2-4-dithiobiuret and Paralysis

In previous studies, 2,4-dithiobiuret (DTB) caused a delayed onset neuromuscular weakness in rats which was associated with decreased quantal content, alterations in postsynaptic ion channel properties, and abnormalities in nerve terminal ultrastructure. The latter include features typical of degenerating or diseased motor endplates as well as a marked proliferation of smooth endoplasmic reticulum (SER), swelling of mitochondria and evidence for a decreased in intraterminal calcium concentrations at early stages of intoxication (Jones, 1989, Acta Neuropathol. 78:72). These in vivo studies do not allow us to distinguish between the initial effects of DTB on the nerve terminal and those evolving as a result of disuse or secondary to its action on the muscle fiber or Schwann cells. To begin to distinguish between primary and secondary effects of DTB, we examined DTB-treated rat PC12 cells for comparable changes. The direct effects of DTB on PC12 cells included signs of general toxicity. Cell death in sparsely- plated cultures increased from 8-9% in controls to 13.7% at 10 microM for 24 hr exposure, and continued to increase in a concentration-dependent fashion to 25% mortality at 25 microM. However, between 25 and 100 microM there was little additional increase in mortality. 10 to 40 microM DTB slightly decreased the ability of both differentiated and undifferentiated cells to adhere to a substrate. This effect was independent of cell mortality. In moderately-differ-entiated cells having processes up to 10 cell diameters and several varicosities, concentrations of DTB as high as those invoking increased cell mortality and comparable to those affecting the rat neuromuscular junction did not cause abnormalities in the structure of the SER. No masses of tubulovesicular profiles were seen with transmission electron microscopy, and large changes in the quantity or distribution were not detected at the light microscope with the fluorescent stains DiOC6 or rhodamine B. Other signs of neuronal degeneration (blebbing of the plasmalemma, large intracellular droplets, mitochondrial abnormalities) preceded or accompanied any evidence for abnormalities in the SER. Thus the effect of DTB on the SER at the rat motor nerve terminal may occur secondary to a more general toxic action on other cell types, or may be dependent on a level of neuronal activity not achieved in sparsely- plated cultures, or may require a greater degree of differentiation of the neuronal cells than p

Topics: Animals; Cell Adhesion; Cell Differentiation; Cell Survival; Endoplasmic Reticulum, Smooth; Glass; Models, Neurological; Nerve Growth Factors; Neurons; Neurotoxins; Paralysis; PC12 Cells; Polylysine; Rats; Thiourea

2,4-Dithiobiuret (DTB) causes a delayed-onset neuromuscular weakness when given chronically to rats. Mechanisms underlying this effect involve disruptions of acetylcholine (ACh) release and possibly effects on the ACh receptor channel complex. Previous experiments demonstrated a decrease in decay time constants for end-plate currents and miniature end-plate currents of muscles from rats exhibiting DTB-induced muscle weakness compared with those of controls. The purpose of the present study was to determine whether the alteration in rise and decay times for synaptic currents was due to direct effects of DTB on ACh receptor channels. Currents carried through single ACh-activated channels were recorded using patch voltage-clamp techniques in G8 mouse myotubes exposed to DTB in their growth medium and from intact hemidiaphragm preparations of rats treated with DTB by examining fluctuations in membrane noise during iontophoresis of agonist. Exposure of myotubes to DTB (1 or 10 microM) decreased the mean channel open time induced by suberyldicholine for short durations of exposure, whereas longer exposures (24-48 h) to DTB were required in order for decreased open times for ACh as an agonist to be observed. In the absence of DTB, closed times for single channels of G8 cells were described by a two-exponential fit reflecting intraburst and interburst closures. At 1 microM DTB, the duration of gaps within bursts and of gaps between bursts increased.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Animals; Cells, Cultured; Ion Channels; Iontophoresis; Male; Mice; Muscle, Skeletal; Paralysis; Rats; Rats, Sprague-Dawley; Receptors, Cholinergic; Thiourea; Time Factors

Daily treatment of rats with 2,4-dithiobiuret (DTB, 1 mg/kg/day i.p.) produces a flaccid neuromuscular weakness first observed in the hindlimbs after 5 to 6 days of treatment. This condition is characterized by diminished contractile strength following single shock and tetanic stimulation of the motor nerve, but no effect on contractions evoked by direct muscle stimulation, indicating an apparent impairment of motor axon conduction, junctional transmission or both. The purpose of the present study was to investigate further the neuromuscular depression caused by DTB using conventional intracellular microelectrode recording techniques. All experiments were conducted using the extensor digitorum longus muscle isolated from male rats treated for 6 to 7 days with 1 mg/kg/day i.p. of DTB or with 0.9% NaCl (1 ml/kg/day) as control. End-plate potentials (EPPs) and miniature end-plate potentials (MEPPs) were recorded from single junctions of DTB-poisoned or NaCl-treated paired controls. Muscles were transected ("cut muscle") to prevent contraction after peroneal nerve stimulation. EPP amplitude was decreased at the time of observable muscle weakness in DTB-treated rats. Endplate resting membrane potential was not affected. Decreased EPP amplitude was associated with a decrease in mean quantal content. Quantal content was depressed to an equivalent extent in DTB-treated rats when stimulus frequency was increased from 0.5 to 2, 5, 25 and 50 Hz. However, as the stimulus frequency was increased, preparations from DTB-treated rats were characterized by failures of nerve impulses to elicit an EPP.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Acetylcholine; Animals; Evoked Potentials; Male; Motor Endplate; Neuromuscular Junction; Paralysis; Rats; Rats, Inbred Strains; Thiourea

Urinary excretion of porphyrin precursors delta-aminolevulinic acid (ALA) and porphobilinogen (PBG) and total porphyrins was measured during intoxication of rats with 2,4-dithiobiuret (DTB), a chemical which produces delayed-onset neuromuscular weakness, in an attempt to ascertain whether or not DTB poisoning in the rat would serve as an animal model of the neurologic symptoms of acute intermittent porphyria. Daily administration of DTB (1 mg/kg/day, i.p.) produced flaccid skeletal muscle weakness first detected after 4 to 5 days of treatment. Onset of skeletal muscle weakness was associated with a significant increase in urinary excretion of ALA. The excretion of PBG and total porphyrin was also increased; however, the increase was not significant. The increase in porphyrins and porphyrin precursors was due to increased urine output which coincided with the onset of neuromuscular weakness; urinary concentrations of ALA, PBG, and porphyrins were not increased by DTB. Measurements of free-erythrocyte protoporphyrin, taken after 7 days of DTB treatment, indicated a significant elevation of free erythrocyte protoporphyrin concentration. The pattern of alterations in the heme precursors associated with DTB-induced paralysis in rats is quite different from that observed in humans afflicted with acute intermittent porphyria. Therefore, we conclude that DTB-induced paralysis in the rat does not represent an accurate animal model of acute intermittent porphyria.

Topics: Aminolevulinic Acid; Animals; Disease Models, Animal; Levulinic Acids; Male; Muscle Hypotonia; Neuromuscular Diseases; Paralysis; Porphobilinogen; Porphyrias; Porphyrins; Rats; Thiourea; Time Factors