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Biomedical subjects

S Thesleff

Publications and source records attributed to S Thesleff.

At least 73 records · Page 4Linked to original sources
PubMed1977

Protamine induced intracellular uptake of horseradish peroxidase and vacuolation in mouse skeletal muscle in vitro.

The uptake in vitro of horseradish peroxidase (HRP) in mouse skeletal muscle was examined by electron microscopy and chemical determination. In muscles exposed to an HRP solution for 60 min at +37 degrees C, HRP infiltrated the basal lamina of muscle fibres and caused an intense labelling of their sarcolemma. In addition HRP was found within the transverse tubules. Exposure to HRP for 30 min at +37 degrees C followed by HRP together with a polycationic protein (protamine) for 30 min at +37 degrees C caused an intracellular vesicular uptake of HRP. Intracellular HRP was found in numerous vesicles, membrane limited bodies and vacuoles. Protamine also induced focal autophagic vacuolation with progressive muscle fibre degeneration. An intracellular HRP uptake or muscle cell vacuolation could not be detected in the absence of protamine or when the incubation temperature was +4 degrees C. Chemical determination of HRP uptake was in general agreement with the morphological results. The uptake of HRP in the presence of protamine was stimulated at +37 degrees C and blocked at +4 degrees C. The results suggest that in skeletal muscle in vitro intracellular uptake of macromolecules occurs by endocytosis.

Animals↗
PubMed1977

Antagonism of the paralysis produced by botulinum toxin in the rat. The effects of tetraethylammonium, guanidine and 4-aminopyridine.

The injection of botulinum toxin type A into the hind-leg of adult rats causes complete paralysis of the leg lasting for several weeks. In the extensor digitorum longus (EDL) muscle transmitter release is reduced to a level of less than 1% of normal. Tetraethylammonium (TEA) and guanidine in concentrations of about 3 mM restore, in EDL muslces in vitro, neuromuscular transmission to about the normal level, provided that the external calcium concentration is 4 mM or higher. 4-Aminopyridine (4-AP) has similar restorative effect but is about 20-30 times more potent. Unlike TEA and guanidine, 4-AP is effective when the ambient calcium concentration is 2 mM; this drug is therefore also active in vivo. The intravenous injection of 4-AP (5 mg/kg body weight) restores neuromuscular transmission from complete paralysis by botulinum toxin to a normal level as shown by the recording of almost normal twitch and tetanic tensions in the EDL muscle. In rats paralysed by a lethal dose of botulinum toxin, the intraperitoneal administration of 4-AP restores general motor activity, the effect lasting 1-2 hours. A study of the effects of these drugs on spontaneous and evoked transmitter release suggests that all three compounds increase the level of free calcium inside the nerve terminals. In botulinum poisoning the transmitter release mechanism appears to be intact, but a reduced sensitivity to calcium has been shown (Cull-Candy et al. 1976), and this could explain why the drugs restore evoked transmitter release in botulinum poisoning.

Action Potentials↗
PubMed1976

Taipoxin, an extremely potent presynaptic neurotoxin from the venom of the australian snake taipan (Oxyuranus s. scutellatus). Isolation, characterization, quaternary structure and pharmacological properties.

Taipoxin (taipan toxin), purified from the venom of the Australian taipan (Oxyuranus s. scutellatus) by gel filtration on Sephadex G-75 followed by column zone electrophoresis, is the most lethal neurotoxin yet isolated from any snake venom. The LD50 is 2 mug/kg in the mouse. The main physiological effect is a gradual reduction to complete stop of evoked and spontaneous release of acetylcholine from motor nerve terminals. Intoxicated animals die of asphyxia caused by neuromuscular blockage of the respiratory muscles. Taipoxin is a moderately acidic sialo-glycoprotein (pI 5) with a molecular weight of 45 600 as calculated from composition data or 46 800 as determined by meniscus depletion sedimentation equilibrium. Taipoxin is a 1:1:1 ternary complex of subunits designated alpha, beta and gamma which dissociate completely at low pH and high ionic strength or in 6 M guanidine hydrochloride. The dissociation by guanidine at neutral pH is reversible, while the acid-induced dissociation is not . The alpha and beta components consist of 120 amino acid residues cross-linked by seven disulfide bridges, whereas the gamma component has 135 residues and eight disulfides. The very basic (pI Greater than 10) alpha component contains 13 residues of arginine and is the only subunit displaying lethal neurotoxicity (mouse LD50 approximately 300 mug/kg). The neutral beta fraction was separated by ion-exchange chromatography into two iso-component, beta 1 and beta2, which differ slightly in amino acid composition. The very acidic gamma component contains all of the carbohydrate, which includes 4-5 residues of sialic aid. The three subunits are homologous in sequence although the gamma component is eight residues longer on the N-terminus and must also contain extra amino acids elsewhere.

Acetylglucosamine↗
PubMed1976

Motor end-plates in regenerating rat skeletal muscle exposed to botulinum toxin.

Botulinum toxin type A, when applied to the extensor digitorum longus muscle of rats, reduces spontaneous and evoked transmitter release to a few per cent of the normal level. The toxin, however, fails to affect the morphological redifferentiation of the postsynaptic part of the neuromuscular junction following muscle degeneration-regeneration induced by a selective myotoxic agent, bupivacaine. This indicates that neither transmitter release nor resulting muscle activity are necessary for the differentiation of the sole plate.

Anesthetics, Local↗
PubMed1976

The effects of taipoxin and notexin on the function and fine structure of the murine neuromuscular junction.

The isolated neurotoxins taipoxin and notexin from the venoms of the Elapidae, Oxyuranus scutellatus and Notechis scutatus scutatus respectively cause a neuromuscular block when administered to the mouse in vivo or to the phrenic nerve-hemidiaphragm preparation in vitro. The block is preceded by a latency period during which the toxins bind irreversibly to the nerve. The period is shortened by nerve activity. The frequency of the miniature end-plate potentials is gradually reduced, almost to zero, and their amplitude distribution is altered; small and very large miniature endplate potentials appearing. Ultrastructurally the endplates are altered in the presynaptic portion but not in the postsynaptic part. In an early stage of poisoning the axolemma has an increased number of omega-shaped indentations similar in size to synaptic vesicles. At a later stage, when the animals die of respiratory paralysis, the axolemmal indentations are more numerous and the synaptic vesicles greatly reduced in number, the remaining vesicles having a variable and frequently larger than normal size. When impulse activity in the phrenic nerve is stopped by cutting the nerve before the administration of toxin there is no reduction in the number of synaptic vesicles, only the appearance of an increased number of axolemmal indentations. It is suggested that taipoxin and notexin irreversibly interfere with the formation of synaptic vesicles by arresting vesicle membrane recycling at the level of the axolemma. When the pre-existing store of vesicles is depleted, by nerve activity, a neuromuscular block results.

Acetylcholine↗
PubMed1976

Spontaneous activity in denervated mouse diaphragm muscle.

Intracellular electrodes were used to study the discrete depolarizations which trigger fibrillation potentials in chronically denervated mouse diaphragm muscles. Provided that the muscles were perfused on both sides spontaneous activity was maintained in vitro. 2. Discrete spontaneous depolarizations, present only in the centre of the muscle, were recorded from the third day of denervation reaching a maximum in prevalence 9-12 days after sectioning the nerve. These potentials had random occurrence and nearly constant amplitude and frequency within a fibre, dependence of amplitude and frequency on membrane potential, and low temperature dependence. 3. The spontaneous activity was enhanced and could be initiated in previously quiescent fibres by lowering the external Ca concentration. The activity was reduced by increasing external Ca and was abolished at 15mM-[Ca] 0. Tetrodotoxin (10-(7)M) blocked spontaneous activity. 4. The spontaneous activity was enhanced by the catecholamines isoprenaline and adrenaline (0.5-10 mug/ml.). This effect of isoprenaline was accompanied by an increase in the rate of rise and the amount of overshoot of the action potential. 5. Ouabain (10-(6)-10-(4)M) of K+-free solutions reversibly blocked spontaneous activity. Ouabain (10-(4)M) reduced the rate of rise and the amount of overshoot of the action potential. 6. Detubulation of muscle fibres with glycerol of the presence of hypertonic solutions abolished spontaneous activity which could not be restarted by reducing Ca or by the addition of isoprenaline. 7. The results support the suggestion that the spontaneous discrete depolarizations which give rise to fibrillation potentials in denervated muscle result from regenerative sodium conductance increases within the transverse tubular system of the muscle fibres. Catecholamines and ouabain could affect this activity either directly, through an action on membrane excitability, or indirectly via the Na+-K+ pump.

Action Potentials↗
PubMed1976

Effects of botulinum toxin on neuromuscular transmission in the rat.

1. Botulinum toxin (BoTx) type A partially blocks spontaneous transmitter release from nerve terminals in the rat. Minature end-plate potentials (m.e.p.p.s) are present at all end-plates, initially with a low frequency but increasing with time after posoning. Their amplitude distribution is at first skew with a predominace of very small m.e.p.p.s but, after a few days, larger than normal m.e.p.p.s appear. 2. Tetanic nerve stimulation, Black Widow Spider Venom, the Caionophore A 23187 or mechanical damage to nerve terminals increases the frequency of m.e.p.p.s and alters the amplitude distribution of m.e.p.p.s towards a normal Gaussian one; the m.e.p.p. size approaches that seen at normal end-plates. This was seen at any time after poisoning. 3. Nerve stimulation gives rise to end-plate potentials (e.p.p.s) of low amplitude and high failure rate. Statistical analysis indicates that evoked release is quantal in nature and follows Poisson statistics, quantum size being initially very small, but after a few days approaching normal size. Short-term tetanic nerve stimulation reversibly increases the quantum content of e.p.p.s and during early stages of paralysis long-term (2 hr) stimulation causes an apparently permanent increase in quantum size. 4. Raising the extracellular Ca concentration from 2 to 16 mM increases the frequency of m.e.p.p.s in normal muscle but not in BoTx poisoned ones. K-free medium or ouabain, which are believed to raise the intracellular Ca concentration in nerve terminals, similarly increases m.e.p.p. frequency in normal but not in poisoned muscles. When the Ca-ionophore A 23187 is used together with high extracellular Ca (greater than 4 mM) massive release of transmitter occurs from poisoned terminals. 5. The extracellular Ca concentration which causes a certain level of transmitter release in reponse to nerve impulses is considerably higher at BoTx poisoned end-plates than at normal ones. The slope value for Ca dependence of transmitter release is about 1-5 compared with about 3 at normal end-plates. 6. Tetraethylammonium (TEA) greatly increases the amount of transmitter released by nerve impulses and restores neuromuscular transmission during all stages of poisoning, although it has not effect on spontaneous transmitter release. In the presence of TEA the power relation between Ca concentration and quantum content at the BoTx poisoned end-plate is similar to that seen at normal end-plates. 7. It is suggested that in BoTx poisoning the mechanism for transmitter release has a reduced sensitivity to Ca, and the level for activation by intracellular Ca is elevated. Once the intracellular concentration of Ca is raised to this level, by tetanic nerve stimulation, mechanical injury to nerve terminals, the Ca-ionophore or the prolongation of the nerve action potential with TEA, augmented transmitter release occurs, similar to that which occurs in normal nerve terminals at a lower level of Ca.

Animals↗
PubMed1975

Guanidine and neuromuscular transmission. I. Effect on transmitter release occurring spontaneously and in response to single nerve stimuli.

The effect of guanidine on neuromuscular transmission was studied in human intercostal muscle and mouse diaphragm preparations in vitro. Guanidine greatly increased the number of acetylcholine (ACh) quanta released by a single motor nerve action potential. This effect of guanidine was greater at junctions with a low quantum content. The spontaneous release of ACh quanta was not substantially changed by guanidine. No change was found in the postsynaptic sensitivity to ACh released from the motor nerve or iontophoretically applied to the muscle fiber. Effects of the drug had slow onset and were very long-lasting and resistant to wash.

Acetylcholine↗
PubMed1975

Guanidine and neuromuscular transmission. II. Effect on transmitter release in response to repetitive nerve stimulation.

The effect of guanidine on the neuromuscular transmission in human intercostal and mouse diaphragm muscle in vitro during repetitive nerve stimulation was studied. The drug greatly increased the release of acetylcholine (ACh) quanta by nerve impulses at low frequencies of nerve stimulation and at the beginning of tetani at high frequencies of stimulation. The effect was shown to be produced by an increase in fractional release from an unchanged store of ACh quanta available fro immediate release. This seems to explain why guanidine has a poor therapeutic effect in myasthenia gravis but a good effect in the myasthenic syndrome.

Acetylcholine↗
PubMed1975

Studies on the mechanism of fibrillation potentials in denervated muscle.

1. Intracellular electrodes were used to study the origin of fibrillation potentials in chronically denervated rat muscle. 2. Fibrillation potentials were observed to start from spontaneous biphasic membrane potential oscillations. Each action potential was followed by an after-hyperpolarization which in turn served as a pre-potential for the next spike. The critical level (threshold) for the initiation of the first spike in a train was lower than that of the next and subsequent spikes. 3. A correlation was found between the level of membrane potential and the critical level for action potential generation. This relation was most marked around the resting membrane potential (minus 60 to minus 80 mV) where 10 mV hyperpolarization caused a 9 m V increase in the critical potential level. At higher membrane potentials the correlation was less pronounced. In innervated muscles a similar correlation existed but it was less marked and was present only at membrane polarizations below the resting potential. 4. Increasing the external calcium concentration from 2 to 8 mM reduced the membrane potential-critical level relationship in denervated fibres towards that of innervated ones. 5. As critical level changes with membrane hyperpolarization, the rate of rise of the action potential increased, suggesting a progressive removal of sodium inactivation. 6. It is suggested that a mechanism similar to anode break excitation is important for the induction and maintenance of fibrillation potentials.

Action Potentials↗