Biomedical subjects

A C Wareham

Publications and source records attributed to A C Wareham.

At least 19 recordsLinked to original sources

PubMed1999

Increased quantal release of acetylcholine at the neuromuscular junction following scald injury in the rat.

Following severe burns, patients frequently develop a profound resistance to nondepolarizing neuromuscular blockers. Several mechanisms have been proposed to account for this, including upregulation of nicotinic acetylcholine receptors. We investigated the effects of a 30% body surface area (BSA) scald on neuromuscular transmission in slow-twitch soleus (SOL) and fast-twitch extensor digitorum longus (EDL) of rats. Rats were sacrificed 72 h after the injury, a time at which sepsis is unlikely and body weight gain and core temperature have returned to normal. Further groups of rats were sham operated and either pair fed to the scalded rats or freely fed to assess the influence of food restriction. When compared with muscle from pair-fed control rats, scald resulted in an almost 50% increase in miniature endplate potential (mEPP) frequency in both SOL and EDL. However, scald did not increase mean mEPP amplitude in SOL, although it did cause a 10% increase in EDL. Scald injury did produce a significant increase in the size of the evoked endplate potential in SOL (33%) and EDL (37%). These data indicate that a significant increase in the quantal content of evoked transmitter released in SOL (38%) and EDL (30%) occurred by 72 h after scald. Such an increase may contribute to the resistance to nondepolarizing neuromuscular blockers documented in patients following thermal injury.

Acetylcholine↗

PubMed1998

Electrophysiology of the neuromuscular junction of the laminin-2 (merosin) deficient C57 BL/6J dy2J/dy2J dystrophic mouse.

The C57 BL/6J dy2J/dy2J dystrophic mouse expresses an abnormal truncated form of the alpha2 subunit of the protein laminin-2 (or merosin), which is unable to form a stable link between the extracellular matrix and the dystrophin-associated proteins, resulting in muscular dystrophy. Morphological abnormalities of the peripheral nervous system and neuromuscular junction have also been reported. The electrophysiological properties of the neuromuscular junctions of diaphragm, extensor digitorum longus (EDL), and soleus from C57 BL/6J dy2J/dy2J mice and controls are described. No evidence for the presence of denervated fibres were found. Mean MEPP amplitudes were significantly increased in EDL and soleus but reduced in the diaphragm from affected mice. Mean MEPP frequencies were raised in all the dy2J/dy2J muscles studied. dy2J/dy2J muscles were paralysed by low concentrations of mu-conotoxin suggesting that embryonic (tetrodotoxin and mu-conotoxin resistant) sodium channels are not widespread on dy2J/dy2J muscle as has previously been reported. EPP latencies were significantly prolonged in the diaphragm and EDL but not soleus from dy2J/dy2J mice. Quantal contents were higher in all dy2J/dy2J muscles. In the dy2J/dy2J diaphragm failures in neurotransmission occurred and a faster rate of rundown of EPPs were apparent. Some changes appear from a direct effect of dystrophy, whilst increased MEPP frequency and quantal content, and failures in neurotransmission indicate neuronal abnormalities.

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↗

PubMed1995

Effects of clenbuterol and ICI118551, a selective beta 2-antagonist, on the growth of skeletal muscle of suckling rats.

The beta 2-adrenergic agonist, clenbuterol, was administered to lactating rats (4 mg/kg diet) from post-partum day 1 to day 19, or directly injected into neonate rats (0.1 and 1.0 mg/kg body weight) from post-partum day 3 until day 15. Changes in body weight and the skeletal muscles soleus (SOL) and extensor digitorum longus (EDL) were studied in both dams and suckling offspring. Drug treatment consistently increased body weight in dams whilst significantly reducing the growth of their suckling pups. In dams treated with clenbuterol (4 mg/kg of diet) muscle weights and protein contents were significantly increased. Total protein content increased by 16% in SOL and 47% in EDL after 19 days of treatment. In contrast, in their suckling pups, there was a 22% and 26% reduction in protein content of SOL and EDL respectively. Administration of the beta 2-antagonist ICI118551 to these pups failed to prevent these reductions in body and muscle weights. Hence, if clenbuterol did reach the pups via the milk from treated mothers it did not act via conventional beta 2-receptors. Injection of pups with clenbuterol (1.0 mg/kg every 12 h) from litters suckling from untreated dams also resulted in significant reductions in muscle weights and protein contents. Protein content was reduced by 10% in SOL and 13% in EDL after 12 days of treatment. No alteration in fibre type proportion in SOL or EDL resulted from this treatment. Further work is required to determine whether the growth suppression in the two situations occurs via the same mechanism.

Adrenergic beta-2 Receptor Antagonists↗

PubMed1995

Effect of the beta 2-adrenergic agonist clenbuterol on the growth of fast- and slow-twitch skeletal muscle of the dystrophic (C57BL6J dy2J/dy2J) mouse.

Clenbuterol (4mg/kg in diet for 21 days) had no statistically significant effect on whole body growth. It did cause a significant increase (18.2%) in wet weight of the fast twitch muscle extensor digitorum longus (EDL) and a corresponding 14.9% increase in total muscle protein. In transverse sections through dystrophic muscle fibre sizes were more variable than in normal muscle. Clenbuterol treatment resulted in a reduction in the proportion of small diameter fibres, and therefore an increase in mean fibre diameter, in dystrophic EDL. Clenbuterol had no significant effect upon the slow twitch muscle soleus.

Administration, Oral↗

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↗

PubMed1994

Rundown and reactivation of ATP-sensitive potassium channels (KATP) in mouse skeletal muscle.

Dissociated single fibers from the mouse flexor digitorum brevis (FDB) muscle were used in patch clamp experiments to investigate the mechanisms of activation and inactivation of KATP in mammalian skeletal muscle. Spontaneous rundown of channel activity, in many excised patches, occurred gradually over a period of 10-20 min. Application of 1.0 mM free-Ca2+ to the cytoplasmic side of the patch caused irreversible inactivation of KATP within 15 sec. Ca(2+)-induced rundown was not prevented by the presence of 1.0 microM okadaic acid or 2.0 mg ml-1 of an inhibitor of calcium-activated neutral proteases, a result consistent with the conclusion that phosphatases or calcium-activated neutral proteases were not involved in the rundown process. Application of 1.0 mM Mg.ATP to Ca(2+)-inactivated KATP caused inhibition of residual activity but little or no reactivation of the channels upon washout of ATP, even in the presence of the catalytic subunit of cyclic AMP-dependent protein kinase (10 U ml-1). Mg.ATP also failed to reactivate KATP, even after only partial spontaneous rundown, despite the presence of channels that could be activated by the potassium channel opener BRL 38227. Nucleotide diphosphates (500 microM; CDP, UDP, GDP and IDP) caused immediate and reversible opening of Ca(2+)-inactivated KATP. Reactivation of KATP by ADP (100 microM) increased further upon removal of the nucleotide. In contrast to KATP from cardiac and pancreatic cells, there was no evidence for phosphorylation of KATP from the surface sarcolemma of dissociated single fibers from mouse skeletal muscle.(ABSTRACT TRUNCATED AT 250 WORDS)

Adenosine Triphosphate↗

PubMed1994

Low quantal content of the endplate potential reduces safety factor for neuromuscular transmission in the diaphragm of the newborn rat.

We have used phrenic nerve-hemidiaphragm preparations from rats aged 11-28 days to determine the electrophysiological basis of the three-fold increase in sensitivity of the neonatal neuromuscular junction to non-depolarizing neuromuscular blocking drugs. The mean resting membrane potential (RMP) of intact muscle fibres at 19-21 degrees C increased from -67.1 (SEM 0.6) mV at 11 days to -75.8 (0.06) mV at 28 days. Over the same age range the mean size of miniature endplate potentials (MEPP) decreased from 1.7 (0.1) mV to 1.4 (0.03) mV, and the threshold depolarization to produce a muscle action potential increased from 6.8 (0.4) mV to 9.5 (0.6) mV. The quantal content of the evoked endplate potential (EPP) calculated from EPP and MEPP measured in cut muscle fibres was only 6.6 (0.9) at 11 days, but this increased to 21.1 (2.3) at 21 days. As a result of the low quantal content of the EPP, the safety factor for neuromuscular transmission was only 1.7 at 11 days, compared with 2.8 at 20 days. A safety factor of 1.7 in the youngest rats indicates that block of only about 40% of postjunctional acetylcholine receptors would result in failure of neuromuscular transmission. Hence, a low quantal content of EPP, leading to a low safety factor for neuromuscular transmission, probably underlies the increased sensitivity to non-depolarizing neuromuscular blocking drugs of young rats and, by inference, human neonates.

Action Potentials↗

PubMed1992

Effect of clenbuterol on skeletal muscle atrophy in mice induced by the glucocorticoid dexamethasone.

1. The ability of clenbuterol to antagonize the catabolic effect of the glucocorticoid dexamethasone on the skeletal muscles, soleus, gastrocnemius and extensor digitorum longus was studied in mice. 2. Daily injections of 5 mg dexamethasone/kg body weight over 10 days caused a significant (20%) loss of muscle weight and protein content in fast twitch but not in slow twitch muscles. 3. Inclusion of clenbuterol (4 mg/kg) in the diet for the period of dexamethasone treatment partly prevented glucocorticoid-induced muscle atrophy, and increasing the concentration of clenbuterol to 8 mg/kg diet totally prevented glucocorticoid-induced protein loss in all muscles.

Animals↗

PubMed1992

Age-dependent variation in response to tubocurarine in the isolated rat diaphragm.

The EC50 of tubocurarine was determined in phrenic nerve-hemidiaphragm preparations obtained from 35 Sprague-Dawley rats aged 0-46 days. We measured also the ratio of the fourth to the first twitch in the train-of-four (T4:T1) when the first twitch of the train was depressed to 50% of control. The preparation was not unduly sensitive to tubocurarine at 0 days and there was little evidence of T4:T1 fade. However, by age 11 days the preparation exhibited fade and a three-fold sensitivity to tubocurarine similar to that in the human neonate. We conclude that the phrenic nerve-hemidiaphragm preparation from 11-day-old rats should be a suitable model in which to investigate the biochemical and electrophysiological basis of the sensitivity seen in humans.

Age Factors↗

PubMed1990

Effect of clenbuterol on normal and denervated muscle growth and contractility.

The reported anabolic action of some beta 2 agonists may have clinical applications in certain muscle wasting states. Administration of clenbuterol (2 mg/kg diet for 14 days) to rats resulted in a limited degree of hypertrophy of normal muscles; the effect was more pronounced on fast-twitch muscles than on slow-twitch muscles. The anabolic effect was greatest in denervated muscles, where it was significantly more effective on the slow-twitch type. Clenbuterol significantly improved the contractile properties of denervated slow-twitch muscle, reverting them toward normal, but had little effect on contractile properties of denervated fast-twitch muscle. Such differential effects of clenbuterol must be taken into consideration in the evaluation of any future human intervention study.

Animals↗

PubMed1990

Sarcolemmal K+ channel activity in developing rat skeletal muscle membranes.

A method has been adapted to produce membrane vesicles suitable for routine membrane patch clamping from neonate rat skeletal muscle. Single K+ channel activity was recorded from cell-free inside-out patches. Most Ca2(+)-activated voltage sensitive channels had large conductances of up to 300 pS, as determined from their current/voltage relationship, and an open probability (Po) approaching unity at positive membrane potentials. A lower conductance K+ channel, probably responsible for inward rectification, had a lower conductance of about 100 pS. Outward rectifying K+ channels were also observed with the lowest conductance, about 40 pS. 0.1 mM ATP when applied to the inner membrane surface reduced or blocked activity, drastically reducing Po without altering single channel conductance. Such an effect has been reported in other preparations but was different in the neonate preparation in that it blocked channels with conductances as high as 300 pS. The simple preparation described, which we have also used successfully on mature rat and mouse skeletal muscle, has potential in the analysis of channel activities in various conditions and pathologies without the need for tissue culture to produce suitable membrane preparations.

Adenosine Triphosphate↗

PubMed1990

Potassium channel activity in sarcolemmal vesicles formed from skeletal muscle fibres of normal and dystrophic mice.

Sarcolemmal vesicles were produced from adult mouse extensor digitorum longus muscle (EDL) by treating swollen muscle fibres with collagenase. Vesicles formed from dystrophic (C57BL/6J dy/dy) and phenotypically normal animals were patch clamped and the single channel activity was recorded. Three types of K+ channel were observed in excised patches taken from normal and dystrophic muscle. A large conductance (300 pS) Ca2(+)-dependent K+ channel (KCa) was the most frequently observed of the K+ channels in both types of muscle preparation. In a number of patches taken from dystrophic muscle the open probability-voltage relationship for the KCa channel was markedly different from that in normal muscle, suggesting a possible reduction in Ca2+ sensitivity. An ATP-sensitive K+ channel (90 pS) was common to both normal and dystrophic muscle vesicles and was present in a large number of patches. An inwardly rectifying K+ channel (40 pS) was also observed in both types of sarcolemmal vesicles. The properties of all three K+ channels types were broadly consistent with other observations of skeletal muscle K+ channels, though all had higher conductances than had previously been noted in other species.

Adenosine Triphosphate↗

PubMed1987

Effects of denervation and immobilisation during development upon [3H]ouabain binding by slow- and fast-twitch muscle of the rat.

The effect of innervation and of muscle inactivity upon the normal production of Na+-K+-ATPase sites, assayed by [3H]ouabain binding, in muscle surface membranes has been determined for the rat. In both slow-twitch soleus (SOL) and fast-twitch extensor digitorum longus (EDL) muscles a large increase was found to occur in [3H]ouabain binding per unit weight of muscle over the first 3 weeks of life. Interruptions of development, brought about by fixation of muscles at different lengths at 5 days of age, had no significant effect upon [3H]ouabain binding by EDL. In contrast, fixation led to a decrease in binding in SOL. When fixed in a shortened position profound morphological changes occurred, although these were not apparent when SOL was fixed in a stretched position. Denervation of SOL at 5 days of age significantly reduced the age related increase in the density of [3H]ouabain binding, whilst denervation of EDL had little effect. It was concluded that normal development of SOL is dependent upon innervation and possibly the resulting muscle activity, whereas development of EDL was relatively independent of innervation.

Animals↗

PubMed1986

Effect of denervation during development upon membrane potential and intracellular potassium and sodium activities of skeletal muscle of the rat.

Potassium and sodium ion-selective microelectrodes were used in vitro to investigate the effect of denervation of fast-twitch skeletal muscle (extensor digitorum longus) at 5 days of age upon subsequent development of the resting membrane potential. Normally, during the first 3 weeks of life the balance of intracellular potassium and sodium changed to elevate potassium and to lower sodium. These changes were reflected in the development of a more hyperpolarized resting membrane. Periods of denervation delayed, or prevented, the changes in ion activities and membrane hyperpolarization from occurring. The results are compared with those found after denervation of adult skeletal muscle.

Animals↗

PubMed1986

Membrane potential and intracellular potassium and sodium activities after denervation of soleus muscles of neonatal rats.

Potassium and sodium ion-selective microelectrodes were used in vitro to investigate the depolarization of slow-twitch skeletal muscle fibers (soleus) following denervation at 5 to 6 days of age. The normal increase in intracellular potassium ion activity and decrease in sodium ion activity which occurs during the first 3 weeks of life were delayed or prevented by denervation. The results are compared with those found after similar periods of denervation of neonatal rat fast-twitch muscle.

Animals↗

PubMed1985

Changes in membrane potential and potassium and sodium activities during postnatal development of mouse skeletal muscle.

Dual-channel potassium-selective and single-channel sodium-selective microelectrodes were used to investigate the cause of changes in resting membrane potential of muscle fibers of the mouse during early development. The resting membrane of extensor digitorum longus fibers hyperpolarized during the postnatal period from -41.8 mV at 4 days of age to -76.4 mV at 27 days. During this period intracellular potassium activity increased by 42.1% from 82.5 mM at 8 days to 117.2 mM at 29 days. Intracellular sodium activity was high at 8 days, 23.7 mM, but decreased rapidly to adult values by 27 days when it was 9.98 mM, a 57.9% reduction in sodium activity. The time course of the change in resting membrane potential was different from that of the potassium equilibrium potential calculated from the data. If only potassium and sodium ions were to make significant contributions to the potential, then it was calculated that the permeability ratio PNa:PK would have to change from a value of 0.0659 at 8 days to 0.0227 at 27 days. The results indicated that other factors might be involved in generating the membrane potential inasmuch as, although both intracellular potassium and sodium activities did not change significantly after 27 to 30 days, the membrane potential had not attained adult values at that time. The possibility that increases in muscle activity during the postnatal period might initiate the changes in membrane polarization and intracellular ion activities is discussed together with possible complications in interpretation due to great variations in fiber diameters.

Animals↗

PubMed1985

[3H]Ouabain binding in normal and dystrophic mouse skeletal muscles and the effect of age.

The numbers of Na+-K+ ATPase sites in skeletal muscles of normal and dystrophic mice between 3 and 17 months of age have been estimated using [3H]ouabain binding assays. In normal mice, at all ages, slow twitch muscle, soleus (SOL), bound significantly more [3H]ouabain than fast-twitch muscle, extensor digitorum longus (EDL). [3H]Ouabain binding did not alter in either SOL or EDL from normal mice over the age range studied. The numbers of Na+-K+ ATPase sites did alter in muscles taken from dystrophic mice (C57BL/6J dy2J/dy2J). In EDL there was an increase and in SOL a decrease in [3H]ouabain binding. This may be related to a change in muscle fibre metabolism from glycolytic to oxidative or to an altered activity pattern. Increasing age resulted in a progressive reduction in [3H]ouabain binding of both SOL and EDL from dystrophic mice. Part of this reduction may be only apparent and due to an increase in connective tissue composition of dystrophic muscles. A limited study of muscles from neonate dystrophic mice indicated that abnormal [3H]ouabain binding was not present in EDL before two weeks of age.

Age Factors↗