A 31P-n.m.r. study of the acute effects of beta-blockade on the bioenergetics of skeletal muscle during contraction.

1. The effects of beta-adrenoceptor antagonist administration on skeletal muscle contractile performance and bioenergetics in vivo have been investigated during unilateral sciatic nerve stimulation in the rat. 2. Two muscle stimulation protocols have been used: supramaximal stimulation at 4 Hz, or incremental supramaximal stimulation at 1, 2 and 4 Hz. Changes in high-energy phosphate concentrations were followed using 31P-n.m.r., and gastrocnemius muscle twitch characteristics were monitored continuously. 3. Under all conditions investigated, DL-propranolol administration (2.5 mg/kg body wt.) caused a significant decrease in cyclic AMP concentrations in resting and stimulated gastrocnemius muscle, prevented an increase in heart rate upon muscle stimulation, but did not affect plasma glucose, fatty acid or lactate concentrations in comparison with values obtained in control experiments. 4. Administration of DL-propranolol 5 min or 35 min before unilateral stimulation of 4 Hz had no effect on changes in muscle phosphocreatine, ATP or Pi concentrations, intracellular pH or contractile performance. 5. In contrast, animals receiving DL-propranolol 5 min before unilateral stimulation of 1, 2 and 4 Hz showed a significant deterioration in gastrocnemius muscle tension development during 2 and 4 Hz stimulation compared with control animals. Concurrent with this change in contractile performance was a higher muscle concentration of phosphocreatine, a lower concentration of Pi and no significant change in intramuscular pH compared with control experiments. 6. The changes in muscle performance and bioenergetics observed during the incremental stimulation protocol were not observed when D-propranolol was administered and could be completely circumvented by a short period of muscle stimulation of 4 Hz prior to initiation of the incremental stimulation protocol. 7. Mechanisms are discussed which may account for the failure of gastrocnemius muscle to generate the expected force during the incremental stimulation protocol in the presence of beta-blockade.     

A quartz crystal biosensor for measurement in liquids.

The detection of anti-human immunodeficiency virus (HIV) antibodies by means of synthetic HIV peptide immobilized on a piezoelectric quartz sensor is demonstrated. The measurement set-up consists of an oscillator circuit, a suitably modified AT-cut thickness-shear-mode quartz crystal with gold electrodes, which is housed in a special reaction vessel, and a computer-controlled frequency counter for the registration of the measured frequency values. The quartz crystal is adapted for a steady operation in liquids at a frequency of 20 MHz. In phosphate-buffered saline solution the oscillator reaches a stability of about 0.5 Hz within a few seconds, of about 2 Hz within 10 min and about 30 Hz within 1 h. The frequency shift due to the adsorption of various proteins to the uncoated sensor surface has been investigated. It can be shown that a stable adsorptive binding of proteins to an oscillating gold surface is feasible and can be used for the immobilization of a receptor layer (e.g. HIV peptide). Specific binding of the anti-HIV monoclonal antibody to the HIV peptide immobilized on the quartz sensor is demonstrated. Control experiments show, however, additional unspecific binding. According to the experiments, the Sauerbrey formula gives a sufficiently accurate value for the decrease of the resonant frequency due to adsorption or binding of macromolecular proteins on the quartz crystal surface.     

13C solid-state NMR of gramicidin A in a lipid membrane.

The natural-abundance 13C NMR spectrum of gramicidin A in a lipid membrane was acquired under magic-angle spinning conditions. With fast sample spinning (15 kHz) at approximately 65 degrees C the peaks from several of the aliphatic, beta-, alpha-, aromatic, and carbonyl carbons in the peptide could be resolved. The resolution in the 13C spectrum was superior that observed with 1H NMR under similar conditions. The 13C linewidths were in the range 30-100 Hz, except for the alpha- and beta-carbons, the widths of which were approximately 350 Hz. The beta-sheet-like local structure of gramicidin A was observed as an upfield shift of the gramicidin alpha and carbonyl resonances. Under slow sample spinning (500 Hz), the intensity of the spinning sidebands from 13C in the backbone carbonyls was used to determine the residual chemical shift tensor. As expected, the elements of the residual chemical shift tensor were consistent with the single-stranded, right-handed beta6.3 helix structure proposed for gramicidin A in lipid membranes.     

Biological effects of low-frequency pulsed magnetic fields on the embryonic central nervous system development. A histological and histochemical study

Numerous experiments have yielded contradictory results on the harmful action of magnetic fields on embryonic development. Pulsed magnetic fields appear to be able to delay normal development of embryos. In the present study, fertilized Gallus domesticus eggs were exposed during incubation to pulsed magnetic fields (harmonic signals of 10 μT for 1 second with silences of 0.5 seconds) of 50 or 100 Hz frequency. Embryos extracted at 45 h of exposure to fields of 50 Hz or 100 Hz frequency had significantly (p<0.05) fewer somite pairs compared with controls of the same age. At 15 days of incubation, only embryos exposed to a 10 μT- 50 Hz field had a significantly (p<0.05) higher somatic weight. At 21 days of incubation, a significantly lower somatic weight (p<0.01) and development stage (p<0.05) was found in embryos exposed to a 10 μT-100 Hz field than in controls, while a lower development stage (p<0.05) alone was observed in those exposed to a 10 μT-50 Hz field. In addition, animals showed higher expression of the neural marker NSE (neural specific enolase) after 21 days of development as determined by immunohistochemistry, with very low expression of glycosaminoglycans identified by alcyan blue staining. These results suggest that pulsed magnetic fields may be able to hinder normal embryonic development in vivo and to alter normal neural function, at least at the intensities and frequencies analyzed in the present study.     

A cyclic model for bimodal activation of calcium activated potassium channels in radish vacuoles.

This paper presents the mathematical framework of a cyclic model proposed for describing the transition between a fast and a slow mode (fast-slow effect) induced by the application of step membrane potentials to ion channels from radish vacuoles. A voltage stimulation pulse with frequency in the range of 2 Hz or higher increased the activation time (slow mode) of the recorded currents. When the frequency of the stimulation pattern was restored to 0.1 Hz the activation time decreased twofold (fast mode). This experimental result cannot be explained by classical kinetic theory. The model, based on a simple extension of the Hodgkin and Huxley chain, describes the whole current experimental data and provides hints on the structural conformation of ion channels.     

Temperature and auditory thresholds: Bioacoustic studies of the frogsRana r. ridibunda,Hyla a. arborea andHyla a. savignyi (Anura,amphibia)

Summary The auditory thresholds of three frogs-two subspecies of the genusHyla (H. a. arborea, H. a. savignyi) and one of the genusRana (R. r. ridibunda)—were measured at 5°, 12°, 20° and 28°C, by recording multi-unit activity from the torus semicircularis. In the tree frogs, the upper limit of the audible range is 7,000 Hz. At 5°C the best frequency is 3,000 Hz; the threshold (expressed in dB SPL in all cases) at this frequency is 49 dB (males) and 43 dB (females) forH. a. arborea and 42 dB (males) and 48 dB (females) forH. a. savignyi. At 12°C the thresholds are lower, and they are lower still at 20°, reaching a minimum, at 3,000 Hz, of 42 dB (males) and 38 dB (females) forH. a. arborea and 41 dB (males) and 40 dB (females) forH. a. savignyi. At frequencies of 1,000 Hz and lower, thresholds are high at 5°C; in part of this range they are considerably lowered at 20°C, whereas at 28°C there is a reduction in sensitivity to most frequencies inH. a. arborea, amounting to more than 10 dB in the males.H. a. savignyi differs in this regard; at 28° sensitivity is no less than at lower temperatures, and in fact is greater in the range 1,000–1,400 Hz. The audible range ofR. r. ridibunda is more restricted than that of the tree frogs, but it is more sensitive within this range. The highest frequency is 4,500 Hz. At 5°C the thresholds of the males are lowest at 500–600 Hz (42 dB) and 1,400–1,900 Hz (ca. 39 dB). The best frequencies of the females are 700 Hz (38 dB) and 1,400 Hz (36 dB). At 12°C the thresholds at 300 Hz and 1,000 Hz are markedly lowered, by 10–18 dB. The thresholds of the females at 20°C are still lower over almost the entire audible range, whereas in the males only part of the range is affected. This difference persists at 28°C, the threshold curve of the males being slightly raised, while that of the females is unchanged. Latencies are dependent upon temperature and sound pressure. With a rise in temperature from 5° to 20°C the latency falls by ca. 8 ms. An increase in sound pressure from 5 dB to 30 dB SPL shortens the latency by ca. 10 ms. These changes were found in all the frogs studied.     

A new micromethod for mechano-chemical research.

A new micromethod to study activity of enzymes in crystalline and amorphous solid samples subjected to tension is described. Both static (up to 200%) and dynamic (with an amplitude up to 50% and frequency of 10(-3)-50 Hz) deformation may be applied to the sample during the activity measurement. Strain-induced changes in activity of thin (3-10 microns) needle-like strips (0.2-0.5 microns long), cut out of the microtome sections of cross-linked crystals of pancreatic carboxypeptidase A is measured in a droplet of substrate solution (with volume of 10(-6)-10(-9) l) with a pH-microelectrode technique.     

The effect of sampling frequency on EMG measures of occupational mechanical exposure.

The purpose of the present study was to determine how much it is possible to downsample EMG without loosing the accuracy of some EMG measures of occupational mechanical exposure frequently used in the literature. The EMG signals of four muscles (two trapezius, right deltoid and right extensor digitorum) were collected (sampling frequency: 1024 Hz) from 20 subjects while performing a 20 min computer work task. The EMG RMS amplitude was computed from 0.125 s successive time-windows for the original (1024 Hz) and four additional digitally resampled (at 512, 256, 128 and 64 Hz) EMG signals. Three of the most frequently used data reduction methods (1. gaps analysis, 2. amplitude probability distribution function and 3. exposure variation analysis) were used to summarise the 20 min EMG activation profiles. The results from the exposure variation analysis were summarized into three variables using a new method detailed here. In general, the ANOVA for repeated measures demonstrated that a decrease of the sampling frequency significantly changed the EMG measures (relative to the 1024 Hz sampling condition) most of the time at 64 and 128 Hz, occasionally at 256 Hz, but practically never at 512 Hz. An analysis of the maximal errors (relative to the 1024 Hz condition) across all subjects supported these findings. Consequently, it was concluded that 512 Hz is quite conservative and should practically never lead to invalid EMG parameters estimations. Conversely, 256 Hz represents the lowest limit tolerable for some EMG parameters (gaps analysis, amplitude probability distribution function) while it is unacceptable for others (modified exposure variation analysis).     

Tetanic depression in fast motor units of the cat gastrocnemius muscle.

Ability of muscle fibers to generate force is decreased when higher frequency of stimulation of motor units immediately follows lower frequency. This phenomenon called tetanic depression was found in rat medial gastrocnemius. However, it was not clear whether tetanic depression occurred only in rat muscle or it concerns all mammals. This study was conducted on motor units of cat medial gastrocnemius. Analyses were made at three successive trains of stimulation: 30 Hz, 20 and 30 Hz and again 30 Hz (the first pattern) or 40 Hz, 25 and 40 Hz and 40 Hz (the second pattern). In all fast units force generated within the middle tetanus was lower than force generated at the same, but constant frequency of stimulation applied earlier or later. The mean tetanic depression in 30 Hz tetani amounted to 10.9% for fast fatigable (FF) and 15.9% for fast resistant (FR) motor units, whereas in 40 Hz tetani mean values were 5.6% and 7.3% for FF and FR motor units, respectively. In slow motor units tetanic depression was not observed. These results proved the existence of tetanic depression in the feline muscle and indicated that its intensity depends on the fusion of tetanus. It has been concluded, that the tetanic depression is a general property of fast motor units in mammals.     

31P NMR of Mg-ATP in dilute solutions: complexation and exchange.

1. Monovalent-cation [(CH3)4N+, K(I), Na(I)] ATP, 1 mM in nucleotide, in aqueous solutions at pH 7.2, 24 degrees C, generates 2 different 31P NMR spectra, depending upon the salt content of the solution. At salt concentrations below 10 mM, the 31P NMR signals are chemically-shifted upfield (Na salt: alpha, -11.44 delta; beta, -22.91 delta; gamma, -8.36 delta) and the beta- and gamma-groups are broadened (at half-height: alpha, 3.5 Hz; beta, 9.6 Hz; gamma, 69 Hz). Above 10 mM salt, the signals are shifted downfield and are narrow (Na salt: alpha, -11.09 delta, 1.9 Hz; beta, -21.75 delta, 3.3 Hz; gamma, -6.30 delta, 3.9 Hz). 2. The Na-Mg-ATP complex, corresponding to the composition Na6Mg1ATP2, yields a single set of 31P resonances at concentrations of nucleotide of 100 mM, that upon dilution to 0.2 mM, resolve into 2 sets of ATP resonances characterized by low-field and high-field beta- and gamma-group resonance pairs. This set of ATP resonances, in contrast to the resonance set at 100 mM ATP, are broad (100 mM in ATP: alpha, -10.7 delta, 3.7 Hz; beta, -20.1 delta, 15 Hz; gamma, -5.7 delta, 7.3 Hz. 0.2 mM in ATP: alpha, -10.7 delta, 47 Hz; beta, -18.8 and -21.6 delta, 316 and 274 Hz; gamma, -5.5 and -8.7 delta, 460 and 374 Hz). 3. This new data, in combination with data derived from a survey of metal-ion-ATP studies, are interpreted in terms of ATP dimers, incorporating 2 molecules of ATP and 2 metal cations, that exist in water under the physiological conditions of neutral pH, high salt content [135 mM K(I)] and ATP concentrations in the range of 3 mM. 4. A compilation of 31P in vivo and ex vivo data compared to a reference Mg-ATP chemical shift vs Mg/ATP ratio plot indicates that ATP is not fully Mg-saturated in living systems and that 41% exists as the Mg(ATP)2 complex.     

"Resonance-like" frequencies of sensorimotor areas evoked by repetitive tactile stimulation.

The aim of the present study was to investigate resonance-like frequencies of somatosensory steady-state evoked potentials (SSSEPs) during mechanical stimulation of the left and right index finger. The resonance effects were studied in a group of normal subjects at 8 frequencies between 17 Hz and 31 Hz. Band power was computed in 2 s epochs in eight 2 Hz beta bands starting from 16-18 Hz up to 30-32 Hz and referred to a baseline prior to stimulation. For each subject the band with the largest band power increase was determined on electrodes overlying the contralateral hand area. The band power maxima and therewith the "resonance-like" frequencies were found around 27 Hz.     

A low frequency active filter with plug-in tuning impedances.

A new approach to the construction of low frequency (1 Hz -- 20 kHz) active filters, based on the application of a pair of IC operational amplifiers forming a separate module and a plug-in unit containing RC tuning circuits, is described. The electronic circuits and the design of the filters are presented with all details.     

Electroacupuncture analgesia could be mediated by at least two pain-relieving mechanisms; endorphin and non-endorphin systems.

This present paper shows different levels of electroacupuncture analgesia (antinociceptive effect) induced by three different frequencies of stimulation (i.e. 0.2, 4 and 200 Hz); highest analgesia is induced at 200 Hz and lowest at 0.2 Hz. Naloxone (1 mg/kg) completely reverses the electroacupuncture effects at low frequency stimulation (4 Hz) but produces no inhibition at high frequency stimulation (200 Hz). Conversely, parachlorophenylalanine (320 mg/kg) partially blocks the high-frequency (200 Hz) analgesia but produces no effect on the low-frequency (4 Hz) electroacupuncture analgesia. This suggests that electroacupuncture analgesia induced by low frequency stimulation may be mediated by endorphins while high frequency stimulation is not endorphinergic but may be partly due to serotonin.     

弱极低频磁场对Actin骨架组装效率的频率窗口效应初探

前期研究发现,50 Hz弱磁场辐照能明显降低细胞的微丝含量和组装效率,对actin骨架形态也有明显影响.电磁生物学效应是否与辐照场频率相关,一直受到研究者的关注.单体球状肌动蛋白(G-actin)是带电结构,电磁场频率会影响其振荡频率并对微丝聚合效率产生影响.本文从细胞骨架形态和蛋白质两层次,采用免疫荧光技术考察0.4 m T,在35~140 Hz范围内5个频率的极低频磁场(ELF-MF)对FL细胞中纤维状肌动蛋白(F-actin)含量的影响,并采用荧光共振能量转移技术(FRET)验证效应最明显的频率对离体G-actin组装效率的干扰程度.结果显示,相比假辐照组,细胞中F-actin含量在50 Hz辐照组下降了(34.66±3.14)%,110 Hz次之,而另外3组(35、70和140 Hz)无显著性差异.同时利用FRET方法验证,在50 Hz磁场辐照下,离体环境中G-actin组装成F-actin的效率较假辐照组、35和70 Hz组显著降低.经初步分析,G-actin在弱ELF-MF中受到以洛伦兹力和感生电场力的合力为主的相关电磁力干扰,致使组装效率下降,且由于工频磁场周期与微丝组装周期的特殊相干性,在50 Hz频率附近可能存在一个外磁场干扰actin骨架组装的频率窗口.     

Characterization of enzyme-bound ligand dynamics by solid-state NMR in the presence of ligand exchange: L-phenylalanine on carboxypeptidase A.

Deuterium NMR spectra were obtained for L-phenylalanine-d5, deuterated on the phenyl ring, in cross-linked polycrystalline samples of carboxypeptidase A containing different amounts of water. The deuterium powder pattern line shapes are simulated by extension of the theory to include both a local reorientational motion of the bound L-phenylalanine phenyl ring and exchange of the L-phenylalanine with an intracrystalline isotropic environment. The spectral simulations are consistent with the phenyl ring of the phenylalanine executing pi-flips in the bound environment at rates that vary from 3 x 10(4) Hz at 6% water content to 1 x 10(5) Hz at 21% water content. At all water contents studied, the ligand exchanges with an essentially isotropic environment in the crystal with a rate constant of approximately 2.5 x 10(-3) Hz. Although the dissociation constant for the L-phenylalanine is only 18 mM, the spectral simulations that reproduce the experimental line shape well do not require significant wobble of the phenyl ring rotation axis, which is consistent with the binding interactions identified by x-ray crystallography.     

Neonatal murine heart slices. A robust model to study ventricular isometric contractions.

BACKGROUND/AIMS: Cardiac function is increasingly studied using murine models. However, current multicellular preparations to investigate contractile properties have substantial technical and biological limitations and are especially difficult to apply to the developing murine heart. METHODS: Newborn murine hearts were cut with a vibratome into viable tissue slices. The structural and functional integrity of the tissue was shown by histology, ATP content and sharp electrode recordings. RESULTS: Within the first 48 hours after slicing structure remained intact without induction of apoptosis. ATP concentrations and action potential parameters were comparable to those of physiological tissue. Isometric force measurements demonstrated a physiological force-frequency relationship with a ;primary-phase' negative force-frequency relationship up to 1-2 Hz and a ;secondary-phase' positive force-frequency relationship up to 8 Hz. (-)-Isoproterenol (10(-6) mol/l) increased active force to 251 +/- 35% (n=15) of baseline values and shortened relaxation times indicating a preserved beta-adrenergic regulation of contraction. Changes of the force-frequency relationship after application of ryanodine and nifedipine indicated functionality of calcium release from the sarcoplasmic reticulum and of L-type calcium channels. CONCLUSION: Generation of viable, physiological intact ventricular slices from neonatal hearts is feasible and provides a robust model to study loaded contractions.     

Control of glycogenolysis and hemodynamics in perfused rat liver by the sympathetic innervation. Dependence on stimulation frequency and duration

Perivascular stimulation of the hepatic nerves in the in situ perfused rat liver with a constant frequency of 20 Hz over a constant period of 5 min had previously been shown to cause an increase of glucose output, a shift from lactate uptake to release, a reduction in perfusion flow (Hartmann et al. (1982) Eur. J. Biochem. 123, 521-526) and an overflow of noradrenaline into the hepatic vein (Beckh et al. (1982) FEBS Lett. 149, 261-265). In the present study the dependence of the metabolic and hemodynamic effects on the frequency between 1 and 30 Hz and duration of stimulation between 0.5 and 5 min was investigated. Over a constant stimulation period of 5 min the alteration in glucose exchange was maximal with a frequency of 10 Hz and half-maximal with 4 Hz. The corresponding values for the exchange of lactate were 5 Hz and 2 Hz, respectively, and for the perfusion flow 2.5 Hz and 1.5 Hz, respectively. An increase of noradrenaline overflow was not observed with the lower frequencies of 1 and 2.5 Hz; it was maximal at 10 Hz and half-maximal at 6.5 Hz. At a constant frequency of 20 Hz the increase in glucose release was maximal with a total stimulation period of 1 min and half-maximal with a period of 0.4 min. An essentially maximal alteration of lactate exchange and perfusion flow as well as of noradrenaline overflow was also effected by a stimulation period of 1 min.(ABSTRACT TRUNCATED AT 250 WORDS)     

Diaphyseal bone formation in murine tibiae in response to knee loading.

Mechanical stimulation is critical for bone architecture and bone mass. The aim of this study was to examine the effects of mechanical loads applied to the knee. The specific question was whether loads applied to the tibial epiphysis would enhance bone formation in the tibial diaphysis. In C57/BL/6 mice, loads of 0.5 N were applied for 3 min per day for 3 days at 5, 10, or 15 Hz. Bone samples were harvested 13 days after the last loading. The strains were measured 13 +/- 2 microstrains at 5 Hz in the diaphysis. The histomorphometric data in the diaphysis clearly showed enhanced bone formation. First, compared with nonloaded control the cross-sectional cortical area was increased by 11% at 5 Hz and 8% at 10 Hz (both P < 0.05). Second, the cortical thickness was elevated by 12% at 5 Hz (P < 0.01) and 8% at 10 Hz (P < 0.05). Third, mineralizing surface (MS/BS), mineral apposition rate (MAR), and bone formation rate (BFR/BS) were increased at 5 Hz (P < 0.01 for MS/BS; P < 0.001 for MAR and BFR/BS) and at 10 Hz (P < 0.05 for MS/BS; P < 0.01 for MAR and BFR/BS). Bone formation was enhanced more extensively in the medial side than the lateral or the posterior side. The results reveal that knee loading is an effective means to enhance bone formation in the tibial diaphysis in a loading-frequency dependent manner without inducing significant in situ strain at the site of bone formation.     

Study of H-2 mutations in mice. IV. A comparison of the mutants M505 and Hz1 by skin grafting and serological techniques

The line B6.M505 is congenic with C57BL/6JY and carries a mutant form of theH-2 ^b haplotype designatedH-2 ^bd . The mutant site 505 was located by the F₁ tests in theK end of theH-2 gene complex. The M505 mice are histoincompatible with the B6.C(Hz1) line (haplotypeH-2 ^ba ) carrying another mutation in theK end ofH-2 ^b . Inability of M505 to complement Hz1 in tests with B6 skin grafting is considered as an evidence that the same gene was altered by both mutations. The gained H antigens of two mutants can cross-react in vivo as revealed by accelerated rejection of Hz1 skin grafts by B6 recipients presensitized with M505 spleen cells. The lost antigenic determinants are not identical as shown by accelerated rejection of B6 skin grafts by Hz1 hosts preimmunized with M505 spleen cells. Absorptions of the antiserum ASY-015, (d×a) anti-i, anti-H-2.33 with M505 spleen cells did not clear forH-2 ⁱ ,H-2 ^b andH-2 ^ba , and absorptions with Hz1 did not clear forH-2 ⁱ ,H-2 ^b , andH-2 ^bd . These results show that changes of histocompatibility determinants may be accompanied by loss of some haptenic determinants in the Hz1 and M505 mutations.     

听觉脑干诱发电位的频谱分析和相关分析

本文研究了正常受试者和有听力障碍病人的听觉脑干诱发电位反应(ABR)的频谱分析及相关分析。正常受试者的ABR频谱中有三个主要峰,其中心频率位置大约在200Hz,500Hz及800～1000Hz左右,而在1200Hz以上则很少成分。 有听力障碍病人的ABR,其频谱幅值比正常受试者的要小得多,整个频谱向低端偏移,频谱频率成分主要在600Hz以下。 正常受试者和某个基准ABR之间的互相关函数,在最早的0.5ms内达到最大值,然后逐渐减小。异常ABR和基准ABR之间的互相关函数则没有明显的最大值,整个曲线比较平坦。