Method to Measure Tone of Axial and Proximal Muscle

The control of tonic muscular activity remains poorly understood. While abnormal tone is commonly assessed clinically by measuring the passive resistance of relaxed limbs¹, no systems are available to study tonic muscle control in a natural, active state of antigravity support. We have developed a device (Twister) to study tonic regulation of axial and proximal muscles during active postural maintenance (i.e. postural tone). Twister rotates axial body regions relative to each other about the vertical axis during stance, so as to twist the neck, trunk or hip regions. This twisting imposes length changes on axial muscles without changing the body''s relationship to gravity. Because Twister does not provide postural support, tone must be regulated to counteract gravitational torques. We quantify this tonic regulation by the restive torque to twisting, which reflects the state of all muscles undergoing length changes, as well as by electromyography of relevant muscles. Because tone is characterized by long-lasting low-level muscle activity, tonic control is studied with slow movements that produce "tonic" changes in muscle length, without evoking fast "phasic" responses. Twister can be reconfigured to study various aspects of muscle tone, such as co-contraction, tonic modulation to postural changes, tonic interactions across body segments, as well as perceptual thresholds to slow axial rotation. Twister can also be used to provide a quantitative measurement of the effects of disease on axial and proximal postural tone and assess the efficacy of intervention.     

Axial skeletal and Hox expression domain alterations induced by retinoic acid, valproic acid, and bromoxynil during murine development

Retinoic acid (RA) alters the developmental fate of the axial skeletal anlagen. "Anteriorizations" or "posteriorizations," the assumption of characteristics of embryonic areas normally anterior or posterior to the affected tissues, are correlated with altered embryonal expression domains of Hox genes after in utero RA treatment. These "homeotic" changes have been hypothesized to result from alterations of a "Hox cod" which imparts positional identity in the axial skeleton. To investigate whether such developmental alterations were specific to RA, or were a more general response to xenobiotic exposure, CD-1 pregnant mice were exposed to RA, valproic acid (VA), or bromoxynil (Br) during organogenesis. Additionally, the expression domains of two Hox genes, Hoxa7 and Hoxa10, were examined in gestation day (GD) 12.5 embryos obtained from control, RA, VA, or Br, treated gravid dams exposed on GD 6, 7, or 8. The anterior expression boundary of Hoxa7 is at the level of the C7/T1 vertebrae and that of Hoxa10 is at L6/S1. Compound-induced changes in the incidence of skeletal variants were observed. These included supernumerary cervical ribs (CSNR) lateral to C7, 8 vertebrosternal ribs, supernumerary lumbar ribs (LSNR) lateral to L1, extra presacral vertebrae, and the induction of vertebral and/or rib malformations. RA and VA administration on GD 6 caused posteriorization in the cervico-thoracic region (CSNR) while GD 8 exposure to any of the three compounds resulted in anteriorizations in the thoraco-lumbar area (LSNR and an increase in the number of presacral vertebrae). These effects occurred across regions of the axial skeleton. Analysis of gene expression demonstrated changes in the anterior boundaries of Hoxa7 expression domains in embryos treated on GD 6 and 8 with RA. VA and Br did not induce any statistically significant alterations in Hoxa7 and none of the compounds caused alterations in Hoxa10 expression domains. The studies indicate that RA GD 6 treatment-induced Hoxa7 shifts were rostral (posteriorization) while the RA-induced GD 8 anterior expression boundary shift was caudal (anteriorization), correlating with the axial skeletal changes noted. These data suggest that xenobiotic compounds such as VA and Br may induce similar axial skeletal changes by affecting different components of the developmental processes involved in the patterning of the axial skeleton.     

Ophthalmologic changes related to radiation exposure and age in adult health study sample, Hiroshima and Nagasaki

A 2-year ophthalmologic study of age and radiation-related ophthalmologic lesions among the atomic bomb survivors in Hiroshima and Nagasaki was conducted in 1978-80. The study sample in both cities was composed of all persons exposed to 100+ rad, their controls, and all other persons with a previous record of axial opacities or posterior subcapsular changes. Most of the losses were due to persons who refused to participate or for whom it was not possible to arrange for an ophthalmologic examination at the time of the regularly scheduled medical examination. It should be emphasized, however, that the loss of persons in both the control and the 100+ rad groups did not change systematically with increasing age by city. Increased lenticular opacities, other lens changes, and loss of visual acuity and accommodation occurred with increasing age in both exposed and control subjects as manifestations of the normal aging process. A highly significant excess risk for all age categories in the 300+ rad group in comparison to those in the control group was observed for both axial opacities and posterior subcapsular changes in Hiroshima, but not in Nagasaki. A stronger radiosensitive aging effect for persons who were under 15 years old at the time of the bombing (ATB) was observed for both axial opacities and posterior subcapsular changes in Hiroshima.     

ClpP: a structurally dynamic protease regulated by AAA+ proteins

Proteolysis is an important process for many aspects of bacterial physiology. Clp proteases carry out a large proportion of protein degradation in bacteria. These enzymes assemble in complexes that combine the protease ClpP and the unfoldase, ClpA or ClpX. ClpP oligomerizes as two stacked heptameric rings enclosing a central chamber containing the proteolytic sites. ClpX and ClpA assemble into hexameric rings that bind both axial surfaces of the ClpP tetradecamer forming a barrel-like complex. ClpP requires association with ClpA or ClpX to unfold and thread protein substrates through the axial pore into the inner chamber where degradation occurs. A gating mechanism regulated by the ATPase exists at the entry of the ClpP axial pore and involves the N-terminal regions of the ClpP protomers. These gating motifs are located at the axial regions of the tetradecamer but in most crystal structures they are not visible. We also lack structural information about the ClpAP or ClpXP complexes. Therefore, the structural details of how the axial gate in ClpP is regulated by the ATPases are unknown. Here, we review our current understanding of the conformational changes that ClpA or ClpX induce in ClpP to open the axial gate and increase substrate accessibility into the degradation chamber. Most of this knowledge comes from the recent crystal structures of ClpP in complex with acyldepsipeptides (ADEP) antibiotics. These small molecules are providing new insights into the gating mechanism of this protease because they imitate the interaction of ClpA/ClpX with ClpP and activate its protease activity.     

Identification of apolipoprotein A-I as a "STOP" signal for myopia

Good visual acuity requires that the axial length of the ocular globe is matched to the refractive power of the cornea and lens to focus the images of distant objects onto the retina. During the growth of the juvenile eye, this is achieved through the emmetropization process that adjusts the ocular axial length to compensate for the refractive changes that occur in the anterior segment. A failure of the emmetropization process can result in either excessive or insufficient axial growth, leading to myopia or hyperopia, respectively. Emmetropization is mainly regulated by the retina, which generates two opposite signals: "GO/GROW" signals to increase axial growth and "STOP" signals to block it. The presence of GO/GROW and STOP signals was investigated by a proteomics analysis of the retinas from chicken with experimental myopia and hyperopia. Of 18 differentially expressed proteins that were identified, five displayed an expression profile corresponding to GO/GROW signals, and two corresponded to STOP signals. Western blotting confirmed that apolipoprotein A-I (apoA-I) has the characteristics of a STOP signal both in the retina as well as in the fibrous sclera. In accordance with this, intraocular application of the peroxisome proliferator-activated receptor alpha agonist GW7647 resulted in up-regulation of apoA-I levels and in a significant reduction of experimental myopia. In conclusion, using a comprehensive functional proteomics analysis of chicken ocular growth models we identified targets for ocular growth control. The correlation of elevated apoA-I levels with reduced ocular axial growth points toward a functional relationship with the observed morphological changes of the eye.     

LiCl disrupts axial development in mouse but does not act through the beta-catenin/Lef-1 pathway

Chimera and cell marking studies suggest that axial determination in mouse embryos occurs at postimplantation stages. In contrast, Xenopus laevis axes are determined early due to the asymmetric distribution of maternally derived factors in the one-cell zygote. In our earlier study we used lithium chloride (LiCl) to perturb development of mouse axes. Here we investigate whether the lithium induced axial defects in mouse are being mediated by the beta-catenin/Lef-1 pathway as in Xenopus laevis. In lithium treated embryos we did not observe any changes in the amount or localization of beta-catenin protein. Furthermore, the lack of Lef-1 mRNA in treated and untreated embryos indicates the LiCl induced axial defects in the mouse are not mediated by the beta-catenin/Lef-1 pathway.     

Microfibril orientation in differentiating and maturing fibre and parenchyma cell walls in culms of bamboo (Phyllostachys viridi-glaucescens (Garr.) Riv. & Riv.)

Results of trials using chemical and enzymatic wall extractants for the removal of matrix materials for in situ observations of newly deposited microfibrils are described. Observations were then made of the orientation of microfibrils on the inner walls of differentiating and maturing fibres and parenchyma cells under the FESEM. Orientation changes were similar in both cell types. During very early primary wall development, deposition of microfibrils was in more or less axial alignment, which was later superseded by microfibrils in transverse orientation (90^o to the long axis). A transverse orientation of microfibrils remained throughout much of primary wall synthesis, until an abrupt shift occurred to a sloped orientation during late primary wall synthesis. Microfibrils of the first secondary wall layer were in axial alignment or steeply sloped. In subsequent secondary wall deposition there was an alternation between a transverse and a sloped or axial alignment in maturing fibres and parenchyma cells.     

Structural changes of actin-bound myosin heads after a quick length change in frog skeletal muscle

Changes in the x-ray diffraction pattern from a frog skeletal muscle were recorded after a quick release or stretch, which was completed within one millisecond, at a time resolution of 0.53 ms using the high-flux beamline at the SPring-8 third-generation synchrotron radiation facility. Reversibility of the effects of the length changes was checked by quickly restoring the muscle length. Intensities of seven reflections were measured. A large, instantaneous intensity drop of a layer line at an axial spacing of 1/10.3 nm(-1) after a quick release and stretch, and its partial recovery by reversal of the length change, indicate a conformational change of myosin heads that are attached to actin. Intensity changes on the 14.5-nm myosin layer line suggest that the attached heads alter their radial mass distribution upon filament sliding. Intensity changes of the myosin reflections at 1/21.5 and 1/7.2 nm(-1) are not readily explained by a simple axial swing of cross-bridges. Intensity changes of the actin-based layer lines at 1/36 and 1/5.9 nm(-1) are not explained by it either, suggesting a structural change in actin molecules.     

NMR study on the structural changes of cytochrome P450cam upon the complex formation with putidaredoxin. Functional significance of the putidaredoxin-induced structural changes

We investigated putidaredoxin-induced structural changes in carbonmonoxy P450cam by using NMR spectroscopy. The resonance from the beta-proton of the axial cysteine was upfield shifted by 0.12 ppm upon the putidaredoxin binding, indicating that the axial cysteine approaches to the heme-iron by about 0.1 A. The approach of the axial cysteine to the heme-iron would enhance the electronic donation from the axial thiolate to the heme-iron, resulting in the enhanced heterolysis of the dioxygen bond. In addition to the structural perturbation on the axial ligand, the structural changes in the substrate and ligand binding site were observed. The resonances from the 5-exo- and 9-methyl-protons of d-camphor, which were newly identified in this study, were upfield shifted by 1.28 and 0.20 ppm, respectively, implying that d-camphor moves to the heme-iron by 0.15-0.7 A. Based on the radical rebound mechanism, the approach of d-camphor to the heme-iron could promote the oxygen transfer reaction. On the other hand, the downfield shift of the resonance from the gamma-methyl group of Thr-252 reflects the movement of the side chain away from the heme-iron by approximately 0.25 A. Because Thr-252 regulates the heterolysis of the dioxygen bond, the positional rearrangement of Thr-252 might assist the scission of the dioxygen bond. We, therefore, conclude that putidaredoxin induces the specific heme environmental changes of P450cam, which would facilitate the oxygen activation and the oxygen transfer reaction.     

Determination of molecular changes in soft tissues under strain using laser Raman microscopy

The paper presents a non-contact technique to examine the molecular changes in a collagen fibre subjected to in vitro axial tension. Laser Raman microscopy was employed to monitor the vibrational changes in specific assignments of the Raman spectrum of collagen. Results were presented in the form of Raman wavenumber shift as a function of applied tensile strain. Two distinct responses were observed depending on whether the vibrations were axial to, or normal to, the collagen backbone. The former response produced a decrease in wavenumber values, indicating tension, whereas the latter produced an increase, indicating compression. The rate of wavenumber shift with applied strain was non-linear in form, with a marked increase at higher levels of applied strain, for example, a strain 4% in the case of axial vibrations. This technique can prove to be a powerful tool for examining deformation at the molecular level in collagenous tissues.     

Fundamental role of axial stress in compensatory adaptations by arteries

Arteries exhibit a remarkable ability to adapt to diverse genetic defects and sustained alterations in mechanical loading. For example, changes in blood flow induced wall shear stress tend to control arterial caliber and changes in blood pressure induced circumferential wall stress tend to control wall thickness. We submit, however, that the axial component of wall stress plays a similarly fundamental role in controlling arterial geometry, structure, and function, that is, compensatory adaptations. This observation comes from a review of findings reported in the literature and a comparison of four recent studies from our laboratory that quantified changes in the biaxial mechanical properties of mouse carotid arteries in cases of altered cell-matrix interactions, extracellular matrix composition, blood pressure, or axial extension. There is, therefore, a pressing need to include the fundamental role of axial wall stress in conceptual and theoretical models of arterial growth and remodeling and, consequently, there is a need for increased attention to evolving biaxial mechanical properties in cases of altered genetics and mechanical stimuli.     

Measuring,reversing, and modeling the mechanical changes due to the absence of Fibulin-4 in mouse arteries

Mice with a smooth muscle cell (SMC)-specific deletion of Fibulin-4 (SMKO) show decreased expression of SMC contractile genes, decreased circumferential compliance, and develop aneurysms in the ascending aorta. Neonatal administration of drugs that inhibit the angiotensin II pathway encourages the expression of contractile genes and prevents aneurysm development, but does not increase compliance in SMKO aorta. We hypothesized that multidimensional mechanical changes in the aorta and/or other elastic arteries may contribute to aneurysm pathophysiology. We found that the SMKO ascending aorta and carotid artery showed mechanical changes in the axial direction. These changes were not reversed by angiotensin II inhibitors, hence reversing the axial changes is not required for aneurysm prevention. Mechanical changes in the circumferential direction were specific to the ascending aorta; therefore, mechanical changes in the carotid do not contribute to aortic aneurysm development. We also hypothesized that a published model of postnatal aortic growth and remodeling could be used to investigate mechanisms behind the changes in SMKO aorta and aneurysm development over time. Dimensions and mechanical behavior of adult SMKO aorta were reproduced by the model after modifying the initial component material constants and the aortic dilation with each postnatal time step. The model links biological observations to specific mechanical responses in aneurysm development and treatment.     

Lattice spacing changes accompanying isometric tension development in intact single muscle fibers.

The myosin lattice spacing of single intact muscle fibers of the frog, Rana temporaria, was studied in Ringer's solution (standard osmolarity 230 mOsm) and hyper- and hypotonic salines (1.4 and 0.8 times standard osmolarity respectively) in the relaxed state, during "fixed end" tetani, and during shortening, using synchrotron radiation. At standard tonicity, a tetanus was associated with an initial brief lattice expansion (and a small amount of sarcomere shortening), followed by a slow compression (unaccompanied by sarcomere length changes). In hypertonic saline (myosin lattice compressed by 8.1%), these spacing changes were suppressed, in hypotonic saline (lattice spacing increased by 7.5%), they were enhanced. During unloaded shortening of activated fibers, a rapid lattice expansion occurred at all tonicities, but became larger as tonicity was reduced. This expansion was caused in part by the change in length of the preparation, but also by a recoil of a stressed radial compliance associated with axial force. The lattice spacing during unloaded shortening was equal to or occasionally greater than predicted for a relaxed fiber at that sarcomere length, indicating that the lattice compression associated with activation is rapidly reversed upon loss of axial force. Lattice recompression occurred upon termination of shortening under standard and hypotonic conditions, but was almost absent under hypertonic conditions. These observations indicate that axial cross-bridge tension is associated with a compressive radial force in intact muscle fibers at full overlap; however, this radial force exhibits a much greater sensitivity to lattice spacing than does the axial force.     

Axial flow velocity patterns in a pulmonary artery model with varying degrees of valvular pulmonic stenosis: pulsatile in vitro studies

With the advent of noninvasive clinical techniques which can measure blood flow velocities (Doppler ultrasound), it is suggested that a fundamental knowledge of the axial flow velocity patterns in the pulmonary artery, and the changes caused by stenosis, may be used to support accurate diagnosis of valvular pulmonic stenosis. The present study was designed to characterize the axial flow velocity patterns in an in vitro model of a human adult pulmonary artery with varying degrees of valvular pulmonic stenosis. A two-dimensional laser Doppler anemometer (LDA) system was used to map the flow fields in the main (MPA), left (LPA), and right (RPA) branches of the pulmonary artery model. The study was conducted in the Georgia Tech. right heart pulse duplicator system. It was observed that the axial flow velocity patterns in the MPA and the LPA change dramatically with increasing degree of valvular stenosis. This indicates that the axial flow velocity patterns in these two branches are strongly influenced by the degree of valvular stenosis. The axial flow velocity patterns in the RPA, however, do not change much with varying degrees of valvular stenosis, indicating that the axial flow fields in the RPA are mainly influenced by the geometry of the bifurcation. It may be concluded therefore, that the changes in the axial flow velocity patterns in the MPA and LPA (rather than in the RPA) could be sensitive and reliable indicators of the severity of the defect.     

Estimates of ventilation from body surface measurements in unrestrained subjects

To make estimates of ventilation from measurements of body surface movements in unrestrained subjects, we measured changes in linear dimensions and cross-sectional areas of the rib cage (RC) and abdomen (AB) of six healthy unrestrained subjects during a variety of maneuvers. RC and AB anteroposterior diameters and abdominal length in the cephalocaudal axis (axial displacement) were measured with magnetometers, and RC and AB cross-sectional areas were measured with a respiratory inductance plethysmograph. Flow was measured at the mouth with a pneumotachograph and integrated electrically to give volume. Volume and body surface measurements were analyzed by multiple linear regression. Addition of the axial measurements to either the anteroposterior dimensions or cross-sectional areas of RC and AB improved estimates of tidal volume in all subjects (P less than 0.01). With measurements of axial displacement and cross-sectional area of the RC and AB, tidal volume could be reliably estimated to within 20% of actual ventilation. We conclude that measurement of axial displacements improves estimates of ventilation in unrestrained subjects.     

Living cells in wood. 1. Absence,scarcity and histology of axial parenchyma as keys to function

The diversity of expression in axial parenchyma (or lack of it) in woods is reviewed and synthesized with recent work in wood physiology, and questions and hypotheses relative to axial parenchyma anatomy are offered. Cell shape, location, abundance, size, wall characteristics and contents are all characteristics for the assessment of the physiological functions of axial parenchyma, a tissue that has been neglected in the consideration of how wood histology has evolved. Axial parenchyma occurrence should be considered with respect to mechanisms for the prevention and reversal of embolisms in tracheary elements. This mechanism complements cohesion–tension‐based water movement and root pressure as a way of maintaining flow in xylem. Septate fibres can substitute for axial parenchyma (‘axial parenchyma absent’) and account for water movement in xylem and for the supply of carbohydrate abundance underlying massive and sudden events of foliation, flowering and fruiting, as can fibre dimorphism and the co‐occurrence of septate fibres and axial parenchyma. Rayless woods may or may not contain axial parenchyma and are informative when analysing parenchyma function. Interconnections between ray and axial parenchyma are common, and so axial and radial parenchyma must be considered as complementary parts of a network, with distinctive but interactive functions. Upright ray cells and more numerous rays per millimetre enhance interconnection and are more often found in woods that contain tracheids. Vesselless woods in both gymnosperms and angiosperms have axial parenchyma, the distribution of which suggests a function in osmotic water shifting. Water and photosynthate storage in axial parenchyma may be associated with seasonal changes and with succulent or subsucculent modes of construction. Apotracheal axial parenchyma distribution often demonstrates storage functions that can be read independently of osmotic water shifting capabilities. Axial parenchyma may serve to both enhance mechanical strength or, when parenchyma is thin‐walled, as a tissue that adapts to volume change with a change in water content. Other functions of axial parenchyma (contributing resistance to pathogens; a site for the recovery of physical damage) are considered. The diagnostic features of axial parenchyma and septate fibres are reviewed in order to clarify distinctions and to aid in cell type identification. Systematic listings are given for particular axial parenchyma conditions (e.g. axial parenchyma ‘absent’ with septate fibres substituting). A knowledge of the axial parenchyma information presented here is desirable for a full understanding of xylem function. © 2015 The Linnean Society of London, Botanical Journal of the Linnean Society, 2015, 177 , 291–321.     

Nonuniform volume changes during muscle contraction.

We measured dynamic changes in volume during contraction of live, intact frog skeletal muscle fibers through a high-speed, intensified, digital-imaging microscope. Optical cross-sections along the axis of resting cells were scanned and compared with sections during the plateau of isometric tetanic contractions. Contraction caused an increase in volume of the central third of a cell when axial force was maximum and constant and the central segment was stationary or lengthened slightly. But changes were unequal along a cell and not predicted by a cell's resting area or shape (circularity). Rapid local adjustments in the cytoskeletal evidently keep forces in equilibrium during contraction of living skeletal muscle. These results also show that optical signals may be distorted by nonuniform volume changes during contraction.     

Early ontogeny of vascular cambium

Observations were made on structural changes of procambium and the subsequent appearance of cambium in the developing shoot. The procambium in early stages shows radial seriation of cells as a result of repeated tangential divisions. In tangential view the procambium has initially a rather homogeneous structure and later becomes organized into two distinct systems, one composed of long cells and the other of short cells. The latter cells are arranged tangentially in axial files and transversely in radial files. They show repeated transverse divisions. Some of the short cells elongate to intrude between neighboring cells. Therefore, long cells may be derived both from cells of homogeneous structure in the first stage and from elongation of some of the short cells in axial files. Long cells have mostly tapering end walls and elongate actively. In the subsequent stages, the frequency of transverse divisions of short cells in axial files decreases and these cells expand radially. Short cells in axial files are separated from one another vertically by the elongation of neighboring long cells which break up the axial files as seen in tangential view. The vascular meristem in this stage is believed to initiate the cambium. Eventually, short cells remain mostly single, or form files of two or three cells in height in tangential view. The observations are discussed in relation to the structure of the vascular meristem in other plants.