DADLE blocked the cell death caused by serum deprivation in a naltrexone-sensitive manner. Thus, DADLE, and by extension the endogenous delta opioid peptides and delta opioid receptors, may play an important role in organ and neuronal survival. Here, critical developments concerning these fascinating cell protective properties of DADLE are reviewed.     

Enhanced antioxidant defense due to extracellular catalase activity in Syrian hamster during arousal from hibernation

Mammalian hibernators are considered a natural model for resistance to ischemia-reperfusion injuries, and protective mechanisms against oxidative stress evoked by repeated hibernation-arousal cycles in these animals are increasingly the focus of experimental investigation. Here we show that extracellular catalase activity provides protection against oxidative stress during arousal from hibernation in Syrian hamster. To examine the serum antioxidant defense system, we first assessed the hibernation-arousal state-dependent change in serum attenuation of cytotoxicity induced by hydrogen peroxide. Serum obtained from hamsters during arousal from hibernation at a rectal temperature of 32 degrees C, concomitant with the period of increased oxidative stress, attenuated the cytotoxicity four-fold more effectively than serum from cenothermic control hamsters. Serum catalase activity significantly increased during arousal, whereas glutathione peroxidase activity decreased by 50%, compared with cenothermic controls. The cytoprotective effect of purified catalase at the concentration found in serum was also confirmed in a hydrogen peroxide-induced cytotoxicity model. Moreover, inhibition of catalase by aminotriazole led to an 80% loss of serum hydrogen peroxide scavenging activity. These results suggest that extracellular catalase is effective for protecting hibernators from oxidative stress evoked by arousal from hibernation.     

Radioprotective effect of drug combinations on the small intestine

The study of the radioprotective effect of complexes containing small amounts of cystamine or gammaphos mixed with mexamine, gutimine and ethyrone on mouse small intestine showed high efficiency of the drugs as determined by cellularity and DNA-synthesizing activity of intestinal epithelium. This is connected with the influence of the complex mexamine + gutimine + ethyrone on the pharmacodynamics and pharmacokinetics of cystamine and gammaphos.     

Significance of ED50 and therapeutic indexes of various radiation-protective agent groups]

Radioprotective agents are divided in 3 groups: (1) cystamine, AET, cystaphos, gammaphos, and thiogammaphos with ED50 (the dose that gives a half of the maximal protective effect) of 10(3)-10(1.6) mumol/kg and therapeutic index K = LD50/ED50 = 10(0)-10(1.6); (2) 5-methoxytryptamine, phenylephrine, serotonin, and norepinephrine with ED50 = 10(1)-10(0) mumol/kg and K = 10(1.8)-10(2,6); (3) clonidine and isoprenaline with ED50 = 10(-0.5)-10(-0.8) mumol/kg and K = 10(3)-10(4). Possible causes of these differences and advantages of low ED50 and high K are discussed.     

Polar bears (Ursus maritimus), the most evolutionary advanced hibernators, avoid significant bone loss during hibernation

Some hibernating animals are known to reduce muscle and bone loss associated with mechanical unloading during prolonged immobilisation,compared to humans. However, here we show that wild pregnant polar bears (Ursus maritimus) are the first known animals to avoid significant bone loss altogether, despite six months of continuous hibernation. Using serum biochemical markers of bone turnover, we showed that concentrations for bone resorption are not significantly increased as a consequence of hibernation in wild polar bears. This is in sharp contrast to previous studies on other hibernating species, where for example, black bears (Ursus americanus), show a 3-4 fold increase in serum bone resorption concentrations posthibernation,and must compensate for this loss through rapid bone recovery on remobilisation, to avoid the risk of fracture. In further contrast to black bears, serum concentrations of bone formation markers were highly significantly increased in pregnant female polar bears compared to non-pregnant,thus non-hibernating females both prior to and after hibernation. However, bone formation concentrations in new mothers were significantly reduced compared to pre-hibernation concentrations. The de-coupling of bone turnover in favour of bone formation prior to hibernation, suggests that wild polar bears may posses a unique physiological mechanism for building bone in protective preparation against expected osteopenia associated with disuse,starvation, and hormonal drives to mobilise calcium for reproduction, during hibernation. Understanding this physiological mechanism could have profound implications for a natural solution for the prevention of osteoporosis in animals subjected to captivity with inadequate space for exercise,humans subjected to prolonged bed rest while recovering from illness, or astronauts exposed to antigravity during spaceflight.? 2008 Elsevier Inc. All rights reserved.     

Effect of brain stem lesions on hibernation of the hamster (Cricetus cricetus L.)]

Small lesions in the brain stem (including the hypothalamus) of the European hamster were effective with respect to food intake, hibernatory disposition and thermogenic power (oxygen consumption) as well. Hyperphagia was accompanied by depression of hibernation mostly. Moreover, hibernation was hindered by impairment of the thermogenic capacity. Entrance into hibernation depended on the integrity of the middle and caudal hypothalamic areas and the rostral portions of the pons and midbrain. Hyperphagia resulted from destruction of the middle (ventromedial) hypothalamic and caudal hypothalamic areas, including transition structures to the pons. A depression of thermogenesis against cold was observed after destruction of supramammillary and neighbouring mesencephalic areas. Supplementary results: An annual metabolic rhythm characterized by a minimum in december has been established once more. Urethane anesthesia did not abolish cold thermogenesis, despite the development of a slight hypothermia. Poikilothermia resulting from brain stem damage disappeared during a three-day period. Furthermore, diencephalic lesions did not suppress arousal from hibernation significantly.     

Increased cardiac alpha-myosin heavy chain in left atria and decreased myocardial insulin-like growth factor (Igf-I) expression accompany low heart rate in hibernating grizzly bears

Grizzly bears (Ursus arctos horribilis) tolerate extended periods of extremely low heart rate during hibernation without developing congestive heart failure or cardiac chamber dilation. Left ventricular atrophy and decreased left ventricular compliance have been reported in this species during hibernation. We evaluated the myocardial response to significantly reduced heart rate during hibernation by measuring relative myosin heavy-chain (MyHC) isoform expression and expression of a set of genes important to muscle plasticity and mass regulation in the left atria and left ventricles of active and hibernating bears. We supplemented these data with measurements of systolic and diastolic function via echocardiography in unanesthetized grizzly bears. Atrial strain imaging revealed decreased atrial contractility, decreased expansion/reservoir function (increased atrial stiffness), and decreased passive-filling function (increased ventricular stiffness) in hibernating bears. Relative MyHC-α protein expression increased significantly in the atrium during hibernation. The left ventricle expressed 100% MyHC-β protein in both groups. Insulin-like growth factor (IGF-I) mRNA expression was reduced by ～50% in both chambers during hibernation, consistent with the ventricular atrophy observed in these bears. Interestingly, mRNA expression of the atrophy-related ubiquitin ligases Muscle Atrophy F-box (MAFBx) and Muscle Ring Finger 1 did not increase, nor did expression of myostatin or hypoxia-inducible factor 1α (HIF-1α). We report atrium-specific decreases of 40% and 50%, respectively, in MAFBx and creatine kinase mRNA expression during hibernation. Decreased creatine kinase expression is consistent with lowered energy requirements and could relate to reduced atrial emptying function during hibernation. Taken together with our hemodynamic assessment, these data suggest a potential downregulation of atrial chamber function during hibernation to prevent fatigue and dilation due to excessive work against an optimally filled ventricle, a response unpredicted by the Frank-Starling mechanism.     

The "hibernation induction trigger": specificity and validity of bioassay using the 13-lined ground squirrel

Even though the existence of the blood-borne "hibernation induction trigger" has been reported in the 13-lined ground squirrel, transfusion of plasma from hibernating rodents with other hibernating species as the recipients failed to induce the occurrence of summer hibernation. In order to verify whether the response to the "trigger" substance is species specific, the present study was carried out to compare the effect of plasma from hibernating Richardson's ground squirrels on the incidence of summer hibernation in both juvenile Richardson's and adult 13-lined ground squirrels. In two series of experiments, 13-lined ground squirrels entered hibernation quite readily independent of the treatment. The rate of occurrence of hibernation ranged from 78% after sham injection to 86% after warm saline, fresh summer active plasma, and fresh hibernating plasma, respectively. There were no differences in the number of hibernation bouts and the number of days in hibernation after each treatment. In contrast, none of the juvenile Richardson's ground squirrels entered hibernation after any of the treatments up to the end of the 8-week observation period. These results not only argue against the existence of blood-borne "trigger" substance, at least in the Richardson's ground squirrel, but also caution against the use of the 13-lined ground squirrel as a standard test animal for the bioassay of the "trigger" substance.     

Novel findings on ultrastructural protection of skeletal muscle fibers during hibernation of Daurian ground squirrels: Mitochondria,nuclei, cytoskeleton,glycogen

We examined ultrastructure protective phenomena and mechanisms of slow and fast muscles in hibernating Daurian ground squirrels (Spermophilus dauricus). Some degenerative changes such as slightly decreased sarcomere length and vacuolization occurred in hibernation, but periaxonal capsular borders in intrafusal fibers remained distinct and the arrangement of extrafusal fibers and Z-lines unscathed. In soleus samples, the number of glycogenosomes more than tripled during hibernation. The expression of phosphorylated glycogen synthase remained unaltered while that of glycogen phosphorylase decreased during hibernation. The number of extensor digitorum longus glycogenosomes decreased and the expression of phosphorylated glycogen synthase decreased, while glycogen phosphorylase expression remained unaltered. The nuclei number remained unchanged. Kinesin and desmin, preventors of nuclear loss and damage, were maintained or just slightly reduced in hibernation. The single-fiber mitochondrial concentration and sub-sarcolemmal mitochondrial number increased in both muscle types. The expression of vimentin, which anchors mitochondria and maintains Z-line integrity, was increased during and after hibernation. Also, dynamin-related protein 1, mitochondrial fission factor, and adenosine triphosphate synthase were elevated in both muscle types. These findings confirm a remarkable ultrastructure preservation and show an unexpected increase in mitochondrial capacity in hibernating squirrels.     

Functional capacity of the kidneys in whole-body irradiated rats protected with gammaphos and merkamine

In experiments with Wistar rats it was shown that gammaphos promotes the recovery of the kidneys' filtration function at early times after 7.4 Gy gamma-irradiation and inhibits it after 210 days. Merkamine induces haemodynamic and functional disorders in the exposed rat kidneys within the first 30 days after irradiation.     

Myocardial hibernation: a delicate balance

The pathophysiology of myocardial hibernation is characterized as a situation of reduced regional contractile function distal to a coronary artery stenosis that recovers after removal of the coronary stenosis. A subacute "downregulation" of contractile function in response to reduced regional myocardial blood flow exists, which normalizes regional energy and substrate metabolism but does not persist for more than 12-24 h. Chronic hibernation develops in response to one or more episodes of myocardial ischemia-reperfusion, possibly progressing from repetitive stunning with normal blood flow to hibernation with reduced blood flow. An upregulation of a protective gene program is seen in hibernating myocardium, putting it into the context of preconditioning. The morphology of hibernating myocardium is characterized by both adaptive and degenerative features.     

光照黑暗循环条件下达乌尔黄鼠的冬眠模式和能量消耗

为研究冬眠季节的光照条件对贮脂类冬眠动物入眠的影响,在达乌尔黄鼠腹腔埋植体温记录元件iButton,在体重高峰后的下降阶段置于5℃和12L:12D的光照条件下,观察测定其冬眠模式和能量消耗。达乌尔黄鼠冬眠模式出现深冬眠型、少冬眠型和不冬眠型,蛰眠阵包括深冬眠阵、短冬眠阵和日眠阵。不同冬眠阵中最低体温、冬眠阵的持续时间、阵间产热的持续时间、冷却速率和复温速率差异显著;阵间产热的最高体温基本相同。平均每日能量消耗在日眠阵中最高、短冬眠阵中居中、深冬眠阵中最低。入眠时间多集中于黑暗时相,觉醒时间多集中于光照时相。本实验结果提示,在冬眠季节施加光照黑暗循环条件可减少达乌尔黄鼠冬眠的时间,升高阵间最低体温,缩短冬眠阵与阵间产热的持续时间,降低复温速率;增加冬眠期间能量消耗。入眠与觉醒受光照条件影响,具有明显的光暗节律。     

Hibernating myocardium: Its pathophysiology and clinical role

Myocardial hibernation, as first defined by Rahimtoola, is a state of chronic contractile dysfunction in patients with coronary artery disease which is fully reversible upon reperfusion. Clinical conditions consistent with the existence of myocardial hibernation include unstable and stable angina, myocardial infarction heart failure, and anomalous origin of coronary arteries. The mechanisms of hibernation are not known. Morphological alterations have been described in the hibernating area of patients, but these information are strongly affected by the diagnostic criteria utilized to screen patients. It has been postulated that hibernation is an adaptive phenomenon occurring during ischemia. In this context, downregulation of contraction is not regarded as a consequence of energetic deficit, but as a regulatory event aimed at reducing energy expenditure, thereby maintaining integrity and viability. Thus, hibernation might bear a relationship to the phenomenon of low-flow perfusion-contraction matching, or repetitive stunning or preconditioning. Clear-cut evidence for the mechanism of hibernation in the clinical setting seems likely to remain elusive, because of the nature of the studies needed to document it. Current experimental evidence supports the view that hibernation, stunning, preconditioning, or their coexistence can be responsible for regional myocardial contractile dysfunction which is reversible upon reperfusion. These are all adaptive and protective phenomena independent of their terminology and strict definitions and do not always apply to the extremely complex situation of myocardial ischemia in man.     

脑室注射6—羟多巴胺对黄鼠科眠入眠的影响

The effect of forced depletion of brain norepinephrine (NE) on the onset of hibernation was observed in the ground squirrel (Citellus dauricus) by intraventricular injection of 6-hydroxydopamine (6-OHDA). The results showed: (1) Intraventricular injection of 100-200 micrograms 6-OHDA, which depleted 50-60% NE, markedly facilitated the onset of hibernation, i.e. the average induction period for hibernation in the treated animals was significantly shorter than that of the natural hibernating animals. (2) The average total torpor time in the treated animals was longer than that of natural hibernating animals. (3) All hibernating animals treated with 6-OHDA were able to wake up from deep hibernation spontaneously and undergo normal hibernation bouts. The results indicate the decrease of NE system activity in brain is one important factor in triggering the onset of hibernation.     

Periodic arousal from hibernation is necessary for initiation of immune responses in ground squirrels

Golden-mantled ground squirrels (Spermophilus lateralis) undergo seasonal hibernation during which core body temperature (T(b)) values are maintained 1-2 degrees C above ambient temperature. Hibernation is not continuous. Squirrels arouse at approximately 7-day intervals, during which T(b) increases to 37 degrees C for approximately 16 h; thereafter, they return to hibernation and sustain low T(b)s until the next arousal. Over the course of the hibernation season, arousals consume 60-80% of a squirrel's winter energy budget, but their functional significance is unknown and disputed. Host-defense mechanisms appear to be downregulated during the hibernation season and preclude normal immune responses. These experiments assessed immune function during hibernation and subsequent periodic arousals. The acute-phase response to bacterial lipopolysaccharide (LPS) was arrested during hibernation and fully restored on arousal to normothermia. LPS injection (ip) resulted in a 1-1.5 degrees C fever in normothermic animals that was sustained for > 8 h. LPS was without effect in hibernating squirrels, neither inducing fever nor provoking arousal, but a fever did develop several days later, when squirrels next aroused from hibernation; the duration of this arousal was increased sixfold above baseline values. Intracerebroventricular infusions of prostaglandin E(2) provoked arousal from hibernation and induced fever, suggesting that neural signaling pathways that mediate febrile responses are functional during hibernation. Periodic arousals may activate a dormant immune system, which can then combat pathogens that may have been introduced immediately before or during hibernation.     

Impaired Skeletal Muscle Regeneration in the Absence of Fibrosis during Hibernation in 13-Lined Ground Squirrels

Skeletal muscle atrophy can occur as a consequence of immobilization and/or starvation in the majority of vertebrates studied. In contrast, hibernating mammals are protected against the loss of muscle mass despite long periods of inactivity and lack of food intake. Resident muscle-specific stem cells (satellite cells) are known to be activated by muscle injury and their activation contributes to the regeneration of muscle, but whether satellite cells play a role in hibernation is unknown. In the hibernating 13-lined ground squirrel we show that muscles ablated of satellite cells were still protected against atrophy, demonstrating that satellite cells are not involved in the maintenance of skeletal muscle during hibernation. Additionally, hibernating skeletal muscle showed extremely slow regeneration in response to injury, due to repression of satellite cell activation and myoblast differentiation caused by a fine-tuned interplay of p21, myostatin, MAPK, and Wnt signaling pathways. Interestingly, despite long periods of inflammation and lack of efficient regeneration, injured skeletal muscle from hibernating animals did not develop fibrosis and was capable of complete recovery when animals emerged naturally from hibernation. We propose that hibernating squirrels represent a new model system that permits evaluation of impaired skeletal muscle remodeling in the absence of formation of tissue fibrosis.     

Differential regulation of glomerular and interstitial endothelial nitric oxide synthase expression in the kidney of hibernating ground squirrel.

Hibernating animals transiently reduce renal function during their hypothermic periods (torpor), while completely restoring it during their periodical rewarming to euthermia (arousal). Moreover, structural integrity of the kidney is preserved throughout the hibernation. Nitric oxide (NO) generated by endothelial nitric oxide synthase (eNOS) is a crucial vasodilatory mediator and a protective factor in the kidney. We investigated renal NOS expression in hibernating European ground squirrels after 1 day and 7 days of torpor (torpor short, TS, and torpor long, TL, respectively), at 1.5 and at 10 h of rewarming (arousal short, AS, and arousal long, AL, respectively), and in continuously euthermic animals after hibernation (EU). For that purpose, we performed NOS activity assay, immunohistochemistry and real-time PCR analysis. Immunohistochemistry revealed a decreased glomerular eNOS expression in hibernating animals (TS, TL, AS, and AL) compared to non-hibernating animals (EU, p < 0.05), whereas no difference was found in the expression of interstitial eNOS. Expression of iNOS and nNOS did not differ between all groups. The reduced glomerular eNOS was associated with a significantly lower eNOS mRNA levels and NOS activity of whole kidney during torpor and arousal (TS, TL, AS, and AL) compared to EU. In all methods used, torpid and aroused squirrels did not differ. These results demonstrate differential regulation of eNOS in glomeruli and interstitium of hibernating animals, which is unaffected during arousal. The differential regulation of eNOS may serve to reduce ultrafiltration without jeopardizing tubular structures during hibernation.