Effects of spinal cord and other tissue extracts on resting and action potentials of organ-cultured mouse skeletal muscle

doi:10.1016/0006-8993(77)90095-6

Brain Research

Volume 119, Issue 1, 1 January 1977, Pages 133-140

https://doi.org/10.1016/0006-8993(77)90095-6 Get rights and content

Summary

The trophic effects of spinal cord extract on resting potential, action potential and sensitivity of action potential to tetrodotoxin (TTX) in organ-cultured mouse skeletal muscle were examined. Spinal cord extract blocked decrease in the maximal rates of rise and fall of the action potential caused by cultivation and also partially prevented decrease in its TTX-sensitivity and in the resting potential. In another experiment, spinal cord extract was added to organ cultures of skeletal muscle that had been previously denervated for 3 days in vivo. Spinal cord extract partially reversed post-denervation fall in the maximal rate of rise of action potential and its TTX-sensitivity. Extracts of non-nervous tissues (kidney, spleen, pancreas and adrenal gland) were ineffective.

These results indicate that mouse spinal cord appears to possess neurotrophic substance(s) regulating membrane properties of mouse skeletal muscle.

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The slow hyperpolarizing afterpotential (slow HAP) following an evoked action potential in cultured rat muscle cells is inhibited by coculture with spinal cord neurons, spinal cord-conditioned medium (CM) and by a low-molecular-weight fraction (<4000 Da) prepared from this CM. The incidence, amplitude and duration of the slow HAPs decreased significantly after 24 h of incubation. In contrast, chick retinal neuron cocultures or CM did not alter the slow HAP.
Trophic action of nerve extract on denervated skeletal muscle in vivo: Dose dependency, species specificity, and timing of treatment
1983, Experimental Neurology
Atrophy in a denervated muscle results from the disuse caused by paralysis of the muscle, and from the loss of special neurotrophic substances. Proteins extracted from rats' sciatic nerves have been shown to prevent the nondisuse atrophy of denervated rats' extensor digitorum longus (EDL) muscles when administered by intramuscular injection. The present investigation elicited more information about this neurotrophic phenomenon. Extract of liver failed to affect the rate of atrophy of denervated EDL muscles. Injection of various doses of extract of rats' or sheep's sciatic nerves indicated that the action of the extract was dose-dependent but not species-specific. Muscles denervated for 14 days and injected with extract on all 14 days or only on the first or last 7 days all exhibited significantly less atrophy than those that were not injected.
Molecular aspects of the trophic influence of nerve on muscle
1983, Progress in Neurobiology
Neurotrophic substance develops tetrodotoxin-sensitive action potential and increases curare-sensitivity of acetylcholine response in cultured rat myotubes
1981, Developmental Brain Research
We examined the trophic effects of a partially purified trophic substance from mouse spinal cord extract on the tetrodotoxin (TTX)-sensitivity of action potentials and on acetylcholine-sensitivity of rat skeletal myotubes in 7- and 8-day-old cultures. Many myotubes grown in control medium generate action potentials in the presence of TTX (10⁻⁶ M). The addition of fraction E (Fr.E) from a Biogel P2 column, which exhibited trophic activity on adult denervated muscle in organ culture, decreased TTX-resistivity of action potentials of myotubes in cell culture. The trophic substance was also effective when further purified by paperchromatography and electrophoresis. The response to iontophoretically applied acetylcholine of Fr.E-treated myotubes was mure more reduced by d-tubocurarine (10⁻⁷) g/ml) than those of control cultured myotubes. No difference in morphological differentiation, protein synthesis, creatine phosphokinase activity or specific binding of [¹²⁵I]α-bungarotoxin was observed between control and Fr.E-treated cultures. These results suggest that the trophic substance in Fr.E may be involved in the normal development of TTX-sensitive sodium channels and of acetylcholine receptor properties.
Neuronal maturation in mammalian cell culture is dependent on spontaneous electrical activity
1981, Brain Research
Fetal mouse spinal cord (SC) and dorsal root ganglion (DRG) neurons undergo a process of maturation in cell culture lasting a month or more. We have investigated the role of electrical activity in this maturational process with the use of tetrodotoxin (TTX), the specific blocker of the voltage-sensitive sodium channel responsible for action potential generation. This agent completely eliminates the spikes and related synaptic activity which occur abundantly in untreated cultures. Such blockade of electrical activity in the cultures, when begun early (day 1 or day 8 in vitro), results in a 85–95% reduction in the number of large SC neurons, without affecting DRG neuron numbers. TTX treatment initiated when cultures are mature (day 70) has no significant effect on either DRG or SC neurons. Intermediate effects are obtained when treatment is initiated at day 35 in vitro. The activity of the nerve-specific enzyme, choline acetyltransferase, is significantly decreased by early TTX treatment, while DNA and protein content of the cultures (primarily contributed by glial and fibroblastic cells) is not affected.
Partial purification and characterization of neurotrophic substance affecting tetrodotoxin sensitivity of organ-cultured mouse muscle
1979, Brain Research
From mouse spinal cord homogenate, we isolated a trophic substance which reverses the post-denervation decrease in tetrodotoxin sensitivity of action potential in organ-cultured extensor digitorum longus muscle of mouse and characterized its physicochemical properties. The trophic substance was separated from macromolecules in homogenate by gel filtration on Biogel P2 column. The partially purified trophic substance was heat-stable, acid-stable and alkaline-labile. The trophic activity was destroyed by lyophilization at neutral pH but not at acidic pH. The trophic activity was abolished by incubation with pronase or leucine aminopeptidase, but not by trypsin, chymotrypsin, thermolysin or carboxypeptidase A. The trophic substance passed through an ultrafiltration membrane UM10 freely. A small part of the trophic activity passed through a UM2 or UM05, and the rest was retained on the membranes. The trophic substance adsorbed on CM-Sephadex at pH 7.2 but passed through DEAE-Sephadex at pH 8.4.
These results suggest that the trophic substance regulating tetrodotoxin sensitivity of action potential in mouse skeletal muscle is a peptide with a rather low molecular weight of less than 10,000 and that while the N-terminus of the peptide is free, the C-terminus is probably blocked. This peptide differs from other trophic substances reported previously by other investigators.

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Effects of spinal cord and other tissue extracts on resting and action potentials of organ-cultured mouse skeletal muscle

Summary

Exp. Neurol.

Europ. J. Pharmacol.

Brain Research

Exp. Neurol.

J. biol. Chem.

Exp. Neurol.

Europ. J. Pharmacol.

A comparative study of membrane properties of innervated and chronically denervated fast and slow skeletal musclesof the rat

Acta physiol. scand.

The pharmacology of batrachotoxin. IV. Interaction with tetrodotoxin on innervated and chronically denervated rat skeletal muscle

J. Pharmacol. exp. Ther.

Acetylcholine sensitivity of muscle fiber membrane: Mechanism of regulation by motoneurons

Science

Inhibition of denervation changes in skeletal muscle by blockers of protein synthesis

J. Physiol. (Lond.)