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D G Stephenson

Publications and source records attributed to D G Stephenson.

105 records · Page 6Linked to original sources
PubMed1986

Calcium and strontium activation of single skinned muscle fibres of normal and dystrophic mice.

Differences in contractile activation by Ca2+ and Sr2+ between various types of normal and dystrophic murine muscle fibres were investigated using mechanically skinned fibres derived from soleus and extensor digitorum longus (e.d.l.) muscles of normal and dystrophic mice of strain 129ReJ. In terms of contractile activation, the normal e.d.l. muscle was found to consist of one relatively homogeneous population of muscle fibres characterized by steep force-pCa and force-pSr curves, low sensitivity to Ca2+ and very low sensitivity to Sr2+. Normal soleus muscles contained two fibre populations of similar size which could be distinguished on the basis of their contractile activation properties. The first fibre population was characterized mainly by its shallow force-pCa and force-pSr curves, high Ca2+ sensitivity, high Sr2+ sensitivity and the occurrence of large, slow force oscillations of myofibrillar origin. The second fibre population was characterized by force-pCa and force-pSr curves of steepness intermediate between those of normal e.d.l. and those of the first fibre population of normal soleus, by faster myofibrillar force oscillations and by low sensitivity to Ca2+ and Sr2+. The dystrophic e.d.l. fibre population had contractile characteristics which were distinct from those of the three types of normal fibre populations. However, some characteristics of the dystrophic e.d.l. fibres were very similar to those of the normal e.d.l. fibre population. Of all the fibre types investigated, dystrophic e.d.l. fibres were the least sensitive to Ca2+. Dystrophic soleus muscle contained a single homogeneous population of fibres which shared some common contractile activation characteristics with both of the fibre populations present in normal soleus muscle. However, of all fibre types investigated, the dystrophic soleus fibres were the most sensitive to Ca2+. Because of this characteristic, these fibres formed a distinct population. The maximum tensions induced by Ca2+ and Sr2+ were usually smaller in dystrophic fibres than in normal fibres obtained equivalent muscles. In conclusion, various normal murine muscle fibre types can be identified on the basis of differences in the mechanism of force activation by Ca2+ and Sr2+. Furthermore, it is possible to detect significant physiological differences in the mechanism of force activation brought about by murine muscular dystrophy.

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PubMed1986

Activation of the contractile apparatus of skinned fibres of frog by the divalent cations barium, cadmium and nickel.

The contractile apparatus of mechanically skinned muscle fibres of frog can be reversibly activated by Ba2+ and Cd2+. The maximum force induced by both Ba2+ and Cd2+ is the same as that induced by Ca2+ and Sr2+. The ionic concentrations of the divalent cations required to induce 50% of the maximum activated force at 1 mM-Mg2+, pH 7.10, 22 degrees C and 250 mM ionic strength are about 8 X 10(-7) M for Ca2+, 5 X 10(-6) M for Cd2+, 2.6 X 10(-5) M for Sr2+ and 7 X 10(-4) M for Ba2+. Exposure of the skinned fibre to relatively low Ni2+ concentrations (between 10(-6) and 10(-5) M) resulted in a transient force response accompanied by an irreversible change in the ability of the preparation to develop force. The Ba2+- and Cd2+-activation curves are considerably flatter than the corresponding curves for Ca2+ and Sr2+. An increase in Mg2+ concentration from 1 to 3 mM decreased the sensitivity of the contractile apparatus to Ba2+ by a factor of about 1.5 without affecting the maximum force response. The Ca2+-activation curve was modified in the presence of subthreshold concentrations of Ba2+ and the results indicate that Ba2+ could have both an activating and an inhibitory action on the Ca2+-activated force. A kinetic model which can quantitatively explain the results for activation of contraction by Ba2+ and Ca2+, is described.

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PubMed1986

The influence of free calcium on the maximum speed of shortening in skinned frog muscle fibres.

The influence of [Ca2+] on the maximum velocity of shortening (Vmax) was examined in mechanically skinned Rana pipiens and Rana temporaria fibres using improved force clamps and the slack test techniques. All measurements were made at 7.5 degrees C. At low relative loads (P/P0 less than 0.1), maximally activated R. pipiens fibres shortened more rapidly than did submaximally activated fibres. At higher relative loads, however, little difference in the speed of shortening was observed. Vmax (determined by the slack test) of R. pipiens fibres increased as the level of activation increased. Over sarcomere lengths 1.8-2.1 microns it was 2.28 muscle lengths/s (m.l./s) (S.E. of mean +/- 0.25, n = 5) at 20-35% activation, 2.89 m.l./s (+/- 0.22, n = 7) at 40-60% activation, and 4.18 m.l./s (+/- 0.25, n = 6) at 100% activation. At longer sarcomere lengths (2.2-2.6 microns), higher Vmax values were observed at all levels of activation, but the influence of Ca2+ on Vmax persisted. Vmax was 3.54 m.l./s (+/- 0.41, n = 4) at 20-30% activation and 5.15 m.l./s (+/- 0.22, n = 5) at 100% activation. In R. temporaria fibres, Vmax (determined by force clamps over sarcomere lengths 1.8-2.1 micron) also increased as the level of activation increased, from 3.47 m.l./s (+/- 0.06, n = 6) at 13-29% activation to 5.62 m.l./s (+/- 0.17, n = 6) at 100% activation. Vmax was also determined (using the slack test) in mechanically and chemically skinned rabbit soleus fibres. Vmax at 15 degrees C (1.05 m.l./s, +/- 0.11, n = 5) at full activation decreased by more than 3-fold as the level of activation was reduced to 10%. We conclude that the level of activation influences the Vmax of skinned skeletal muscle fibres. This has now been demonstrated in three different preparations and by a variety of techniques. This effect is most pronounced at low relative loads, and might not be observed if there are experimental limitations which prevent making velocity measurements at low relative loads.

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PubMed1985

Temperature-dependent calcium sensitivity changes in skinned muscle fibres of rat and toad.

Single mechanically skinned muscle fibres of different types (fast- and slow-twitch mammalian; slow and twitch amphibian) were successively activated in solutions of various Ca2+ concentrations at different temperatures. An increase in temperature from 5 to 22 degrees C reversibly shifted the isometric steady-state force-pCa curves towards higher Ca2+ concentration for individual fibres of each of the muscle types. A further increase in temperature to 35 degrees C in mammalian fibres resulted in an additional decrease in Ca2+ sensitivity. The temperature dependence of Ca2+ sensitivity was greater in the 'faster' fibre types: fast-twitch greater than slow-twitch; twitch greater than slow. The maximum isometric force response, P0, of both rat and toad skinned fibres was found to be strongly dependent on temperature below 22 degrees C. No detectable force could be induced by Ca2+ in mammalian muscle fibres at 0-1 degree C while in toad fibres P0 decreased by about 90% when temperature dropped from 20 to 0 degree C. Since in mechanically skinned fibres of other amphibians (Bufo bufo, Rana species) P0 is only marginally affected it is likely that the P0-temperature relations are indicative of the range of temperature over which the muscles are normally functional. The P0-temperature relations of skinned muscle fibres closely resembled the P0-temperature relations of tetanically stimulated intact muscle preparations from the same species of animals suggesting that the contractile apparatus is mainly responsible for the variation of force response with temperature in intact muscle.

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PubMed1983

Slow amphibian muscle fibres become less sensitive to Ca2+ with increasing sarcomere length.

Here we report that mechanically skinned muscle preparations from slow amphibian muscle fibres become less sensitive to Ca2+ when the sarcomere length is increased beyond the value corresponding to optimum filament overlap. This effect is opposite to that observed in all other studies on vertebrate myofibrillar preparations from twitch (Endo, 1972a; 1972b; Moisescu & Thieleczek, 1979; Stephenson & Williams, 1982) and cardiac muscles (Fabiato & Fabiato, 1978). This finding shows that stretch-induced increased in Ca2+ sensitivity is not a general property of the contractile apparatus and suggests that differences in the ultrastructure between these muscle fibre types could be responsible for the opposite length effects. Furthermore, these results allow a more detailed understanding of the processes behind the stretch effects observed in intact slow and twitch amphibian muscle fibres (Lännergren, 1975).

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PubMed1983

Effects of caffeine on Ca-activated force production in skinned cardiac and skeletal muscle fibres of the rat.

Skinned fibres prepared by mechanical and/or chemical means from cardiac and skeletal muscles of the rat were activated in solutions strongly buffered for Ca2+ (with 50 mM EGTA) in the absence or presence of caffeine 5-40 mM. In all preparations caffeine was found to reversibly shift the relation between steady-state force and free [Ca2+] toward lower free [Ca2+] in a dose dependent manner. This increase in apparent Ca2+ sensitivity was not antagonized by procaine and was the same, within each muscle type, irrespective of the manner in which the skinned fibre was prepared, and consequently the degree to which it retained cellular membranes. The effect was more pronounced in cardiac and slow-twitch than in fast-twitch, myofibrillar preparations. At equivalent concentrations theophylline mimicked this effect of caffeine in all preparations, however, addition of exogenous cyclic AMP neither mimicked nor modified, in any way, the effect. Maximum Ca2+ -activated force production was not affected by caffeine below 20 mM but was depressed by concentrations of 20 mM and above. The increase in apparent Ca2+ sensitivity produced by caffeine can not be the result of a mobilization of some cellular store of Ca2+ but must arise from a direct effect of caffeine on the myofilaments which leads to a change in the apparent affinity constant of the force controlling sites for Ca2+.

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PubMed1982

Effects of sarcomere length on the force-pCa relation in fast- and slow-twitch skinned muscle fibres from the rat.

1. Steady-state force-pCa relations were determined for mechanically skinned fibres of fast- and slow-twitch rat skeletal muscle, extensor digitorum longus (e.d.l.) and soleus respectively, at varied sarcomere lengths and temperatures (3-35 degrees C).2. This relation was different for the two fibre types at all sarcomere lengths, with soleus fibres having lower Ca(2+) concentration threshold for activation and requiring a wider Ca(2+) concentration range to reach force saturation.3. An increase in sarcomere length within the range 2.2-3.6 mum reversibly shifted the steady-state force-pCa curves for both fast- and slow-twitch fibres toward lower Ca(2+) without affecting the steepness of the curves.4. The results are consistent with the idea that the sensitivity of the contractile apparatus to Ca(2+) is increased with increasing sarcomere length.5. The change in apparent Ca(2+) sensitivity (measured as the change in pCa corresponding to 50% Ca-activated force (DeltapCa(50))) was approximately proportional to the change in sarcomere length (DeltaSL) at all temperatures for both types of muscle fibre.6. The proportionality coefficient (DeltapCa(50)/DeltaSL) was higher for slow-twitch fibres than for fast-twitch fibres by a factor of 1.5-2.0 at all temperatures investigated.7. The force-pCa relation was less affected by changes in sarcomere length at lower (3-5 degrees C) than at higher temperatures (22-35 degrees C).8. The sarcomere length-maximum force relation obtained with skinned fibres for sarcomere lengths between 2.10 and 4.05 mum was similar for both fibre types at all temperatures despite large temperature dependent variations in maximum Ca-activated force response. Maximum force decreased linearly to zero between ca. 2.8 and 3.95 mum; remained high and essentially constant between ca. 2.40 and 2.80 mum, and decreased significantly below 2.40 mum.9. Myofilament-generated tension oscillations produced by partially activated soleus skinned fibres are highly length dependent, disappearing completely when sarcomere length is increased above 3.15 mum.

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PubMed1981

Studies on the luminescent response of the Ca2+-activated photoprotein, obelin.

1. The kinetics and stoicheiometry of the Ca2+-activated luminescent reaction of the photoprotein obelin were studied at different temperatures and in the presence of various substances, including the physiologically occurring cations K+, Na+, Ca+, Mg2+ and H+. 2. The results suggest Ca2+-independent rates of rise and fall in obelin luminescence following sudden changes in [Ca2+] and indicate that changes in [Ca2+] over the range 1 x 10(-6) - 3 x 10(-4) M are followed significantly faster by the obelin response (approx. 3 ms delay at 20 degrees C) than by the aequorin response (approx. 10 ms delay at 20 degrees C). 3. Obelin was found to emit low-intensity light (less than 10(-6) of the maximum Ca2+-activated response), which was independent of Ca2+ at concentrations below about 10(-7) M. The level of this Ca2+-independent light emission is sensitive to temperature and the ionic composition of the solution. 4. The log-log plot of light intensity against ionized Ca indicates a maximum slope of 2.5, suggesting the involvement of three Ca ions in the luminescent reaction. 5. Increase in the concentration of K+, Na+, Mg2+ and H+ generally shift the Ca2+ activation curve for obelin toward higher Ca2+ concentrations. These cations can also affect the maximum rate of obelin utilization at more extreme concentrations. 6. The maximal rate of obelin utilization was also affected to varying degrees by the presence of uncharged substances such as glucose, sucrose and polyvinylpyrrolidone. However, neither the sensitivity of obelin to Ca2+ nor the quantum yield were modified by the substances. 7. Caffeine (less than 20 mM), procaine (less than 20 mM) and sodium dantrolene (saturated solution), substances known to modify cellular Ca2+ movements, had little effect on the Ca2+-induced luminescent reaction. The general anaesthetics chlorpromazine and halothane appeared to lower greatly the quantum yield without, however, modifying the maximum rate of obelin utilization. 8. A scheme of reaction for obelin activation by Ca2+ is presented which adequately explains the experimental observations and allows one to make accurate predictions regarding the relative obelin response under a variety of ionic conditions at room temperature.

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PubMed1981

Non-uniform ion distributions and electrical potentials in sarcoplasmic regions of skeletal muscle fibres.

We report here electrophysiological and ion distribution results obtained with mechanically skinned skeletal muscle fibres which indicate that a Donnan equilibrium, characterized by large ionic concentration and electrical potential differences, is established between the myofibrillar space and the rest of the sarcoplasm surrounding the myofibrils. The more negative electrical potential within the myofibrils (-15 to -29 mV for conditions similar to those in vivo) can strongly influence: (1) intracellular Ca2+ movements associated with contraction; (2) the location of sarcoplasmic enzymes, their substrates and products of reaction; (3) measurements of the electrical potential difference between the extra- and intracellular space and estimation of relative membrane permeabilities for certain ions.

Animals↗
PubMed1981

Calcium-activated force responses in fast- and slow-twitch skinned muscle fibres of the rat at different temperatures.

1. Force responses from mechanically skinned fibres of rat skeletal muscles (extensor digitorum longus and soleus) were measured at different temperatures in the range 3-35 degrees C following sudden changes in Ca2+ concentration in the preparations. 2. At all temperatures there were characteristic differences between the slow- and fast-twitch muscle fibres with respect to the relative steady-state force-[Ca2+] relation: such as a lower [Ca2+] threshold for activation and a less steep force-pCa curve in slow-twitch muscle fibres. 3. At 3-5 degrees C the force changes in both types of muscle fibres lagged considerably behind the estimated changes in [Ca2+] within the preparations and this enabled us to perform a comparative analysis of the Ca2+ kinetics in the process of force development in both muscle fibre types. This analysis suggest that two and six Ca2+ ions are involved in the regulatory unit for contraction of slow- and fast-twitch muscle fibres respectively. 4. The rate of relaxation following a sudden decrease in [Ca2+] was much lower in the slow-twitch than in the fast-twitch muscle at 5 degrees C, suggesting that properties of the contractile apparatus could play an essential role in determining the rate of relaxation in vivo. 5. There was substantial variation in Ca2+ sensitivity between muscle fibres of the same type from different animals at each temperature. However the steepness of the force-[Ca2+] relation was essentially the same for all fibres of the same type. 6. A change in temperature from 5 to 25 degrees C had a statistically significant effect on the sensitivity of the fast-twitch muscle fibres, rendering them less sensitive to Ca2+ by a factor of 2. However a further increase in temperature from 25 to 35 degrees C did not have any statistically significant effect on the force-[Ca2+] relation in fast-twitch muscle fibres. 7. The effect of temperature on the Ca2+ sensitivity of slow-twitch muscle fibres was not statistically significant, mainly because of the large variation in sensitivity amongst these preparations at room temperature. 8. Two types of oscillatory processes not associated with intracellular membranes were observed in the force response of all slow-twitch muscle fibres when submaximally activated (less than 60% maximum force) at 25 and 35 degrees C, but never at 3-5 degrees C. The frequency of oscillations increased with temperature. 9. Maximum Ca2+-activated force in both muscle fibre types was greatly dependent upon temperature over the range 0-25 degrees C, but increased only slightly above 25 degrees C. 10. Experiments on the rigor state suggest that the number of possible actomyosin interacting sites diminishes considerably as temperature is decreased below 25 degrees C.

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PubMed1980

Different isometric force - [Ca2+] relationships in slow and fast twitch skinned muscle fibres of the rat.

Skinned muscle fibres prepared from fast and slow twitch muscles of rat have been activated in Ca2+-buffered solutions using a new activation procedure (Moisescu, D.G. and Thieleczek, R. (1978) J. Physiol. 275, 241--262). The results indicate that (i) the Ca2+ activation curve is less steep for slow fibres, (ii) physiologically relevant force levels are attained considerably faster at constant [Ca2+] in fast fibres, and (iii) active force becomes noticeable at lower [Ca2+], but reaches saturation at higher [Ca2+] for slow fibres.

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PubMed1980

Activation of skinned arthropod muscle fibres by Ca2+ and Sr2+.

Mechanically skinned skeletal muscle fibres of three crustaceans (barnacle, crayfish and crab) and two insects (cockroach and cricket) were activated in Ca2+- and Sr2+-buffered solutions of different concentrations and the isometric force response was determined. The maximum force response induced by Sr2+ (P0Sr) was only 0-10% of that induced by Ca2+ (P0Ca) in all crustacean muscles, but approached 90% in insects. Experiments on barnacle muscle fibres activated simultaneously by Ca2+ and Sr2+ suggested that Sr2+ competes with Ca2+ for binding onto the regulatory sites without, however, being able to turn all of them 'on' as efficiently as Ca2+. Interestingly, the ratio P0Sr/P0Ca and the sensitivity for both Sr2+ and Ca2+ increased substantially after 4-6 h following the dissection of the animals in most intact decapod muscle fibres and after 24 h in most barnacle muscle fibres. The steepness of the activation curves for both Ca2+ and Sr2+ was similar for each muscle regardless of the age of the fibre and implied that more than 2 Ca2+ (2 Sr2+) were involved in the activation process of each muscle. A Ca2+-induced Ca2+ release mechanism of physiological importance was found to operate in all arthropod muscle fibres investigated.

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PubMedDate not supplied

Molecular cogs in machina carnis.

1. This review explores the complexity of skeletal muscle function mainly from the perspective of work performed by the author over the past two decades.

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