Calcium- and myosin-dependent changes in troponin structure during activation of heart muscle

Each heartbeat is triggered by a pulse of intracellular calcium ions which bind to troponin on the actin-containing thin filaments of heart muscle cells, initiating a change in filament structure that allows myosin to bind and generate force. We investigated the molecular mechanism of calcium regulation in demembranated trabeculae from rat ventricle using polarized fluorescence from probes on troponin C (TnC). Native TnC was replaced by double-cysteine mutants of human cardiac TnC with bifunctional rhodamine attached along either the C helix, adjacent to the regulatory Ca²⁺-binding site, or the E helix in the IT arm of the troponin complex. Changes in the orientation of both troponin helices had the same steep Ca²⁺ dependence as active force production, with a Hill coefficient ( n _H) close to 3, consistent with a single co-operative transition controlled by Ca²⁺ binding. Complete inhibition of active force by 25 μ m blebbistatin had very little effect on the Ca²⁺-dependent structural changes and in particular did not significantly reduce the value of n _H. Binding of rigor myosin heads to thin filaments following MgATP depletion in the absence of Ca²⁺ also changed the orientation of the C and E helices, and addition of Ca²⁺ in rigor produced further changes characterized by increased Ca²⁺ affinity but with n _H close to 1. These results show that, although myosin binding can switch on thin filaments in rigor conditions, it does not contribute significantly under physiological conditions. The physiological mechanism of co-operative Ca²⁺ regulation of cardiac contractility must therefore be intrinsic to the thin filaments.     

Temperature dependence of the force-generating process in single fibres from frog skeletal muscle

Generation of force and shortening in striated muscle is due to the cyclic interactions of the globular portion (the head) of the myosin molecule, extending from the thick filament, with the actin filament. The work produced in each interaction is due to a conformational change (the working stroke) driven by the hydrolysis of ATP on the catalytic site of the myosin head. However, the precise mechanism and the size of the force and length step generated in one interaction are still under question. Here we reinvestigate the endothermic nature of the force-generating process by precisely determining, in tetanised intact frog muscle fibres under sarcomere length control, the effect of temperature on both isometric force and force response to length changes. We show that raising the temperature: (1) increases the force and the strain of the myosin heads attached in the isometric contraction by the same amount (∼70 %, from 2 to 17 °C); (2) increases the rate of quick force recovery following small length steps (range between −3 and 2 nm (half-sarcomere)⁻¹) with a Q ₁₀ (between 2 and 12 °C) of 1.9 (releases) and 2.3 (stretches); (3) does not affect the maximum extent of filament sliding accounted for by the working stroke in the attached heads (10 nm (half-sarcomere)⁻¹). These results indicate that in isometric conditions the structural change leading to force generation in the attached myosin heads can be modulated by temperature at the expense of the structural change responsible for the working stroke that drives filament sliding. The energy stored in the elasticity of the attached myosin heads at the plateau of the isometric tetanus increases with temperature, but even at high temperature this energy is only a fraction of the mechanical energy released by attached heads during filament sliding.     

Temperature change does not affect force between regulated actin filaments and heavy meromyosin in single-molecule experiments

The temperature dependence of sliding velocity, force and the number of cross-bridges was studied on regulated actin filaments (reconstituted thin filaments) when they were placed on heavy meromyosin (HMM) attached to a glass surface. The regulated actin filaments were used because our previous study on muscle fibres demonstrated that the temperature effect was much reduced in the absence of regulatory proteins. A fluorescently labelled thin filament was attached to the gelsolin-coated surface of a polystyrene bead. The bead was trapped by optical tweezers, and HMM–thin filament interaction was performed at 20–35°C to study the temperature dependence of force at the single-molecule level. Our experiments showed that there was a small increase in force with temperature  ( Q ₁₀= 1.43)  and sliding velocity  ( Q ₁₀= 1.46)  . The small increase in force was correlated with the small increase in the number of cross-bridges  ( Q ₁₀= 1.49)  , and when force was divided by the number of cross-bridges, the result did not depend on the temperature  ( Q ₁₀= 1.03)  . These results demonstrate that the force each cross-bridge generates is fixed and independent of temperature. Our additional experiments demonstrate that tropomyosin (Tm) in the presence of troponin (Tn) and Ca²⁺ enhances both force and velocity, and a truncated mutant, Δ23Tm, diminishes force and velocity. These results are consistent with the hypothesis that Tm in the presence of Tn and Ca²⁺ exerts a positive allosteric effect on actin to make actomyosin linkage more secure so that larger forces can be generated.     

Actomyosin energy turnover declines while force remains constant during isometric muscle contraction

Energy turnover was measured during isometric contractions of intact and Triton-permeabilized white fibres from dogfish ( Scyliorhinus canicula ) at 12°C. Heat + work from actomyosin in intact fibres was determined from the dependence of heat + work output on filament overlap. Inorganic phosphate (P_i) release by permeabilized fibres was recorded using the fluorescent protein MDCC-PBP, N-(2-[1-maleimidyl]ethyl)-7-diethylamino-coumarin-3 carboxamide phosphate binding protein. The steady-state ADP release rate was measured using a linked enzyme assay. The rates decreased five-fold during contraction in both intact and permeabilized fibres. In intact fibres the rate of heat + work output by actomyosin decreased from 134 ± s.e.m. 28 μW mg⁻¹ ( n = 17) at 0.055 s to 42% of this value at 0.25 s, and to 20% at 3.5 s. The force remained constant between 0.25 and 3.5 s. Similarly in permeabilized fibres the P_i release rate decreased from 5.00 ± 0.39 mmol l⁻¹ s⁻¹ at 0.055 s to 39% of this value at 0.25 s and to 19% at 0.5 s. The steady-state ADP release rate at 15 s was 21% of the P_i rate at 0.055 s. Using a single set of rate constants, the time courses of force, heat + work and P_i release were described by an actomyosin model that took account of the transition from the initial state (rest or rigor) to the contracting state, shortening and the consequent work against series elasticity, and reaction heats. The model suggests that increasing P_i concentration slows the cycle in intact fibres, and that changes in ATP and ADP slow the cycle in permeabilized fibres.     

Inadequate heat release from the human brain during prolonged exercise with hyperthermia

Brain temperature appears to be an important factor affecting motor activity, but it is not known to what extent brain temperature increases during prolonged exercise in humans. Cerebral heat exchange was therefore evaluated in seven males during exercise with and without hyperthermia. Middle cerebral artery mean blood velocity (MCA V _mean) was continuously monitored while global cerebral blood flow (CBF) and cerebral energy turnover were determined at the end of the two exercise trials in three subjects. The arterial to venous temperature difference across the brain (v-a D _temp) was determined via thermocouples placed in the internal jugular vein and in the aorta. The jugular venous blood temperature was always higher than that of the arterial blood, demonstrating that heat was released via the CBF during the normothermic as well as the hyperthermic exercise condition. However, heat removal via the jugular venous blood was 30 ± 6 % lower during hyperthermia compared to the control trial. The reduced heat removal from the brain was mainly a result of a 20 ± 6 % lower CBF (22 ± 9 % reduction in MCA V _mean), because the v-a D _temp was not significantly different in the hyperthermic (0.20 ± 0.05 °C) compared to the control trial (0.22 ± 0.05 °C). During hyperthermia, the impaired heat removal via the blood was combined with a 7 ± 2 % higher heat production in the brain and heat was consequently stored in the brain at a rate of 0.20 ± 0.06 J g⁻¹ min⁻¹. The present results indicate that the average brain temperature is at least 0.2 °C higher than that of the body core during exercise with or without hyperthermia.     

Oxygen Partial Pressure in Outer Layers of Skin of Human Finger Nail Folds

To gain insight into oxygen transport by the cutaneous microcirculation, we have developed oxygen-sensitive microelectrodes (tip diameter ∼5 μm) to measure the distribution of P _O2 in dermal papillae of the finger nail folds of healthy human subjects. Oxygen entry into the tissue was minimised by covering the skin with a layer of paraffin oil. The finger was held under a dissecting microscope and microelectrodes were guided into position. P _O2 varied from 5–25 % of its atmospheric value, P _air (∼160 mmHg), depending on the location within the papilla. Along the axis of a papillary loop, P _O2 decreased from 40.0 ± 4.8 mmHg (mean ± s.e.m. ,   n = 6  ) at the base to 30.4 ± 5.2 mmHg (   n = 6  ) at the tip. The lowest values of P _O2, in the range of 5 % of P _air, were measured in the epidermis where the metabolism of cells was highest and the steepest P _O2 gradients were recorded in the vicinity of the epidermal–dermal boundary. When the local circulation was abruptly reduced or stopped, P _O2 fell exponentially with time, with a time constant of 8.4 ± 1.5 s (   n = 7  ). When flow was reinstated, P _O2 rose exponentially to a new value with a time constant of 4.8 ± 0.8 s (   n = 6  ). The steady state P _O2 following reperfusion was ∼23 % higher than the pre-occlusion value ( P < 0.05, ANOVA and two-tailed Student's t test) indicating localised reactive hyperaemia.     

Acute dopamine/noradrenaline reuptake inhibition enhances human exercise performance in warm, but not temperate conditions

Nine healthy endurance-trained males were recruited to examine the effect of a dual dopamine/noradrenaline reuptake inhibitor on performance, thermoregulation and the hormonal responses to exercise. Subjects performed four trials, ingesting either a placebo (pla) or 2 × 300 mg bupropion (bup), prior to exercise in temperate (18°C) or warm (30°C) conditions. Trials consisted of 60 min cycle exercise at 55% W _max immediately followed by a time trial (TT). TT performance in the heat was significantly improved by bupropion (pla: 39.8 ± 3.9 min, bup: 36.4 ± 5.7 min; P = 0.046), but no difference between treatments was apparent in temperate conditions (pla: 30.6 ± 2.2 min, bup: 30.6 ± 1.9 min; P = 0.954). While TT power output was consistently lower in the heat when compared to temperate conditions, this decrement was attenuated by bupropion. At the end of the TT in the heat, both core temperature (pla 39.7 ± 0.3°C, bup 40.0 ± 0.3°C; P = 0.017) and HR (pla 178 ± 7 beats min⁻¹, bup 183 ± 12 beats min⁻¹; P = 0.039), were higher in the bupropion trial than in the placebo. Circulating pituitary and adrenal hormone concentrations increased throughout exercise in all trials. Circulating serum prolactin was elevated above temperate levels during exercise in a warm environment ( P < 0.001). These data indicate that performance in warm conditions is enhanced by acute administration of a dual dopamine/noradrenaline reuptake inhibitor. No such effect was apparent under temperate conditions. It appears that bupropion enabled subjects to maintain a greater TT power output in the heat with the same perception of effort and thermal stress reported during the placebo trial, despite the attainment of a higher core temperature.     

Change in contractile properties of human muscle in relationship to the loss of power and slowing of relaxation seen with fatigue

Slow relaxation from an isometric contraction is characteristic of acutely fatigued muscle and is associated with a decrease in the maximum velocity of unloaded shortening ( V _max) and both these phenomena might be due to a decreased rate of cross bridge detachment. We have compared the change in relaxation rate with that of various parameters of the force–velocity relationship over the course of an ischaemic series of fatiguing contractions and subsequent recovery using the human adductor pollicis muscle working in vivo at approximately 37°C in nine healthy young subjects. Maximal isometric force ( F ₀) decreased from 91.0 ± 1.9 to 58.3 ± 3.5 N (mean ± s.e.m. ). Maximum power decreased from 53.6 ± 4.0 to 17.7 ± 1.2 (arbitrary units) while relaxation rate declined from −10.3 ± 0.38 to −2.56 ± 0.29 s⁻¹. V _max showed a smaller relative change from 673 ± 20 to 560 ± 46 deg s⁻¹ and with a time course that differed markedly from that of slowing of relaxation, showing very little change until late in the series of contractions. Curvature of the force–velocity relationship increased ( a/F ₀ decreasing from 0.22 ± 0.02 to 0.11 ± 0.02) with fatigue and with a time course that was similar to that of the loss of power and the slowing of relaxation. It is concluded that for human muscle working at a normal physiological temperature the change in curvature of the force–velocity relationship with fatigue is a major cause of loss of power and may share a common underlying mechanism with the slowing of relaxation from an isometric contraction.     

Reactive oxygen species reduce myofibrillar Ca2+ sensitivity in fatiguing mouse skeletal muscle at 37°C

The mechanisms of muscle fatigue were studied in small muscle bundles and single fibres isolated from the flexor digitorum brevis of the mouse. Fatigue caused by repeated isometric tetani was accelerated at body temperature (37°C) when compared to room temperature (22°C). The membrane-permeant reactive oxygen species (ROS) scavenger, Tiron (5 m m ), had no effect on the rate of fatigue at 22°C but slowed the rate of fatigue at 37°C to that observed at 22°C. Single fibres were microinjected with indo-1 to measure intracellular calcium. In the accelerated fatigue at 37°C the tetanic [Ca²⁺]_i did not change significantly and the decline of maximum Ca²⁺-activated force was similar to that observed at 22°C. The cause of the greater rate of fatigue at 37°C was a large fall in myofibrillar Ca²⁺ sensitivity. In the presence of Tiron, the large fall in Ca²⁺ sensitivity was abolished and the usual decline in tetanic [Ca²⁺]_i was observed. This study confirms the importance of ROS in fatigue at 37°C and shows that the mechanism of action of ROS is a decline in myofibrillar Ca²⁺ sensitivity.     

Acceleration of AMPA receptor kinetics underlies temperature-dependent changes in synaptic strength at the rat calyx of Held

It is well established that synaptic transmission declines at temperatures below physiological, but many in vitro studies are conducted at lower temperatures. Recent evidence suggests that temperature-dependent changes in presynaptic mechanisms remain in overall equilibrium and have little effect on transmitter release at low transmission frequencies. Our objective was to examine the postsynaptic effects of temperature. Whole-cell patch-clamp recordings from principal neurons in the medial nucleus of the trapezoid body showed that a rise from 25°C to 35°C increased miniature EPSC (mEPSC) amplitude from −33 ± 2.3 to −46 ± 5.7 pA ( n = 6) and accelerated mEPSC kinetics. Evoked EPSC amplitude increased from −3.14 ± 0.59 to −4.15 ± 0.73 nA with the fast decay time constant accelerating from 0.75 ± 0.09 ms at 25°C to 0.56 ± 0.08 ms at 35°C. Direct application of glutamate produced currents which similarly increased in amplitude from −0.76 ± 0.10 nA at 25°C to −1.11 ± 0.19 nA 35°C. Kinetic modelling of fast AMPA receptors showed that a temperature-dependent scaling of all reaction rate constants by a single multiplicative factor ( Q ₁₀= 2.4) drives AMPA channels with multiple subconductances into the higher-conducting states at higher temperature. Furthermore, Monte Carlo simulation and deconvolution analysis of transmission at the calyx of Held showed that this acceleration of the receptor kinetics explained the temperature dependence of both the mEPSC and evoked EPSC. We propose that acceleration in postsynaptic AMPA receptor kinetics, rather than altered presynaptic release, is the primary mechanism by which temperature changes alter synaptic responses at low frequencies.     

The mechanism of the force response to stretch in human skinned muscle fibres with different myosin isoforms

Force enhancement during lengthening of an active muscle, a condition that normally occurs during locomotion in vivo , is attributed to recruitment of myosin heads that exhibit fast attachment to and detachment from actin in a cycle that does not imply ATP splitting. We investigated the kinetic and mechanical features of this cycle in Ca²⁺ activated single skinned fibres from human skeletal muscles containing different myosin heavy chain (MHC) isoforms, identified with single-fibre gel electrophoresis. Fibres were activated by using a new set-up that allows development of most of the tension following a temperature jump from 0–1°C to the test temperature (∼12°C). In this way we could prevent the development of sarcomere non-uniformity and record sarcomere length changes with a striation follower in any phase of the mechanical protocol. We found that: (i) fibres with fast MHC isoforms develop 40–70% larger isometric forces than those with slow isoforms, as a result of both a larger fraction of force-generating myosin heads and a higher force per head; (ii) in both slow and fast fibres, force enhancement by stretch is due to recruitment of myosin head attachments, without increase in strain per head above the value generated by the isometric heads; and (iii) the extent of recruitment is larger in slow fibres than in fast fibres, so that the steady force and power output elicited by lengthening become similar, indicating that mechanical and kinetic properties of the actin–myosin interactions under stretch become independent of the MHC isoform.     

Inward-rectifying anion channels are expressed in the epithelial cells of choroid plexus isolated from ClC-2 'knock-out' mice

Choroid plexus epithelial cells express inward-rectifying anion channels which have a high HCO₃⁻ permeability. These channels are thought to have an important role in the secretion of cerebrospinal fluid. The possible relationship between these channels and the ClC-2  Cl⁻  channel was investigated in the present study. RT-PCR, using specific ClC-2 primers, amplified a 238 bp fragment of mRNA from rat choroid plexus, which was 99 % identical to the 5' sequence of rat ClC-2. A 2005 bp clone was isolated from a rat choroid plexus cDNA library using a probe for ClC-2. The clone showed greater than 99 % identity with the sequence of rat ClC-2. Inward-rectifying anion channels were observed in whole-cell recordings of choroid plexus epithelial cells isolated from ClC-2 knock-out mice. The mean inward conductance was 19.6 ± 3.6 nS (   n = 8  ) in controls (3 heterozygote animals), and 22.5 ± 3.1 nS (   n = 10  ) in three knock-out animals. The relative permeability of the conductances to I⁻ and  Cl⁻  ( P _I : P _Cl) was determined. I⁻ was more permeant than  Cl⁻  in both heterozygotes ( P _I: P _Cl= 4.0 ± 0.9, n = 3) and knock-out animals ( P _I : P _Cl= 4.1 ± 1.4, n = 3). These results indicate that rat choroid plexus expresses the ClC-2 variant that was originally reported in other tissues. ClC-2 does not contribute significantly to inward-rectifying anion conductance in mouse choroid plexus, which must therefore express a novel inward-rectifying anion channel.     

Nitric oxide plays a role in the regulation of adrenal blood flow and adrenocorticomedullary functions in the llama fetus

The effect of temperature on isometric tension with and without the regulatory proteins tropomyosin and troponin was studied in bovine myocardium using a thin filament removal and reconstitution protocol. In control bovine myocardium, isometric tension increased linearly with temperature in the range 5–40 °C: isometric tension at 10 and 30 °C was 0.65 and 1.28 times that at 20 °C, respectively, with a Q ₁₀ of about 1.4. In actin filament-reconstituted myocardium without regulatory proteins, the temperature effect on isometric tension was less: isometric tension at 10 and 30 °C was 0.96 and 1.17 times that at 20 °C, respectively, with a Q ₁₀ of about 1.1. The temperature dependence of the apparent rate constants was studied using sinusoidal analysis. The temperature dependence of 2π b (rate constant of delayed tension phase) did not vary significantly with the regulatory proteins under the standard activating condition (5 m m MgATP, 8 m m P_i, 200 m m ionic strength, pCa 4.66, pH 7.00). Q ₁₀ for 2π b in control and actin filament-reconstituted myocardium was 3.8 and 4.0, respectively. There were two phases to the temperature dependence of 2π c (rate constant of quick recovery). In control and thin filament-reconstituted myocardium, Q ₁₀ for 2π c was approximately 5.5 in the low temperature range (≤ 25 °C) and 2.7 in the high temperature range (≥ 30 °C). In actin filament-reconstituted myocardium, Q ₁₀ for 2π c was 8.5 in the low temperature range and 3.6 in the high temperature range. The above results demonstrate that regulatory proteins augment the temperature dependence of isometric tension, indicating that the regulatory proteins may modify the actomyosin interaction.     

Structural changes in myosin motors and filaments during relaxation of skeletal muscle

Structural changes in myosin motors and filaments during relaxation from short tetanic contractions of intact single fibres of frog tibialis anterior muscles at sarcomere length 2.14 μm, 4°C were investigated by X-ray diffraction. Force declined at a steady rate for several hundred milliseconds after the last stimulus, while sarcomere lengths remained almost constant. During this isometric phase of relaxation the intensities of the equatorial and meridional M3 X-ray reflections associated with the radial and axial distributions of myosin motors also recovered at a steady rate towards their resting values, consistent with progressive net detachment of myosin motors from actin filaments. Stiffness measurements confirmed that the fraction of motors attached to actin declined at a constant rate, but also revealed a progressive increase in force per motor. The interference fine structure of the M3 reflection suggested that actin-attached myosin motors are displaced towards the start of their working stroke during isometric relaxation. There was negligible recovery of the intensities of the meridional and layer-line reflections associated with the quasi-helical distribution of myosin motors in resting muscle during isometric relaxation, and the 1.5% increase in the axial periodicity of the myosin filament associated with muscle activation was not reversed. When force had decreased to roughly half its tetanus plateau value, the isometric phase of relaxation abruptly ended, and the ensuing chaotic relaxation had an exponential half-time of ca 60 ms. Recovery of the equatorial X-ray intensities was largely complete during chaotic relaxation, but the other X-ray signals recovered more slowly than force.     

Independent vasomotor control of rat tail and proximal hairy skin

Quantitative differences are known to exist between the vasomotor control of hairy and hairless skin, but it is unknown whether they are regulated by common central mechanisms. We made simultaneous recordings from sympathetic cutaneous vasoconstrictor (CVC-type) fibres supplying back skin (hairy) and tail (hairless) in urethane-anaesthetized, artificially ventilated rats. The animal's trunk was shaved and encased in a water-perfused jacket. Both tail and back skin CVC-type fibres were activated by cooling the trunk skin, and independently by the resultant fall in core (rectal) temperature, but their thresholds for activation differed (skin temperatures 38.8 ± 0.4°C versus 36.8 ± 0.4°C, core temperatures 38.1 ± 0.2°C versus 36.8 ± 0.2°C, respectively; P < 0.01). Back skin CVC-type fibres were more responsive to skin than to core cooling, while the reverse applied to tail fibres. Back skin CVC-type fibres were less responsive than tail fibres to prostaglandin E₂ (PGE₂) microinjected into the preoptic area. Spectral analysis showed no significant coherence between tail and back skin CVC-type fibre activities during cooling. After preoptic PGE₂ injection, a coherent peak at 1 Hz appeared in some animals; this disappeared after partialization with respect to ventilatory pressure, indicating that it was attributable to common ventilatory modulation. Neuronal inhibition in the rostral medullary raphé by microinjected muscimol (2 m m , 60–120 nl) suppressed both tail and back skin CVC-type fibre activities, and prevented their responses to subsequent skin cooling. These results indicate that thermoregulatory responses of hairless and hairy skin vessels are controlled by independent neural pathways, although both depend on synaptic relays in the medullary raphé.     

Tetrahydrobiopterin augments endothelium-dependent dilatation in sedentary but not in habitually exercising older adults

Endothelium-dependent dilatation (EDD) is impaired with ageing in sedentary, but not in regularly exercising adults. We tested the hypotheses that differences in tetrahydrobiopterin (BH₄) bioactivity are key mechanisms explaining the impairment in EDD with sedentary ageing, and the maintenance of EDD with ageing in regularly exercising adults. Brachial artery flow-mediated dilatation (FMD), normalized for local shear stress, was measured after acute oral placebo or BH₄ in young sedentary (YS) ( n = 10; 22 ± 1 years, mean ± s.e.m. ), older sedentary (OS) ( n = 9; 62 ± 2), and older habitually aerobically trained (OT) ( n = 12; 66 ± 1) healthy men. At baseline, FMD was ∼50% lower in OS versus YS (1.12 ± 0.09 versus 0.57 ± 0.09 (Δmm (dyn cm⁻²)) × 10⁻², P < 0.001; 1 dyn = 10⁻⁵ N), but was preserved in OT (0.93 ± 0.08 (Δmm (dyn cm⁻²)) × 10⁻²). BH₄ administration improved FMD by ∼45% in OS (1.00 ± 0.10 (Δmm (dyn cm⁻²)) × 10⁻², P < 0.01 versus baseline), but did not affect FMD in YS or OT. Endothelium-independent dilatation neither differed between groups at baseline nor changed with BH₄ administration. These results suggest that BH₄ bioactivity may be a key mechanism involved in the impairment of conduit artery EDD with sedentary ageing, and the EDD-preserving effect of habitual exercise.     

腰椎间盘突出患者腰椎CT三维重建影像的解剖学测量及其在单边双通道内镜手术中的临床意义

目的 探讨腰椎间盘突出症患者腰椎CT三维重建影像解剖学特点及其在单边双通道内镜手术中的意义。方法 横断面调查研究。纳入安徽医科大学第二附属医院骨科2019年1月—2020年6月收治的腰椎间盘突出症的患者50例,其中男32例、女18例,年龄23~69（64.1±13.4）岁。病变节段：L_2/3 1例,L_4/5 18例,L₅/S₁ 23例,L_3/4合并L_4/5 1例,L_4/5合并L₅/S₁ 6例。均采用单边双通道内镜辅助下腰椎间盘突出髓核摘除术治疗。术前患者均行腰椎CT扫描及三维容积重建,观察患者腰椎整体形态结构,使用放射工作站软件测量两侧椎板夹角、椎板宽度、关节突关节矢状化程度以及椎板下移程度,并进行腰椎节段间及侧别间比较。结果 （1）棘突与两侧椎板呈“人”字形平滑连接,两侧椎板向两侧弧形展开。两侧椎板夹角自L₃至L₅逐渐增大,分别为70°±11°、78°±12°和92°±13°,差异有统计学意义（F=42.25,P<0.001）。（2）椎板下缘向两侧延伸,椎板宽度自L₃至L₅逐渐增宽,左侧分别为（14.0±2.4）、（15.7±2.6）和（18.7±2.8）mm,右侧分别为（14.4±2.4）、（16.4±2.3）和（19.2±2.8）mm,各节段椎板宽度右侧均大于左侧,差异均有统计学意义（t=2.13、4.00、2.78,P值均<0.05）。（3）椎板末端逐渐增宽膨大成关节面,关节面L_3/4~L₅/S₁逐渐由矢状化转变为冠状化,左侧分别为60°±10°、50°±9°和42°±11°,右侧分别为58°±11°、48°±10°和40°±13°。同一节段左右两侧对比,在L_3/4差异有统计学意义（t=2.92,P=0.005）,在L_4/5及L₅/S₁差异均无统计学意义（t=1.29、1.79,P值均>0.05）。（4）椎板下移程度自L₃至L₅逐渐减小,分别为（6.8±2.1）、（5.1±2.3）和（2.4±2.3）mm,差异有统计学意义（F=50.39,P<0.001）。结论 腰椎间盘突出症患者从L₃到L₅,两侧椎板夹角逐渐增大、椎板宽度逐渐增大、关节突关节逐渐由矢状化转变为冠状化、椎板下移程度逐渐减小。了解腰椎解剖的节段性变化有助于单边双通道内镜手术的顺利进行、椎板去除范围的确定、对关节突关节的保护以及椎间盘位置的确定。     

A reduced cerebral metabolic ratio in exercise reflects metabolism and not accumulation of lactate within the human brain

During maximal exercise lactate taken up by the human brain contributes to reduce the cerebral metabolic ratio, O₂/(glucose + 1/2 lactate), but it is not known whether the lactate is metabolized or if it accumulates in a distribution volume. In one experiment the cerebral arterio-venous differences (AV) for O₂, glucose (glc) and lactate (lac) were evaluated in nine healthy subjects at rest and during and after exercise to exhaustion. The cerebrospinal fluid (CSF) was drained through a lumbar puncture immediately after exercise, while control values were obtained from six other healthy young subjects. In a second experiment magnetic resonance spectroscopy (¹H-MRS) was performed after exhaustive exercise to assess lactate levels in the brain ( n  = 5). Exercise increased the AV_O2 from 3.2 ± 0.1 at rest to 3.5 ± 0.2 m m (mean ± s.e.m. ; P < 0.05) and the AV_glc from 0.6 ± 0.0 to 0.9 ± 0.1 m m ( P < 0.01). Notably, the AV_lac increased from 0.0 ± 0.0 to 1.3 ± 0.2 m m at the point of exhaustion ( P < 0.01). Thus, maximal exercise reduced the cerebral metabolic ratio from 6.0 ± 0.3 to 2.8 ± 0.2 ( P < 0.05) and it remained low during the early recovery. Despite this, the CSF concentration of lactate postexercise (1.2 ± 0.1 m m ; n = 7) was not different from baseline (1.4 ± 0.1 m m ; n = 6). Also, the ¹H-MRS signal from lactate obtained after exercise was smaller than the estimated detection limit of ∼1.5 m m . The finding that an increase in lactate could not be detected in the CSF or within the brain rules out accumulation in a distribution volume and indicates that the lactate taken up by the brain is metabolized.     

Interstitial exclusion of albumin in rabbit lung during development of pulmonary oedema

The modifications of the macromolecular sieving properties of the pulmonary extracellular tissue matrix were studied in adult anaesthetized rabbits ( n = 10) exposed to increased tissue hydration. Exclusion of albumin from the perivascular pulmonary interstitial space was determined by using the continuous infusion method coupled with direct sampling of interstitial fluid performed through the wick technique. The rabbits underwent an intravenous infusion of saline amounting to 10 ( n = 5) or 20 % ( n = 5) body weight. Extracellular albumin distribution volume was derived from the steady state tissue concentration of radioactive rabbit serum albumin (¹²⁵I-RSA). Pulmonary extracellular and intravascular fluid volumes ( V _x and V _v, respectively) were measured as distribution volumes of ⁵¹Cr-EDTA and ¹³¹I-RSA, respectively, and interstitial fluid tracer concentrations were determined in interstitial fluid collected through implanted wicks. At the highest degree of hydration, interstitial fluid volume ( V _i= V _x− V _v) and extravascular albumin distribution volume ( V _a,w) significantly increased by 38.5 and 240.2 %, respectively, compared to control. Albumin-excluded volume ( V _e,a= V _i− V _a,w) was 398.9 ± 17 μl (g dry tissue weight)⁻¹; the albumin-excluded volume fraction ( F _e,a= V _e,a/ V _i) was 0.23 ± 0.01, 33.2 % of the control value. Data indicate that, at variance with what is observed in tissues like skin and muscle, pulmonary F _e,a is highly sensitive to tissue fluid content.     

Exercise Increases Ca2+–Calmodulin-Dependent Protein Kinase II Activity in Human Skeletal Muscle

There is evidence in rodents that Ca²⁺-calmodulin-dependent protein kinase II (CaMKII) activity is higher in contracting skeletal muscle, and this kinase may regulate skeletal muscle function and metabolism during exercise. To investigate the effect of exercise on CaMKII in human skeletal muscle, healthy men (   n = 8  ) performed cycle ergometer exercise for 40 min at 76 ± 1 % peak pulmonary O₂ uptake (_O2peak), with skeletal muscle samples taken at rest and after 5 and 40 min of exercise. CaMKII expression and activities were examined by immunoblotting and in vitro kinase assays, respectively. There were no differences in maximal (+ Ca²⁺, CaM) CaMKII activity during exercise compared with rest. Autonomous (- Ca²⁺, CaM) CaMKII activity was 9 ± 1 % of maximal at rest, remained unchanged at 5 min, and increased to 17 ± 1 % (   P < 0.01  ) at 40 min. CaMKII autophosphorylation at Thr²⁸⁷ was 50-70 % higher during exercise, with no differences in CaMKII expression. The effect of maximal aerobic exercise on CaMKII was also examined (   n = 9  ), with 0.7- to 1.5-fold increases in autonomous CaMKII activity, but no change in maximal CaMKII activity. CaMKIV was not detected in human skeletal muscle. In summary, exercise increases the activity of CaMKII in skeletal muscle, suggesting that it may have a role in regulating skeletal muscle function and metabolism during exercise in humans.