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PubMed1981

Isomyosin distribution in normal and pressure-overloaded rat ventricular myocardium. An immunohistochemical study.

We have used affinity-purified antibodies reacting with guinea pig soleus muscle and ventricular myosin heavy chains to analyze the distribution of specific isomyosin in the ventricular myocardium of normal and renal hypertensive rats. Immunofluorescent staining of cardiac tissue sections with the two antimyosins revealed striking variations in reactivity among ventricular muscle fibers, reactive fibers being more numerous in the left compared to the right ventricle and in subendocardial compared to subepicardial layers. The response of the ventricular myocardium changed during development: all fibers were stained in the newborn rat, whereas most fibers were unreactive in 1-month-old animals. The number of reactive fibers increased again in subsequent stages leading to a mixed pattern in adult animals. The normal mixed pattern of reactivity was transformed into a uniformly positive pattern in hypertensive rats 2 months after surgery. This complete transformation was observed in 20 out of 23 hypertensive animals examined. These findings indicate that the two antimyosins cross-react with a particular type of ventricular myosin heavy chain, whose distribution varies in different muscle cells and whose relative concentration changes during development and during cardiac hypertrophy induced by systemic hypertension. We suggest that differences in pressure load may be responsible for both regional variations in isomyosin distribution and for isomyosin changes in hypertensive animals.

Animals↗
PubMed1994

Evaluation of a laser Doppler flowmetry implantable fiber system for determination of threshold thickness for flow detection in bone.

Laser Doppler flowmetry (LDF) with the use of a standard metal shafted probe, has been successfully used to study bone blood flow in a wide variety of settings. The use of the standard probe is limited by the requirement that the probe be replaced onto the bone surface or driven into the bone with a trocar. In response to this, a system of implantable, detachable fibers was developed. This system allows repeated measurement over time without repeated surgical manipulation of the area of interest. This study was performed to evaluate the implantable fiber system by determining threshold thickness for flow detection in bone. A flow chamber with perpendicular and end-on flow was designed to collect results under controlled conditions for flow detection. Threshold thickness for bovine cortical and cancellous bone samples was determined after sequentially grinding the specimens and placing them in the flow chamber. A 2% solution of latex circulated in the chamber and each specimen was exposed to both flow directions and both types of probe. The laser Doppler probe was able to detect flow by resting on top of the sample, with the latex on the other side of the sample. The results showed a significantly greater threshold thickness for the standard probes than for the implantable probes, and a significantly greater threshold thickness for trabecular bone than for cortical bone. Despite the reduced threshold thickness with the implantable fiber, this new system was able to consistently detect depth of perfusion of 80-90% of the values for the standard probe.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed2002

Method for analysis of 4-hydroxy-2-(E)-nonenal with solid-phase microextraction.

A simple analytical method for 4-hydroxy-2-(E)-nonenal (HNE) using solid-phase microextraction (SPME) fiber was developed. HNE or the derivative of HNE formed by reaction with 2,4-dinitrophenylhydrazine (DNPH) was extracted from the sample solution by immersing the SPME fiber into the solution, and the amount of HNE was quantified by HPLC. The extraction conditions of HNE and HNE-DNPH were examined, using standard solutions, with respect to fiber coating, NaCl concentration, rate of stirring, adsorption temperature, and adsorption time. The recovery of HNE reached 80%, and the quantification limits of HNE and HNE-DNPH using standard compounds were 14.1 pmol/10 mL and 486.5 fmol/10 mL, respectively. This method can be applied to the detection of HNE in oxidized oil or samples of porcine liver.

Aldehydes↗
PubMed1995

Cutaneous innervation in sensory neuropathies: evaluation by skin biopsy.

OBJECTIVE: To use punch skin biopsies to evaluate the loss of intra-epidermal nerve fibers in sensory neuropathies. BACKGROUND: Previous assessments of epidermal nerve fibers have been constrained by relatively insensitive staining techniques and variability in quantification. METHODS: Punch skin biopsies were performed on the heel and leg of HIV-seronegative controls, HIV-seropositive individuals without neuropathy, and patients with sensory neuropathies, including HIV-seronegative and HIV-positive individuals. After formalin fixation, 50-microns free-floating sections were stained with a monoclonal antibody to neuron-specific ubiquitin hydrolase, PGP9.5. The number of intraepidermal fibers/mm in at least three sections from each patient was counted by one observer blinded to site and clinical status. RESULTS: Dermal and epidermal nerve fibers were readily identified and quantified. The immunostaining technique reliably demonstrated a dermal plexus of myelinated and unmyelinated fibers parallel to the surface of the skin. In the epidermis, unmyelinated fibers ascended vertically between the keratinocytes to reach the stratum corneum. The number of intra-epidermal fibers/mm in the distal leg (mean +/- SEM) was 17.84 +/- 3.03 in seven HIV-seronegative controls. Epidermal fiber number was significantly reduced (p = 0.01) in five HIV-infected patients with sensory neuropathies associated with didanosine or zalcitabine therapy (1.07 +/- 0.40) and in eight HIV-seronegative patients with sensory neuropathies (3.1 +/- 3.1). Four of five neurologically normal HIV-seropositive subjects had reduced numbers of epidermal fibers, suggesting a subclinical neuropathy. Serial biopsies in one individual demonstrated the evolution of degenerating epidermal fibers after development of zalcitabine-induced sensory neuropathy. CONCLUSION: Skin biopsies stained with the sensitive panaxonal marker anti-PGP9.5 demonstrated significant reduction in intraepidermal fibers in sensory neuropathies. This simple and repeatable technique is a reliable method for quantitation of small cutaneous sensory fibers. In addition, skin biopsies may be useful in assessing the course and spatial distribution of involvement in peripheral nerve disease.

Adult↗
PubMed2000

Effects of hindlimb unloading on neuromuscular development of neonatal rats.

We hypothesized that hindlimb suspension unloading of 8-day-old neonatal rats would disrupt the normal development of muscle fiber types and the motor innervation of the antigravity (weightbearing) soleus muscles but not extensor digitorum longus (EDL) muscles. Five rats were suspended 4.5 h and returned 1.5 h to the dam for nursing on a 24 h cycle for 9 days. To control for isolation from the dam, the remaining five littermates were removed on the same schedule but not suspended. Another litter of 10 rats housed in the same room provided a vivarium control. Fibers were typed by myofibrillar ATPase histochemistry and immunostaining for embryonic, slow, fast IIA and fast IIB isomyosins. The percentage of multiple innervation and the complexity of singly-innervated motor terminal endings were assessed in silver/cholinesterase stained sections. Unique to the soleus, unloading accelerated production of fast IIA myosin, delayed expression of slow myosin and retarded increases in standardized muscle weight and fiber size. Loss of multiple innervation was not delayed. However, fewer than normal motor nerve endings achieved complexity. Suspended rats continued unloaded hindlimb movements. These findings suggest that motor neurons resolve multiple innervation through nerve impulse activity, whereas the postsynaptic element (muscle fiber) controls endplate size, which regulates motor terminal arborization. Unexpectedly, in the EDL of unloaded rats, transition from embryonic to fast myosin expression was retarded. Suspension-related foot drop, which stretches and chronically loads EDL, may have prevented fast fiber differentiation. These results demonstrate that neuromuscular development of both weightbearing and non-weightbearing muscles in rats is dependent upon and modulated by hindlimb loading.

Animals↗
PubMed1995

A non-enzymatic method for lens decapsulation which leaves the epithelial cells attached to the fibers.

We have developed a method to mechanically decapsulate the lens yet leave the epithelial cells attached to the fibers. This method uses divalent ion chelaters to loosen the capsule-epithelial interactions and bumetanide to control cell swelling. Light microscopy, scanning electron microscopy, and in-vitro fluorescence microscopy demonstrate that the capsule is removed and the epithelial cells remain adherent to the fibers when these procedures are used. Ion channel activity and epithelial gap junction communication remain following decapsulation. This decapsulated lens preparation should prove useful for many kinds of lens studies.

Animals↗
PubMed1988

Differential effects of d- and l-pimobendan on cardiac myofilament calcium sensitivity.

The effects is of the optical isomers of pimobendan (UD-CG 115 BS), an inotropic agent, were studied on the electrical and mechanical activity of intact and detergent-skinned preparations of cardiac muscle from guinea pig and dog. Racemic pimobendan has been shown to increase contractile force and to potentiate slow action potentials (AP) induced by stimulation of papillary muscle partially depolarized with 25 mM [K]o. These effects are shown in this study to be mainly due to the l-optical isomer of pimobendan. When slow APs were maximally stimulated by 1 microM isoproterenol, addition of either the d- or l-isomer of pimobendan did not affect the slow AP parameters. However, under these conditions, contractile force was significantly increased to 124% of control by the d-isomer and to 184% of control by the l-isomer. These results suggest that pimobendan may have direct effects on the myofilaments and that these effects are dependent on the optical isomer of the compound. To test this directly, the effects of d- and l-pimobendan were compared on Ca++-activated force developed by detergent-skinned heart muscle fibers. Submaximal force developed at constant Ca++ was increased by both optical isomers, but the l-isomer had a significantly greater Ca++-sensitizing effect. For example at pCa 6.75 force was 270% of control in the presence of the d-isomer and 400% of control in the presence of the l-isomer. At pCa 5, there was no effect of either isomer on force developed by the skinned fiber preparations.(ABSTRACT TRUNCATED AT 250 WORDS)

Actin Cytoskeleton↗
PubMed1994

Regenerating muscle fibers induce directional sprouting from nearby nerve terminals: studies in living mice.

The principal aim of this work was to better understand how regenerating muscle fibers become innervated in adult animals. To induce muscle regeneration, individual identified muscle fibers in a mouse were damaged with a laser focused through a microscope. The muscle fiber that degenerated and the muscle fiber that was formed in its place were followed by viewing the same site repeatedly over a period of 2 d to 40 weeks. Commonly, the nerve terminal innervating the irradiated muscle fiber partially retracted during muscle fiber degeneration, and then sprouted to innervate the regenerating muscle fiber at the same site it had previously innervated the muscle fiber that was damaged. During the early phase of muscle regeneration we also observed sprouts originating from nerve terminals on adjacent muscle fibers. The new nerve growth was a response to the regenerating muscle fiber rather than to the degenerated fiber it replaced because repeated damage of the same site every 2-3 d over a 10 d period (to prevent regeneration) did not cause any sprouting. The direction of the sprouts on adjacent muscle fibers showed a bias toward the regenerating muscle fiber, although they avoided the region occupied by the original nerve terminal. Forty percent of the sprouts managed to reach the regenerated fiber. Nonetheless, by 11 d after muscle fiber damage, all sprouts had regressed, leaving the new fiber innervated by the same motor axon that innervated the fiber that was damaged. On the other hand, when the overlying nerve terminal as well as the muscle fiber was damaged, the sprouts from nearby muscle fibers were both more numerous and more stable, and in five cases we observed two or more new synaptic junctions on the regenerating fiber originating from different axons. In one case we witnessed a protracted competition between the original motor axon as it sprouted back and the sprouts from nearby junctions for sole innervation of the regenerate. Ultimately, the surviving sprouts myelinated and became the permanent and exclusive input to the new fiber. These results indicate that regenerating muscle fibers emit a signal that induces directional sprouting from nearby undamaged nerve terminals. Reinnervation of the regenerating muscle fiber by one axon apparently prevents the maintenance of such neurites. Because the process of muscle regeneration shares many features in common with myogenesis during embryonic development, it is likely that developing muscle fibers present an analogous stimulus to ingrowing motor axons.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed1991

Coordination of skeletal muscle gene expression occurs late in mammalian development.

The acquisition of specialized skeletal muscle fiber phenotypes during development is investigated by systematic measurement of the accumulation of 21 contractile protein mRNAs during hindlimb development in the rat and the human. During early myotube formation in both species there is no coordination of expression of either fast or slow contractile protein isoform genes, but rather some slow, some fast, and some cardiac isoforms are expressed. Some isoforms are not detected at all in early myotubes. From Embryonic Day 19 in the rat, and after 14 weeks in the human, a strong bias toward fast isoform expression is evident for all gene families examined. This results in the establishment of a coordinated fast isoform phenotype at birth in the rat, and by 24 weeks in the human fetus. Unexpectedly, during secondary myotube formation in the rat we observe sudden rises and falls in contractile protein gene output. We interpret these fluctuations in terms of periods of myoblast proliferation followed by synchronized fusion into myotubes. The data presented indicate that each contractile protein gene has its own determinants of mRNA accumulation and that the different myoblast populations which contribute to the developing limb are not intrinsically programmed to produce particular coordinated phenotypes with respect to the non-myosin heavy chain contractile proteins.

Animals↗
PubMed1977

DNA degradation in terminally differentiating lens fiber cells from chick embryos.

During the terminal differentiation of lens fiber cells, nuclear DNA is known to accumulate free 3'-OH ends and is progressively lost from the nucleus. Toward the end of this process, nuclei undergo pycnosis and disappear. The size of the DNA in the epithelia and in early and late stages of fiber cell development was examined by electrophoresis on nondenaturing agarose/polyacrylamide gels. Low molecular weight DNA of discrete sizes appears only at the final stages of nuclear degeneration in central fiber cells and persists after the disappearance of the nuclei. These low molecular weight DNA fragments appear as multiples of a monomeric unit and are similar to the fragments produced by the digestion of epithelial cell nuclei by micrococcal nuclease. The data indicate that in lens fiber nuclei the double-strand breaks in vivo affect the chromatin during nuclear degeneration, and the data suggest that the DNA of these cells is organized into chromatin composed of discrete subunits.

Animals↗
PubMed2005

MRI-based finite-element analysis of left ventricular aneurysm.

Tagged MRI and finite-element (FE) analysis are valuable tools in analyzing cardiac mechanics. To determine systolic material parameters in three-dimensional stress-strain relationships, we used tagged MRI to validate FE models of left ventricular (LV) aneurysm. Five sheep underwent anteroapical myocardial infarction (25% of LV mass) and 22 wk later underwent tagged MRI. Asymmetric FE models of the LV were formed to in vivo geometry from MRI and included aneurysm material properties measured with biaxial stretching, LV pressure measurements, and myofiber helix angles measured with diffusion tensor MRI. Systolic material parameters were determined that enabled FE models to reproduce midwall, systolic myocardial strains from tagged MRI (630 +/- 187 strain comparisons/animal). When contractile stress equal to 40% of the myofiber stress was added transverse to the muscle fiber, myocardial strain agreement improved by 27% between FE model predictions and experimental measurements (RMS error decreased from 0.074 +/- 0.016 to 0.054 +/- 0.011, P < 0.05). In infarct border zone (BZ), end-systolic midwall stress was elevated in both fiber (24.2 +/- 2.7 to 29.9 +/- 2.4 kPa, P < 0.01) and cross-fiber (5.5 +/- 0.7 to 11.7 +/- 1.3 kPa, P = 0.02) directions relative to noninfarct regions. Contrary to previous hypotheses but consistent with biaxial stretching experiments, active cross-fiber stress development is an integral part of LV systole; FE analysis with only uniaxial contracting stress is insufficient. Stress calculations from these validated models show 24% increase in fiber stress and 115% increase in cross-fiber stress at the BZ relative to remote regions, which may contribute to LV remodeling.

Animals↗
PubMed1978

Behavioral and prolactin responses to 5-hydroxytryptophan in rats treated during development with 5,7-dihydroxytryptamine.

The serotonin precursor, 5-hydroxytryptophan (5-HTP), can induce a behavioral syndrome characterized by rigidity, splayed feet, tremor, head weaving, salivation and forepaw treading. This response to 5-HTP was markedly potentiated in adult rats treated intracisternally with 5,7-dihydroxytryptamine (5,7-DHT) during development. Prevention of the 5,7-DHT-induced reduction of brain norepinephrine with pargyline or desipramine did not diminish the potentiation of 5-HTP, suggesting that noradrenergic fibers are not contributing to the altered 5-HTP response. It was also found that treatments with 5,7-DHT potentiated the release of prolactin and the disruption of responding in a fixed-ratio operant task induced by 5-HTP. Other experiments indicated that 5,7-DHT treatments potentiated 5-HTP without affecting the action of L-dihydroxyphenylalanine. In addition, administration of the decarboxylase inhibitor, R0-4-4602, at a dose that inhibits enzyme activity in brain, blocked the 5-HTP-induced behavioral syndrome in 5,7-DHT-treated rats, indicating that 5-HTP must be converted to serotonin for 5-HTP to alter behavior. Thus, the present studies indicate that destruction of serotonergic fibers during development can produce permanent changes in central serotonergic mechanisms.

5,7-Dihydroxytryptamine↗
PubMed2006

Biopotential fiber sensor.

UNLABELLED: We have developed a new biopotential fiber sensor (BFS) technology as an alternative to traditional wet-gel Ag/AgCl electrodes in long-term monitoring applications. Biopotential fiber sensor technology uses proprietary method of bonding copper sulfide to the surface of acrylic fibers, thus creating an electrically conductive medium (10(-1) ohms/cm). Surface modified bundles of acrylic fibers form stable biopotential sensors when doped with proprietary ink comprising organic acids and nano particles of Ag and AgCl. Biopotential fiber sensors are characterized by a small footprint 0.1 mm2 and low mass of 0.005 g. Biopotential fiber sensor systems are disposable 1-piece assemblies combining the functions of the electrodes, lead wires and a patient cable. METHODS: The electric performance characteristics of BFS were obtained according to American National Standards Institute/Association for the Advancement of Medical Instrumentation EC-12:2000 standard and compared with Ag/AgCl wet-gel electrodes. The noise characteristics were determined from 250 hours of 3-lead electrocardiographic (ECG) data recorded with BFS over a period of 10 days from healthy volunteers. RESULTS: The BFS direct current offset voltage was 0.11 mV and alternate current impedance at 10 Hz was 22 ohms. The sensors average longevity on the body surface with no detachments was at least 7 days. BFS demonstrated high immunity to motion and electric field-induced artifacts. CONCLUSIONS: Biopotential fiber sensors are well suited for routine long-term ECG monitoring applications because of increased patient comfort through integration of the electrode, lead wire, and a patient cable, which function into one disposable BFS fiber assembly. Biopotential fiber sensor have electrical characteristics comparable to Ag/AgCl wet electrodes. We observed an improved ECG signal quality, reduced frequent electrode detachments, reduced wire clutter and entanglement, and improved sensor adherence to the skin over longer periods of time.

Action Potentials↗
PubMed1991

Synaptic 5'-nucleotidase is transient and indicative of climbing fiber plasticity during the postnatal development of rat cerebellum.

The transient appearance of 5'-nucleotidase, an adenosine-producing ecto-enzyme, was studied during specific stages of postnatal synaptogenesis in the rat cerebellum. For ultrastructural detection of 5'-nucleotidase activity, an enzyme-cytochemical technique was used. Between postnatal days 4 and 6, enzymatic reaction product was present in the synaptic clefts of climbing fibers containing the perisomatic spines, apical cones and emerging dendrites of Purkinje cells (CF-PC synapses). Labeled parallel fiber synapses were observed on dendritic shafts of cerebellar interneurons. At postnatal days 9 and 12, enzyme-positive parallel fiber terminals were in addition numerous on the spines of peripheral Purkinje branchlets, and gradually disappeared thereafter. Between postnatal days 8 and 15, labeling of perisomatic CF-PC contacts persisted. In contrast, climbing fiber synapses on Purkinje dendrites were only occasionally labeled. Between postnatal days 18 and 21, synaptic reaction product was restricted to mossy fibers. At the same time, association of 5'-nucleotidase with glial profiles was prominent throughout the cerebellar layers. In adult cerebellum (from 24 days onwards) all synapses were devoid of enzymatic activity. Throughout development, basket, stellate and Golgi cell synapses were devoid of enzymatic activity. We conclude that 5'-nucleotidase is present in excitatory cerebellar synapses during part of their generation period. The transient nature of this phenomenon suggests that 5'-nucleotidase may serve as a novel, cytochemical marker for a specific state of synaptic maturation, and in particular for climbing fiber plasticity. A role of 5'-nucleotidase in purinergic neuromodulation and cellular contact formation could be significant in these processes.

5'-Nucleotidase↗
PubMed2005

Bidirectional optical coupler for plastic optical fibers.

We have developed a low-loss bidirectional optical coupler for high-speed optical communication with plastic optical fibers (POFs). The coupler, which is fabricated by an injection molding method that uses poly (methyl methacrylate), has an antisymmetric tapered shape. We show that the coupler has low insertion and branching losses. The tapered shape of the receiving branch reduces beam diameter and increases detection efficiency coupling to a photodetector, whose area is smaller than that of the plastic optical fiber. The possibility of more than 15-m bidirectional transmission with a signaling bit rate up to 500 Mbits/s for simplex step-index POFs is demonstrated.

Journal Article↗
PubMed2003

Mitochondrial distribution and glycogen dynamics suggest diffusion constraints in muscle fibers of the blue crab, Callinectes sapidus.

The scaling of mitochondrial distribution, citrate synthase activity, and post-contractile glycogen recovery was examined in muscle fibers of the blue crab, Callinectes sapidus. The fast-twitch muscle fibers of C. sapidus can reach extremely large dimensions, which may impose constraints on aerobic metabolic processes. However, muscle cells from small crabs are not giant, meaning that during development muscle fibers cross and greatly exceed the surface area to volume (SAV) and diffusion threshold that is adhered to by the cells of most organisms. Cell diameters in the smallest size class were approximately 100 microm, while the largest size class had cell diameters in excess of 500 microm. In the smallest cells, the fractional area of subsarcolemmal and intermyofibrillar mitochondria was similar. However, in the largest cells, mitochondria were almost exclusively subsarcolemmal. Total fractional area of mitochondria was highest in the largest cells due to a proliferation of subsarcolemmal mitochondria. In contrast, citrate synthase activity decreased as cell size increased. Following burst contractile activity, glycogen concentrations decreased significantly and remained depressed for several hours in muscle comprised of giant cells, consistent with previous findings that anaerobic glycogenolysis fuels certain components of post-contractile recovery. However, in muscle composed of the smallest muscle cells, glycogen levels did not decrease significantly following burst activity. While normal scaling of aerobic metabolism would predict a slower aerobic recovery in larger animals, the present results suggest that cellular organization, SAV, and intracellular diffusion distances also impose constraints on aerobic processes in C. sapidus.

Animals↗