Threshold for repetitive activity for a slow stimulus ramp: a memory effect and its dependence on fluctuations

We have obtained new insights into the behavior of a class of excitable systems when a stimulus, or parameter, is slowly tuned through a threshold value. Such systems do not accommodate no matter how slowly a stimulus ramp is applied, and the stimulus value at onset of repetitive activity shows a curious, nonmonotonic dependence on ramp speed. (Jakobsson, E. and R. Guttman. Biophys. J. 1980. 31:293-298.) demonstrated this for squid axon and for the Hodgkin-Huxley (HH) model. Furthermore, they showed theoretically that for moderately slow ramps the threshold increases as the ramp speed decreases, but for much slower ramp speeds threshold decreases as the ramp speed decreases. This latter feature was found surprising and it was suggested that the HH model, and squid axon in low calcium, exhibits reverse accommodation. We have found that reverse accommodation reflects the influence of persistent random fluctuations, and is a feature of all such excitable systems. We have derived an analytic condition which yields an approximation for threshold in the case of a slow ramp when the effect of fluctuations are negligible. This condition predicts, and numerical calculations confirm, that the onset of oscillations occurs beyond the critical stimulus value which is predicted by treating the stimulus intensity as a static parameter, i.e., the dynamic aspect of a ramp leads to a delay in the onset. The condition further demonstrates a memory effect, i.e., firing threshold is dependent on the initial state of the system. For very slow ramps then, fluctuations diminish both the delay and memory effects.(ABSTRACT TRUNCATED AT 250 WORDS)     

Dysregulation of hepatic iron with aging: implications for heat stress-induced oxidative liver injury

Environmental heat stress is associated with an age-related increase in hepatic oxidative damage and an exaggerated state of oxidative stress. The purpose of this investigation was to evaluate the regulation of hepatic iron after heat stress. A secondary aim was to determine a potential role for iron in heat stress-induced liver injury. Hyperthermia-induced alterations in hepatic iron were evaluated in young (6 mo) and old (24 mo) Fischer 344 rats by exposing them to a two-heat stress protocol. Livers were harvested at several time points after the second heating and assayed for labile and nonheme iron. In the control condition, there was no difference in labile iron between age groups. Both labile iron and storage iron were not altered by hyperthermia in young rats, but both were increased immediately after heating in old rats. To evaluate a role for iron in liver injury, hepatic iron content was manipulated in young and old rats, and then both groups were exposed to heat stress. Iron administration to young rats significantly increased hepatic iron content and ferritin but did not affect markers of lipid peroxidation under control conditions or after heat stress. In old rats, iron chelation with deferoxamine prevented the increase in nonheme iron, labile iron, ferritin, and lipid peroxidation after heat stress. These results suggest that iron may play a role in hepatic injury after hyperthermia. Thus, dysregulation of iron may contribute to the gradual decline in cellular and physiological function that occurs with aging.     

Density dependent stimulation of thymidine incorporation in BHK21 cells by active material released from the same cells

Cultures of BHK2l cells continuously release into serum-free medium, active materials which stimulate the incorporation of thymidine in non-confluent, but not in confluent cultures of homotypic cells. The activity is not removed by centrifugation at 30,000 g for four hours, but is non-dialysable and retained by membranes with a nominal limitation for MW 25,000. Activity is stable at 4° for several weeks, but destroyed in 30 minutes at 56°. BHK2l cells also release active material capable of enhancing the effect of added serum. This is also heat labile and its action is density-dependent.     

Surface characters and extracellular toxins involved in the pathogenesis of Aeromonas hydrophila

A small number of serotypically distinct strains of A. hydrophila obtained from diseased freshwater fish were examined for their pathogenic properties comprising of cell surface characteristics and extracellular toxins. Test strains exhibited homogeneity in their cell surface characteristics despite being serologically heterogeneous. Studies on extracellular biological activities revealed qualitative and quantitative differences in production of toxins, probably explaining their antigenic diversity. Three distinct proteases, namely heat stable metallo protease, heat labile serine protease and heat labile metallo protease were identified from the strains.     

Dynamic Properties of Giant Muscle Fibers of the Barnacle

Giant muscle fibers of the barnacle give graded, relativelyslow contractions, A plateau level, termed the unit response,occurs with stimuli of 3 msec, but pulses longer than 10–15msec give much greater tension or shortening. Repetitive stimulationwith pulses of 3 msec leads to a tetanus. The magnitude of activestate was determined, and found to be also graded in natureand slow to develop, though early in onset. The full developmentof active state requires 80–120 msec. A high level ofeffective series-elasticity was associated with the sarcomeresthemselves.     

Investigation of alkaline phosphatase activity in the serum of pregnant rats

An investigation was undertaken to determine if the placental alkaline phosphatase of the rat enters the maternal circulation and to study some of its characteristics. Unlike human, rat placental alkaline phosphatase was found to be heat labile and the alkaline phosphatase activity in the serum of both pregnant and non-pregnant rats was also found to be heat labile. Also unlike the human, the alkaline phosphatase activity in rat serum does not increase as pregnancy progresses to term. In an endeavour to establish if the rat placental enzyme is present in the serum of the pregnant rat, the characteristics of the enzyme in both placental extracts and serum of non-pregnant and 1-, 2- and 3-week pregnant rats were studied using the techniques of heat stability at 56°, gel filtration through Sephadex columns, disc gel electrophoresis, and L-phenylalanine inhibition. The presence of rat placental alkaline phosphatase in maternal serum could not be positively demonstrated by any of these procedures, suggesting that rat placental alkaline phosphatase does not enter the maternal serum.     

Rat platelets contain growth factor(s) distinct from PDGF which stimulate DNA synthesis in primary adult rat hepatocyte cultures

Adult rat hepatocytes in primary cultures are stimulated to synthesize DNA in response to rat serum, whereas rat plasma is considerably less active. Biological activity is present in rat platelets and is secreted during aggregation in response to thrombin. The material secreted by rat platelets is heat labile and is sensitive to digestion with trypsin, suggesting that it is a protein. When assayed on 3T3 cells this material also stimulates DNA synthesis; however, the trypsin-sensitive activity is heat stable (100 degrees C, 10 min). These results indicate that rat platelets contain hepatotrophic activities which by virtue of their heat sensitivity are distinct from heat-stable platelet-derived growth factor (PDGF)-like mitogenic activities required by 3T3 cells for growth. It is possible that hepatotrophic platelet factors might be involved in mediating liver regeneration in the rat after partial hepatectomy.     

On the interaction of 3,4,5,6-tetrahydrouridine with human liver cytidine deaminase.

In contrast to the rapid inhibition of bacterial cytidine deaminase by 3,4,5,6-tetrahydrouridine, the onset of inhibition of the enzyme from human liver was found to be relatively slow. Inhibition was found to be reversible, and the corrected rate constants for binding (kon = 2.4 x 10(4) M-1 sec-1) and release (koff = 5.6 x 10(-4) sec-1) were in reasonable agreement with a Ki value (2.9 x 10(-8) M) measured separately under steady-state conditions, which was several orders of magnitude lower than estimates previously reported in the literature. Rates of binding and release of this potential transition state analogue were not appreciably affected by the substitution of deuterium oxide for solvent water. The slow onset of inhibition, which was also observed for cytidine deaminase from HeLa cells, suggests that structural reorganization precedes the formation of a stable enzyme-inhibitor complex. 6-Azacytidine, which favors a "high-anti" configuration at the glycosidic bond, was found to be active as a substrate for cytidine deaminase, with a turnover number exceeding that of cytidine. 2,2'-Anhydro-1-beta-D-arabinofuranosylcytosine, which is restricted to the "syn" configuration, was found to be without activity as a substrate or an inhibitor.     

Detection, characterisation and purification of a murine liver factor capable of desensitising towards the lethal activity of tumour necrosis factor

Tumour necrosis factor (TNF) is a major mediator in septic shock and several inflammatory diseases such as hepatitis. Galactosamine (GalN) sensitises experimental animals for TNF and the combination TNF/GalN leads to a lethal inflammatory hepatitis. We describe that a single injection of lipopolysaccharide (LPS), interleukin-1 (IL-1) or TNF can desensitise against the lethality induced by TNF/GalN, but also against changes in metabolic parameters such as hypothermia and transaminase release, in a dose responsive way. We also describe the desensitising capacity of a component present in Mouse Liver Extract (MLE). The MLE desensitises mice against the effects of TNF/GalN in a dose responsive way. The activity of the MLE is heat labile and does not involve LPS, TNF, IL-1 or TNF soluble receptors. We describe partial and complete purification of the factor. Partially pure material protects mice against all changes induced by TNF/GalN. The protection is dose dependent and heat labile and also possible in endotoxin-hyporesponsive C3H/HeJ mice. The pure material protects against lethality, hypothermia and AST release and it appears as a heat labile protein of relative molecular weight of 70 kDa probably with a break down product of 35 kDa.     

桂北不同林龄桉树人工林土壤碳库管理指数和碳组分的变化特征

采用时空互代法,以广西北部低山丘陵地区不同林龄(1、2、3、4、5和8 a)桉树人工林为研究对象,探讨林龄对桉树人工林地土壤碳库管理指数的影响及其规律。结果表明:(1)随着林龄的增加,土壤有机碳总体表现为增加的趋势,1～8 a桉树土壤有机碳范围在5.79～15.57 g·kg^-1之间,随着土层的加深而降低; 0～40 cm土层土壤有机碳平均含量表现为8 a>5 a>3 a>4 a>2 a>1 a。(2)土壤非活性有机碳、碳储量随林龄和土层的变化规律与土壤有机碳基本一致。土壤活性有机碳含量大小依次表现为8 a>5 a>4 a>3 a>2a> 1 a,占土壤有机碳的比例随林龄变化无明显规律,8 a和其他林龄间均具有显著差异。(3)碳库管理指数随林龄增加整体呈上升趋势,8 a桉树人工林土壤碳组分含量及碳库管理指数均高于10 a对照马尾松林。碳库管理指数与土壤有机碳、非活性有机碳、活性有机碳、碳储量、碳库活度、全氮、容重呈极显著或显著的相关性,不同林龄和土层间碳库管理指数有差异性。适当延长桉树人工林的轮伐周期,减...     

Temperature Changes in Brown Adipocytes Detected with a Bimaterial Microcantilever

Mammalian cells must produce heat to maintain body temperature and support other biological activities. Methods to measure a cell’s thermogenic ability by inserting a thermometer into the cell or measuring the rate of oxygen consumption in a closed vessel can disturb its natural state. Here, we developed a noninvasive system for measuring a cell’s heat production with a bimaterial microcantilever. This method is suitable for investigating the heat-generating properties of cells in their native state, because changes in cell temperature can be measured from the bending of the microcantilever, without damaging the cell and restricting its supply of dissolved oxygen. Thus, we were able to measure increases in cell temperature of <1 K in a small number of murine brown adipocytes (n = 4–7 cells) stimulated with norepinephrine, and observed a slow increase in temperature over several hours. This long-term heat production suggests that, in addition to converting fatty acids into heat energy, brown adipocytes may also adjust protein expression to raise their own temperature, to generate more heat. We expect this bimaterial microcantilever system to prove useful for determining a cell’s state by measuring thermal characteristics.     

Temperature Changes in Brown Adipocytes Detected with a Bimaterial Microcantilever

Mammalian cells must produce heat to maintain body temperature and support other biological activities. Methods to measure a cell’s thermogenic ability by inserting a thermometer into the cell or measuring the rate of oxygen consumption in a closed vessel can disturb its natural state. Here, we developed a noninvasive system for measuring a cell’s heat production with a bimaterial microcantilever. This method is suitable for investigating the heat-generating properties of cells in their native state, because changes in cell temperature can be measured from the bending of the microcantilever, without damaging the cell and restricting its supply of dissolved oxygen. Thus, we were able to measure increases in cell temperature of <1 K in a small number of murine brown adipocytes (n = 4–7 cells) stimulated with norepinephrine, and observed a slow increase in temperature over several hours. This long-term heat production suggests that, in addition to converting fatty acids into heat energy, brown adipocytes may also adjust protein expression to raise their own temperature, to generate more heat. We expect this bimaterial microcantilever system to prove useful for determining a cell’s state by measuring thermal characteristics.     

Regulation of thymidine kinase activity in the cell cycle by a labile protein

Previous studies have shown that the onset of DNA synthesis in Balb/c 3T3 cells appears to be regulated by a labile protein. We have found that induction of thymidine kinase (TK) activity, after quiescent cells are stimulated by the addition of serum, is similarly regulated by a labile protein. Eight hours after serum stimulation, a 6-h pulse of cycloheximide (CHM) caused an excess delay of 2 h in TK induction. A similar delay also was found in the induction of thymidylate synthase (TS). In contrast, the benzo(a)pyrene transformed 3T3 cell line, BP-A31, which had previously been shown to have no excess delay for the onset of DNA synthesis also had no excess delay for the induction of TK activity after a pulse of CHM. The induction of TK was inhibited by actinomycin D and dichlororibofuranosylbenzimidizole (DRB) suggesting a requirement for new RNA synthesis. It did not appear to depend on DNA synthesis as it was not blocked by aphidicolin. In conclusion, the induction of TK activity appears to be regulated by the same labile cellular signal as the onset of DNA synthesis, and to depend on an increase in the level of TK mRNA in late G1 or early S phase.     

Effects of homozygosity and heterozygosity on certain properties of genetically defined electrophoretic variants of alcohol dehydrogenase isozymes in maize

Summary Genetically defined alcohol dehydrogenase isozymes have been studied. The faster electrophoretically migrating forms of both Adh-1 and Adh-2 have greater specific activity and are more heat stable than the alternative slow forms of the two isozymes. In the heterozygous state, it is confirmed by genetic dosage studies that the Adh-2 F/F homodimer is more catalytically active than the S/S homodimer. The heat stability relationship between the two homodimers is maintained in the heterozygous condition. Adh-1 also has the same heat stability when taken from the heterozygous as well as the homozygous background. The heterodimeric, hybrid molecules (F/S and S/F for Adh-2) have less catalytic activity than the F/F homodimer but more than the homodimer S/S enzyme. This is concluded from studies in the triploid endosperm where the effects of genetic dosing can be investigated. The hybrid enzymes have heat stability similar to the F/F homodimer. The finding that allelic forms of an enzyme can show altered properties of possible physiological advantage to the organism is discussed in relation to fitness of the alternative forms.     

Degradation of type I and II collagen by human activated C1-s

The activated first component of human complement, C1-s, was shown to cleave type I and II collagen and gelatin. The proteolytic activity was heat labile and was inhibited by a monoclonal antibody (M241) which recognized light chain of active human C1-s or by a serine protease inhibitor, DFP, but not by a chelating agent.     

Studies on the Bacterial Formation of a Peptide Antibiotic,Colistin

Colistinase, an enzyme inactivating colistin, which appeared in a fermented broth of Bacillus colistinus attending with decay of colistin production was studied. The enzyme was partially purified and also chromatographed by DEAE-cellulose column. It was a heat labile, mostly basic protein, had optimum pH at 9.0 and was active against peptide antibiotics such as colistin and polymyxin and also against casein. Bacterial proteinase, Nagarse, exhibited colistinase activity exclusively among tested proteinases and immunochemical studies revealed that colistinase was identical with bacterial proteinase Nagarse.     

Motor endplates in fast and slow muscles of the rat: what determines their difference?

Motor endplates in fast and slow skeletal muscles have different functional and morphological characteristics, and for brevity, are termed fast and slow respectively. We have examined the terminal arborization patterns of fast fibular and slow soleus axons 3-4 and 6 months after they reinnervated old preformed endplates or formed new ectopic endplates with denervated rat soleus muscles. Ectopic endplates formed by transplanted fibular and soleus nerves were fast and slow in appearance respectively. Both the fibular and the soleus nerves formed endplates of slow appearance when they reinnervated the original endplates. The fast appearance of ectopic fibular nerve endplates was unaffected by reinnervation of the original endplates by the slow soleus nerve. Dually innervated fibres had intermediate contraction speed compared to the fast fibres reinnervated only by the fibular nerve and the slow fibres reinnervated only by the soleus nerve. Continuous stimulation of the transplanted fibular nerve at 10 Hz for 3-4 months, starting just before the onset of ectopic endplate formation, prevented the increase in contraction speed seen without stimulation. The ectopic endplates of the stimulated axons were much smaller than usual and showed some signs of fast to slow transformation, but the transformation was incomplete and varied in degree between preparations. Transplanted soleus axons were less prone to growing along foreign pathways and to forming ectopic endplates than fibular axons.(ABSTRACT TRUNCATED AT 250 WORDS)     

Human urinary prokallikrein: rapid purification and model activation by trypsin

With only a three-step chromatographic procedure, human urinary prokallikrein has been purified completely. The active kallikrein also could be purified in the process of this purification. The prokallikrein was very rapidly activated by trypsin, followed thereafter by a very slow increase in the kallikrein activity. In the rapidly activated state, the molecular weight and the values of Km and Vmax were very similar to those of the purified active kallikrein. Only the slow increase in the activity was observed by tryptic digestion of the active kallikrein. The results suggest that the initial rapid activation is due to release of the propeptide and the slow reaction is due to a limited hydrolysis of the activated prokallikrein at sites which are not directly related to the active site.     

Characterization of phosphatidate phosphohydrolase activity associated with isolated lamellar bodies.

It has been demonstrated that lamellar bodies isolated from porcine lung have L-alpha-phosphatidate phosphohydrolase (EC 3.1.3.4) activity which cannot be accounted for by microsomal or mitochondrial contamination. Phosphatidate phosphohydrolase activity associated with the lamellar bodies is relatively insensitive to Mg2+ and more heat labile than the activity associated with whole lung microsomes. The enzyme was found to be the most active phosphohydrolase present in isolated lamellar bodies and is inhibited by 5 mM Be2+. Lamellar bodies isolated from human and rat lung tissue were also found to have this activity, and the functional role of the enzyme in lamellar bodies is proposed in relation to glycerophospholipid metabolism.