Biomedical subjects

S I Head

Publications and source records attributed to S I Head.

16 recordsLinked to original sources

PubMed2005

Synaptic plasticity in the dy2J mouse model of laminin alpha2-deficient congenital muscular dystrophy.

Laminin alpha2-deficient congenital muscular dystrophy is a debilitating disease affecting both muscle and neural tissue as a result of mutations in the LAMA2 gene. It presents at or soon after birth with muscle weakness and is further characterised by clinical central nervous system involvement. Laminin alpha2 is part of the extracellular matrix, linked to the cellular cystoskeleton via dystroglycan which is an integral part of the dystrophin-glycoprotein complex (DGC). We examined both short- and long-term synaptic plasticity in the C57BL6J/dy(2J) mouse, an animal model of laminin alpha2 deficient congenital muscular dystrophy. Using a cerebellar slice preparation, we show that the pre-synaptically mediated paired-pulse facilitation (PPF) was no different between dy(2J) and littermate controls. Approximately half (7/12) the dy(2J) Purkinje cells displayed a blunted LTD compared to littermate controls, and one third (4/12) of dy(2J) Purkinje cells displayed LTP. This study demonstrates that a defective laminin alpha2 causes a disruption in long-term synaptic plasticity at the Purkinje cell-parallel fibre synapse.

Animals↗

PubMed2002

Brain function in Duchenne muscular dystrophy.

Duchenne muscular dystrophy (DMD) is the second most commonly occurring genetically inherited disease in humans. It is an X-linked condition that affects approximately one in 3300 live male births. It is caused by the absence or disruption of the protein dystrophin, which is found in a variety of tissues, most notably skeletal muscle and neurones in particular regions of the CNS. Clinically DMD is characterized by a severe pathology of the skeletal musculature that results in the premature death of the individual. An important aspect of DMD that has received less attention is the role played by the absence or disruption of dystrophin on CNS function. In this review we concentrate on insights into this role gained from investigation of boys with DMD and the genetically most relevant animal model of DMD, the dystrophin-deficient mdx mouse. Behavioural studies have shown that DMD boys have a cognitive impairment and a lower IQ (average 85), whilst the mdx mice display an impairment in passive avoidance reflex and in short-term memory. In DMD boys, there is evidence of disordered CNS architecture, abnormalities in dendrites and loss of neurones, all associated with neurones that normally express dystrophin. In the mdx mouse, there have been reports of a 50% decrease in neurone number and neural shrinkage in regions of the cerebral cortex and brainstem. Histological evidence shows that the density of GABA(A) channel clusters is reduced in mdx Purkinje cells and hippocampal CA1 neurones. At the biochemical level, in DMD boys the bioenergetics of the CNS is abnormal and there is an increase in the levels of choline-containing compounds, indicative of CNS pathology. The mdx mice also display abnormal bioenergetics, with an increased level of inorganic phosphate and increased levels of choline-containing compounds. Functionally, DMD boys have EEG abnormalities and there is some preliminary evidence that synaptic function is affected adversely by the absence of dystrophin. Electrophysiological studies of mdx mice have shown that hippocampal neurones have an increased susceptibility to hypoxia. These recent findings on the role of dystrophin in the CNS have implications for the clinical management of boys with DMD.

Adult↗

PubMed2001

The role of nitric oxide in diaphragmatic dysfunction in endotoxemic rats.

In this study we examined the role of nitric oxide (NO) from inducible nitric oxide synthase (iNOS) and adenosine triphosphate (ATP) depletion, using aminoguanidine and 3-aminobenzamide, on diaphragm contractility in a rat model of sepsis. Intraperitoneal lipopolysaccharide (LPS) injection was used to induce septicemia in rats. The LPS treatment caused a decrease in maximal absolute force produced by the diaphragm muscle stimulated at 100 HZ, and the force-frequency curves were right-shifted with a decrease in force at 2, 5 and 15 HZ. LPS administration also made the diaphragm muscle strips more fatigable than controls. The decrease in force in LPS-treated animals was not due to an induction of pathological levels of i NOS. Increased fatigability did not appear to be due to a depletion of ATP through poly-adenosine-diphosphate-ribose polymerase (PARP) activation. This study does not support the hypothesis that the decrease in diaphragm muscle force as a result of sepsis is due to an induction of pathological levels of nitric oxide or ATP depletion.

Adenosine Triphosphate↗

PubMed2001

Effects of the PKA inhibitor H-89 on excitation-contraction coupling in skinned and intact skeletal muscle fibres.

This study investigated the effects of the protein kinase A (PKA) inhibitor, H-89, in mechanically-skinned muscle fibres and intact muscle fibres, in order to determine whether PKA phosphorylation is essential for normal excitation-contraction (E-C) coupling. In skinned EDL fibres of the rat, force responses to depolarization (by ion substitution) were inhibited only slightly by 10 microM H-89, a concentration more than sufficient to fully inhibit PKA. Staurosporine (1 microM), a potent non-specific kinase inhibitor, also had little if any effect on depolarization-induced responses. At 1-2 microM, H-89 significantly slowed the repriming rate in rat skinned fibres, most likely due to it deleteriously affecting the T-system potential. With 100 microM H-89, the force response to depolarization by ion substitution was completely abolished. This inhibitory effect was reversed by washout of H-89 and was not due to block of the Ca2+ release channel in the sarcoplasmic reticulum (SR). In intact single fibres of the flexor digitorum longus (FDB) muscle of the mouse, 1-3 microM H-89 had no noticeable effect on action-potential-mediated Ca2+ transients. Higher concentrations (4-10 microM) caused Ca2+ transient failure in fibres stimulated at 20 Hz in a manner indicative of action-potential failure. At 10-100 microM, H-89 also inhibited net Ca2+ uptake by the SR and affected the Ca2+-sensitivity of the contractile apparatus in rat skinned fibres. All such effects were proportionately greater in toad muscle fibres. These results do not support the hypothesis that phosphorylation is essential for the Ca2+ release channel to open in response to voltage-sensor activation in skeletal muscle fibres.

Animals↗

PubMed2000

Ca2+- and Sr2+-activation properties of muscle fibres from a muscle receptor organ and the associated extrafusal muscle of the crab and crayfish.

In this study on decapod crustaceans, we examined the Ca2+- and Sr2+-activation properties of skeletal muscle fibres from an identified proprioceptor, the thoracic coxal muscle receptor organ (TCMRO) and its extrafusal promotor muscle fibres. Proprioceptors and extrafusal muscles were isolated from a walking leg from the crayfish (Cherax destructor) and the rear swimming leg of the mud crab (Scylla serrata). The crayfish and mud crab TCMROs had very low Hill coefficient (nCa) values (1.86 +/- 0.08 and 1.64 +/- 0.03, respectively). In comparison to other skeletal muscle fibre types these low Hill coefficients would enable the length of the receptor muscles to be finely controlled over a wide range of [Ca2+]. Maximum force was found to be significantly lower in the TCMROs (crayfish: 5.76 +/- 0.98; crab: 4.80 +/- 0.56 Ncm(-2)), compared to their associated extrafusal promotor muscle fibres (crayfish: 10.69 +/- 1.63; crab: 20.07 +/- 1.98 Ncm(-2)), which is consistent with their sensory role. The muscle fibres of the crayfish TCMRO had faster contractile properties than the mud crab TCMRO, we discuss how these contractile properties relate to the type of locomotion undergone by each leg. The mud crab 'red' promotor and all crayfish promotor fibres were characterised as slow with low Hill coefficients (nCa: crayfish: 3.22 +/- 0.29; crab: 3.34 +/- 0.29) and a contractile apparatus with a high sensitivity to Ca2+ (pCa50: crayfish: 6.42 +/- 0.03; crab: 6.18 +/- 0.03). In contrast the 'white' mud crab promotor fibres from the swimming leg had contractile properties that were characteristic of fast fibres with a high mean Hill coefficient (nCa: 5.27 +/- 0.76) and a lower Ca2+ sensitivity (pCa50: 6.03 +/- 0.03). The sensitivity of the contractile apparatus to Sr2+ was very low (range of mean pSr50: 4.23 +/- 0.03-3.48 +/- 0.06) and low force levels were produced in comparison to that produced with Ca2+. The results of this study show that the muscle fibres of the sensory receptor, produce less force and have been adapted to enable the length of the receptor to be finely set in relation to the length of the extrafusal muscle. We discuss how the striated fibres of the receptor have been adapted to perform a sensory role and how this is related to the type of locomotion undergone by the legs. We also discuss how the fibre types of the extrafusal muscle have adapted to the mode of locomotion.

Animals↗

PubMed1998

Effect of clenbuterol on sarcoplasmic reticulum function in single skinned mammalian skeletal muscle fibers.

We examined the effect of the beta2-agonist clenbuterol (50 microM) on depolarization-induced force responses and sarcoplasmic reticulum (SR) function in muscle fibers of the rat (Rattus norvegicus; killed by halothane overdose) that had been mechanically skinned, rendering the beta2-agonist pathway inoperable. Clenbuterol decreased the peak of depolarization-induced force responses in the extensor digitorum longus (EDL) and soleus fibers to 77.2 +/- 9.0 and 55.6 +/- 5.4%, respectively, of controls. The soleus fibers did not recover. Clenbuterol significantly and reversibly reduced SR Ca2+ loading in EDL and soleus fibers to 81.5 +/- 2.8 and 78.7 +/- 4.0%, respectively, of controls. Clenbuterol also produced an approximately 25% increase in passive leak of Ca2+ from the SR of the EDL and soleus fibers. These results indicate that clenbuterol has direct effects on fast- and slow-twitch skeletal muscle, in the absence of the beta2-agonist pathway. The increased Ca2+ leak in the triad region may lead to excitation-contraction coupling damage in the soleus fibers and could also contribute to the anabolic effect of clenbuterol in vivo.

Adrenergic beta-Agonists↗

PubMed1997

Time course of calcium transients derived from Fura-2 fluorescence measurements in single fast twitch fibres of adult mice and rat myotubes developing in primary culture.

In this study, we applied a method to correct for the altered binding kinetics of Fura-2 for Ca2+ in vivo, on Ca2+ fluorescence transients (Ca2+F) measured using Fura-2 in single adult fast twitch skeletal muscle fibres of the mouse, which exhibit very fast [Ca2+] responses, and rat myotubes developing in culture which exhibit slower [Ca2+] responses (rise time [20-80% of peak] of Ca2+F transients: 1.81 +/- 0.17 ms and 16.14 +/- 2.60 ms, respectively). After correction, the [Ca2+] transients (Ca2+C) measured in both the adult mouse fibres and the myotubes rose more rapidly (mean rise time of Ca2+C transients: adult mouse fibres, 0.76 +/- 0.12 ms; rat myotubes, 8.25 +/- 2.83 ms) and often exhibited a Ca2+ spike which exceeded the peak of the Ca2+F transient. In the adult mouse fibres, correction increased the mean peak [Ca2+] of the Ca2+F transients by a factor of 7 from 0.53 +/- 0.08 microM to 3.76 +/- 0.71 microM. The accuracy of the time course of the corrected Ca2+ transients was confirmed by comparison to the time course of Ca2+ transients measured with Mag-Fura-5, which had a similar mean rise time (0.94 +/- 0.10 ms, t-test, P = 0.80). The more slowly rising Ca2+ transients measured in the rat myotubes were less affected by the correction process, increasing in mean peak [Ca2+] by a factor of only 1.2 from 0.82 +/- 0.17 microM to 0.97 +/- 0.15 microM. During the decay phase of the Ca2+ transients elicited in the adult mouse fibres and the myotubes, the corrected Ca2+C signal largely followed the unmodified Ca2+F transient. The correction process was found to have little effect on Ca2+ transients with rise time values greater than 10 ms, which included most of the Ca2+ transients measured in the myotubes.

Animals↗

PubMed1996

Measurement of membrane potential and myoplasmic [Ca2+] in developing rat myotubes at rest and in response to stimulation.

In this study, the membrane potential and cytosolic [Ca2+] were measured in rat myotubes developing in culture from days 6-14. It was found that as the myotubes developed in culture, the resting membrane potential (RMP) became more negative during days 6-8, and then did not significantly change until after day 13, when it started to become less negative. The mean RMP measured at days 8-13 was -59 +/- 1 mV (n = 70). The amplitude of action potentials elicited in the myotubes by anode break stimulation increased in size during development (range: 47.5-119 mV) and this closely correlated with the development of a more negative RMP. Cytosolic [Ca2+] was measured in the rat myotubes using the Ca2+ indicator Fura-2, and no significant change in the resting [Ca2+] was observed during development (days 6-14). Ca2+ responses triggered by action potentials varied from small slow increases in [Ca2+] that failed to return to the baseline to rapid [Ca2+] transients. The size of the [Ca2+] transients positively correlated with both the observed increase in the RMP during development and the size of the action potential. Larger [Ca2+] transients also had more rapid rates of [Ca2+] decay, indicating a tandem increase in the ability of the sarcoplasmic reticulum to release and resequester Ca2+ during development of rat myotubes. Repetitive stimulation (10 Hz) of the myotubes exhibiting small [Ca2+] transients produced a step-like rise in [Ca2+]. Many myotubes exhibiting larger [Ca2+]transients could not be stimulated at 10 Hz by anode break stimulation due to the presence of action potentials with large hyperpolarisations. However, when these myotubes were depolarised at 10 Hz, they produced a tetanic Ca2+ response similar to that seen in adult skeletal muscle.

Animals↗

PubMed1994

Mechanism of action of a K+ channel activator BRL 38227 on ATP-sensitive K+ channels in mouse skeletal muscle fibres.

1. Investigations were made into the effects of BRL 38227, a potassium channel activator, on ATP-sensitive potassium channels (K+ATP channels) in single fibres dissociated from the flexor digitorum brevis muscle of C57BL/6J mice. 2. In cell-attached patches BRL 38227 (100 microM) caused activation of a glibenclamide-sensitive potassium current. Linear slope conductance of the inward current, partial rectification of the outward current and glibenclamide sensitivity indicate that K+ATP channels are the site of action of BRL 38227. 3. In the absence of ATP at the cytoplasmic side of excised inside-out patches, BRL 38227 caused direct and magnesium-dependent activation of K+ATP channels. The degree of activation diminished with successive applications of BRL 38227. 4. BRL 38227 also caused activation of K+ATP channels in the presence of low (< 100 microM) but not high (1.0 mM) ATP, particularly in patches containing large numbers of channels. 5. BRL 38227 and 5 microM MgATP failed to activate channels following complete run-down. 6. Results show that BRL 38227 caused direct activation of K+ATP in skeletal muscle and that this was mediated through a magnesium-dependent binding site rather than alleviation of inhibition by competitive displacement of ATP from the inhibitory site.

Adenosine Triphosphate↗

PubMed1993

Ca2+ levels in myotubes grown from the skeletal muscle of dystrophic (mdx) and normal mice.

1. Myotubes were grown in culture from normal (C57BL/ScSn) and mdx mice and the cytosolic [Ca2+] was monitored through development (5-21 days in culture) using fura-2 loaded via ionophoresis. Simultaneous measurements of the membrane potential and cytosolic [Ca2+] were made in normal and mdx myotubes before, during and after stimulation by action potentials elicited following anode break excitation. All experiments were undertaken at 22 degrees C. All data are expressed as means +/- S.E.M. 2. A new method was developed which enabled accurate determination of the fluorescence characteristics of fura-2 in murine skeletal muscle fibres. In the under in vitro conditions by 14.60 +/- 0.05, 9.40 +/- 0.15 and 6.90 +/- 0.43% respectively. 3. The resting cytosolic [Ca2+] in the mdx myotubes was consistently higher than in the normal myotubes throughout the developmental period measured. Overall, the resting cytosolic [Ca2+] in mdx myotubes (134 +/- 9 nM, n = 22) was twofold higher than in normal myotubes (66 +/- 6 nM, n = 26). After stimulation (one to three action potentials) the cytosolic [Ca2+] of both mdx and normal myotubes remained elevated. The mdx myotubes (236 +/- 55 nM, n = 5) again had approximately double the cytosolic [Ca2+] of normal myotubes (109 +/- 19 nM, n = 9). 4. The time course and amplitude of the Ca2+ responses measured in the mdx and normal myotubes after action potential stimulation were variable. Two categories of Ca2+ response were observed in mdx and normal myotubes, the first consisted of a small, slow rise in [Ca2+] that remained elevated and the second consisted of a rapid (time to peak 7.4 +/- 1.5 ms) (n = 8) rise in [Ca2+] with amplitudes in the range 61-773 nM and a [Ca2+] decay rate constant of 4.35 +/- 1.57 s-1 (n = 8) (range 0.96-15 s-1). 5. In conclusion, the elevated cytosolic [Ca2+] reported here through development of cultured mdx myotubes suggests that this genetic disorder results in a defect which compromises the ability of the myotubes to strictly regulate cytosolic [Ca2+]. The results are consistent with the presence of functionally abnormal Ca2+ channels recently reported in mdx myotubes.

Animals↗

PubMed1993

Contractile properties of skinned muscle fibres from young and adult normal and dystrophic (mdx) mice.

1. Single muscle fibres were enzymatically isolated from the soleus and extensor digitorum longus (EDL) muscles of genetically dystrophic mdx and normal (C57BL/10) mice aged 3-6 or 17-23 weeks. 2. Fibres of both muscles were chemically skinned with the non-ionic detergent Triton X-100 (2% v/v). Ca(2+)- and Sr(2+)-activated contractile responses were recorded and comparisons were made between several contractile parameters of various fibre types of normal and dystrophic mice of similar age. 3. There were no significant differences in the following contractile parameters of skinned fibres of normal and mdx mice of the same age: sensitivity to activating Ca2+ (pCa50) or Sr2+ (pSr50) and differential sensitivity to the activating ions (pCa50-pSr50). However the maximum isometric tension (Po) and the frequency of myofibrillar force oscillations in EDL fast-twitch fibres of young mdx mice were significantly lower than those of soleus fast-twitch fibres of the same animals, or fast-twitch fibres (EDL or soleus) of normal mice. 4. Age-related differences were apparent in some contractile parameters of both normal and mdx mice. In particular the steepness of force-pCa and force-pSr curves increased with age in normal mice, yet decreased with age in fibres of mdx mice. 5. A fluorescent probe, ethidium bromide, which interchelates with DNA, was used with laser-scanning confocal microscopy to determine the distribution of myonuclei in fibres. Fibres isolated from either muscle type of normal animals displayed a characteristic peripheral spiral of myonuclei. Fibres from muscles of mdx mice displayed three major patterns of nuclear distribution; the normal peripheral spiral, long central strands of nuclei, and a mixture of these two patterns. 6. The contractile characteristics of mdx fibres were not markedly influenced by the nuclear distribution pattern in that there were no discernible differences in the major contractile parameters (the Hill coefficients nCa and nSr, which are associated with the steepness of the Ca2+ and Sr2+ activation curves, pCa50, pSr50, pCa50-pSr50) of skinned fibres possessing peripheral or central nuclei. However, except for nSr, these values were all lower in individual fibres which displayed similar proportions of central and peripheral nuclei. The presence of mixed nucleation and absence of fibres with embryonic contractile characteristics in mdx mice suggest that the dystrophin-negative fibres can repair locally occurring muscle damage.

Aging↗

PubMed1993

Membrane potential, resting calcium and calcium transients in isolated muscle fibres from normal and dystrophic mice.

1. Single skeletal muscle fibres were enzymatically isolated from the flexor digitorum brevis muscles (FDB) of dystrophic mdx and control C57BL/10 mice aged 3-9 weeks. In this age range the majority (> 95%) of the mdx fibres were morphologically normal. 2. There was no significant difference between the resting membrane potential (RMP) of mdx and control mice, -71.2 +/- 1.21 (n = 26) and -70.6 +/- 1.15 mV (n = 42), respectively. 3. At RMP more negative than -60 mV the resting calcium (recorded with fura-2, free acid ionophoresed into cell) in the dystrophic mdx cells was not significantly different from the normal animals, 45.7 +/- 4.1 (n = 10) and 46.2 +/- 3.9 nM (n = 9), respectively. 4. The resting cytosolic calcium concentration was measured simultaneously with the RMP. At RMP between -60 to -17 mV there was an increase in the resting calcium concentration in both mdx and control ranging from 79.3 to 252 nM. This increase was most probably due to the activation of the slow calcium current. 5. Fura-2 calcium transients were produced via single action potential stimulation using an intracellular microelectrode both to stimulate the cell and record potential changes. There was no significant difference between the rise time (Tp) or half-decay time (T1/2) at 22 degrees C of the calcium transient in response to a single action potential in mdx compared to normal animals, 5.9 +/- 0.34 (n = 8) and 5.4 +/- 0.36 ms (n = 7); 39.5 +/- 2.9 (n = 8) and 40.75 +/- 3.7 ms (n = 7), respectively.(ABSTRACT TRUNCATED AT 250 WORDS)

Action Potentials↗

PubMed1992

Abnormalities in structure and function of limb skeletal muscle fibres of dystrophic mdx mice.

In this study we have shown that the skeletal muscle fibres from adult (older than 26 weeks) mdx mice have gross structural deformities. We have characterized the onset and age dependence of this feature in mdx mice. The three dimensional structure of these deformities has been visualized in isolated fibres and the orientation of these deformities was determined within the muscle by confocal laser scanning microscopy. We have also shown that the occurrence of morphologically abnormal fibres is greater in muscles with longer fibres (extensor digitorum longus (EDL) and soleus, 6-7.3 mm long), than in muscles with shorter fibres (flexor digitorum brevis (FDB), 0.3-0.4 mm long). A population of post-degenerative fibres, with both central and peripheral nuclei coexistent along the length of the fibre, has also been identified in the muscles studied. We showed that a mild protocol of lengthening (eccentric) contractions (the muscle was stretched by 12% during a tetanic contraction) caused a major reduction in the maximal tetanic force subsequently produced by mdx EDL muscle. In contrast, maximal tetanic force production in normal soleus, normal EDL and mdx soleus muscles was not altered by this protocol. We suggest that the deformed fast glycolytic fibres which are found in adult mdx EDL but not in adult mdx soleus muscles are the population of fibres damaged by the lengthening protocol.

Aging↗

PubMed1991

Reflex actions of one proprioceptor on the motoneurones of a muscle receptor and their central modulation in the shore crab.

1. Reflex efferent control of a muscle stretch receptor by a joint proprioceptor of the same limb was studied in an isolated CNS preparation from the shore crab. The influence of 'fictive locomotor' activity on this interjoint reflex was also examined. 2. The thoracic-coxal muscle receptor organ (TCMRO) and the coxo-basal chordotonal organ (CBCO), which monitor movement and position of the first and second joints of the posterior leg, were isolated together with the whole thoracic ganglion complex. The TCMRO, functionally analogous to a mammalian muscle spindle, has two receptor motoneurones. RM1 innervating the receptor muscle alone and RM2 which also supplies the 'extrafusal' promotor muscle. The CBCO is a typical arthropod elastic strand organ, with many sensory neurones but lacking an efferent supply. The TCMRO was fixed at its mid-length, and stretch-hold-release stimuli were applied to the CBCO. Efferent activity was recorded from the cut nerve roots of the four basal limb muscles and intracellularly as excitatory junction potentials (EJPs) from the receptor muscle. 3. A dynamic increase in the frequency of action potentials in RM1 occurred on both stretch and release of the CBCO. During the hold phase the RM1 activity declined from the dynamic response but remained elevated compared to the resting tonic discharge. RM2, identified by EJPs occurring 1:1 with a unit in the promoter nerve, responded in a similar way. 4. One or more promotor motoneurones were usually co-activated with the two receptor efferents in response to input from the CBCO. In a typical example the average spike frequency of RM1 rose from 0 to 27 Hz during the dynamic phases (stretch and release) of the CBCO stimulus, falling to 2.5 Hz during the hold phase, while the corresponding promotor spike frequencies were 25 and 7.5 Hz, respectively. The other three muscle nerves recorded from generally also showed reflex driving by the CBCO. 5. The totally isolated thoracic ganglion could produce a rhythmic, bursting motor output in the absence of any sensory input. During this centrally generated activity the receptor motor innervation was strongly co-activated with the promotor bursts, and the reflex input from the CBCO was overridden or modulated in a phase-dependent manner. 6. The proximally directed interjoint reflex to the receptor muscle probably functions to maintain the tension on the sensory endings of the TCMRO, and so enable them to respond effectively at all times to movements of the basal leg joint.(ABSTRACT TRUNCATED AT 400 WORDS)

Action Potentials↗

PubMed1990

Properties of enzymatically isolated skeletal fibres from mice with muscular dystrophy.

1. Single intact muscle fibres were enzymatically isolated from the skeletal muscles of the dystrophic mouse 129/ReJ dy/dy and were subjected to a range of physiological interventions. 2. Electrophysiological measurements, diffusion of injected dyes (Lucifer Yellow), microdissection and general appearance in the light microscope have shown that the majority of skeletal fibres isolated from the soleus and extensor digitorum longus (EDL) of adult dystrophic mice (10-14 weeks old) had gross morphological abnormalities. These abnormalities ranged from simple branching of the fibre to interconnections of many fibre branches which form a complex syncitium. 3. Segments from fibres of normal appearance and from fibres with morphological deformities were chemically skinned with Triton X-100 and activated in Ca2(+)- and Sr2(+)-buffered solutions. The different characteristics of the Ca2(+)- and Sr2(+)-activation curves were also used to identify the fibre type. 4. Gross morphological abnormalities were observed both in fibres which had predominantly slow-twitch and fast-twitch characteristics. 5. A new group of fibres was found to exist in the soleus muscle of dystrophic animals and represented about 18% of the entire soleus fibre population. This group of fibres had predominantly fast-twitch characteristics and some of these fibres were also grossly malformed. 6. The activation characteristics of individual branches from the same complex syncitium were similar, indicating that the contractile and regulatory proteins were of one type in one syncitium. 7. Chemically skinned segments from malformed fibres which included a major deformity between the points of attachment were generally unable to sustain near-maximal forces. 8. The proportion of malformed fibres which remained intact decreased markedly after prolonged tetanical stimulation of the intact muscle. This strongly suggests that malformed fibres are also functionally weak and prone to progressive damage when stimulated within the intact muscle. 9. The presence in large proportions of fibres with gross morphological abnormalities may explain the symptoms of severe and progressive muscle weakness and muscle loss which are apparent in the 129/ReJ dy/dy mice and possibly even in the human dystrophies such as Duchenne muscular dystrophy.

Animals↗

PubMed1990

Resting calcium concentrations in isolated skeletal muscle fibres of dystrophic mice.

1. Single, intact muscle fibres were dissociated enzymatically from skeletal muscles of phenotypically normal (+/?) and dystrophic mice (129/ReJ dy/dy: Dystrophia muscularis), and resting Ca2+ levels were measured by image analysis of intracellular Fura-2 fluorescence in distinct parts of the fibres. 2. Fura-2 was introduced into fibres by ionophoresis with glass microelectrodes to concentrations of between 50 and 200 microM. Over this concentration range there was no apparent buffering of intracellular Ca2+ by Fura-2. 3. Fibres isolated from the soleus, flexor digitorum brevis (FDB) and extensor digitorum longus (EDL) muscles of normal animals maintained resting [Ca2+] of 106 +/- 2 nM. Ca2+ distributions within individual fibres were homogeneous. 4. Fibres from dystrophic animals maintained [Ca2+] that was elevated two- to fourfold in comparison to normal fibres. 5. The population of skeletal fibres from dystrophic mice which displayed morphology similar to that of fibres of normal animals were found to have Ca2+ levels that averaged 189 +/- 2 nM. The distribution of Ca2+ within these fibres appeared uniform. 6. The population of dystrophic fibres that possessed morphological abnormalities maintained even higher Ca2+ concentrations (368 +/- 3 nM). Several fibres from this morphological group displayed obvious heterogeneity in Ca2+ distribution with distinct, localized areas of higher Ca2+. 7. These results support the contention that Ca2+ homeostasis is markedly impaired in dystrophic muscle. The elevated Ca2+ levels are near the threshold for contraction and, together with severe morphological fibre abnormalities, are probably centrally involved in fibre necrosis apparent in muscular dystrophy.

Animals↗