Sebastian Conradi

Ultrastructure and synaptology of the initial axon segment of cat spinal motoneurons during early postnatal development.

SummaryThe initial axon segment (IS) of spinal α-motoneurons has been studied in single and serial sections during the first three postnatal weeks in the cat. It was found that boutons of different ultrastructural types made synaptic contact with the IS during the first postnatal week but during the second postnatal week the boutons disappeared. The diameter of the IS was not observed to change from the first to the second postnatal week but a significant increase was noted from the second to the third week. The length and general ultrastructural appearance of the IS did not change significantly over the postnatal period studied. The results are discussed in relation to some aspects of the functional development of spinal motoneurons during the early postnatal period.

Abnormal distribution of lead in amyotrophic lateral sclerosis ☆: Reestimation of lead in the cerebrospinal fluid

The lead concentration in CSF was determined by flameless atomic absorption spectrophotometry in 16 ALS patients and 22 control cases. The mean values were 0.69 +/- 0.55 (ALS) and 0.41 +/- 0.37 (controls), P < 0.01. This confirms our earlier findings of raised CSF lead levels in ALS but the present values are lower than previously reported for both ALS patients and controls.

Elevated levels of transcripts encoding a human retroviral envelope protein (syncytin) in muscles from patients with motor neuron disease.

Retroviral components of both exogenous and endogenous origins have been associated with nervous system diseases in both animals and humans. In the present study, the levels of transcripts from elements in the human endogenous retrovirus (HERV) W family were determined in muscle biopsies from patients with motor neuron disease (MND) and control subjects. Transcripts from the HERV‐W element on chromosome 7q21.2 encoding syncytin and from the SOD1 gene were detected at elevated levels in biopsies from the most affected muscles from MND patients compared to biopsies from control individuals. According to a recent study, syncytin is expressed in microglia in normal brain and can be up‐regulated in macrophages/microglia during inflammation. Although syncytin may have cytotoxic effects, it is therefore more likely that the present findings reflect a macrophage response in the muscles undergoing neurogenic atrophy than a primary pathogenetic event in MND.

Long-time penicillamine-treatment in Amyotrophic Lateral Sclerosis with parallel determination of lead in blood, plasma and urine

Six ALS‐patients were given long‐term penicillamine treatment. A rapid, long‐lasting increase in urinary lead was provoked, whereas whole blood and plasma lead were unchanged. In four patients, the progress did not change markedly, one patient deteriorated rapidly and in one patient the progression slowed down after one and a half years of treatment.

Further studies on the erythrocyte uptake of lead in vitro in amyotrophic lateral sclerosis (ALS) patients and controls

Following our earlier observations on increased plasma concentrations of lead in amyotrophic lateral sclerosis (ALS), the erythrocyte uptake of lead from plasma has been studied in vitro. Whole-blood from ALS patients and controls was incubated after the addition of lead (0.6 mumol/l whole-blood) and plasma lead concentrations were repeatedly determined. Incubation was continued until haemolysis developed. Fairly stable plasma lead concentrations were established at, on the average, 0.5-0.6 mumol/l after 10-30 min and persisted throughout the incubation with no significant difference between ALS- and control samples. Unexpectedly, it was also observed that haemolysis occurred significantly earlier in the ALS- than in the control samples. Plateau levels in plasma lead concentration of the same order as in the present experiments have been observed both in ALS- and control samples in previous experiments with the same technique, where the lead dose added was twice as high, and these plateau levels were about 10 times higher than those observed in vivo in ALS patients and controls. It is therefore suggested that the final plasma lead concentrations in vivo is established by factors other than the erythrocyte uptake and it is improbable that the differences between ALS patients and controls in plasma lead concentration are associated with differences in the degree of lead uptake by the red cells. The increased plasma lead concentrations in ALS patients may instead be caused by increased fragility of the erythrocytes, as manifested by the earlier occurrence of haemolysis in the present experiments. The observation of increased red cell fragility is, however, also of interest as a possible manifestation of a generalized membrane defect.

Cytotoxic activity in the plasma of amyotrophic lateral sclerosis (ALS) patients against normal erythrocytes. Quantitative determinations

Erythrocytes from healthy controls were incubated during 5.5 h under physiological conditions in own plasma, and plasma from blood type-matched ALS patients and diseased controls. Free haemoglobin (Hb) was repeatedly measured spectrophotometrically as a measure of haemolysis. Using a standardized procedure, the interexperimental variation in Hb release during incubation in own control plasma was kept very low in a large series of incubations (n = 77). This indicated that the condition of the erythrocytes had been constant. Parallel incubations in plasma from ALS patients (n = 20) rendered a clear-cut increase in Hb release, which in all cases exceeded the confidence interval of the controls. There was no correlation between Hb release in the ALS group and age, sex, type of disease or duration of symptoms. The cytotoxic effect of ALS plasma was abolished by heating to 56 degrees C. In a material of incubations in plasma of diseased controls (traumatic spinal cord disease, Guillain-Barré syndrome), the haemolysis was slightly larger than in control plasma, but in all cases smaller than in ALS plasma.

Immunoglobulins from patients with amyotrophic lateral sclerosis affect human erythrocyte acetylcholinesterase

Human erythrocyte acetylcholinesterase (AChE) solubilized with Triton X-100 and obtained as a complex with micelles containing Triton and membrane phospholipids was incubated with immunoglobulins (Igs) from patients with amyotrophic lateral sclerosis (ALS) and from normal individuals. The temperature dependence of the AChE activity was determined. Biphasic (broken) Arrhenius plots were obtained with control Igs with the break point at 32.8 +/- 0.3 degrees C (SD, n = 18) indicating that the enzyme changes its conformation at this temperature. With ALS-Igs monophasic (linear) plots were observed in 14 cases and a biphasic in one case. ALS-Igs prevent the conformational change occurring at the break point temperature. The activation energy at physiological temperature increased by 60% from 2.4 to 3.8 kcal/mol (10.0-15.9 kJ/mol) which implies that ALS-Igs inhibit AChE. Thus, ALS-patients have autoantibodies that change the normal behaviour of erythrocyte AChE and which bind to the enzyme molecule or/and to phospholipids associated with the enzyme. At least part of the autoantibodies should be directed against the enzyme molecule, since a change in the Arrhenius plot was also observed in a control experiment with AChE which probably had micelles without any phospholipids. This enzyme was isolated by affinity chromatography and was washed with a buffer containing Triton X-100 before desorption from the affinity column, a treatment known to remove all phospholipids from erythrocyte AChE.

Selective vulnerability of alpha motor neurons in ALS: Relation to autoantibodies toward acetylcholinesterase (AChE) in ALS patients

The degenerative process in amyotrophic lateral sclerosis (ALS) concerns primarily alpha motor neurons in the spinal cord and brain stem, and neurons forming descending pathways to the cord, especially in the pyramidal tract. Some degeneration of large peripheral sensory nerve fibers often occurs too, but preganglionic autonomic neurons and gamma motor neurons are most often spared in the disease. The vulnerability of alpha motor neurons compared to other types of neurons in ALS is discussed in relation to retrograde axoplasmic transport from peripheral blood of foreign noxious macromolecules, interneuronal transport of such molecules, and neuronal surface structure properties relevant to uptake for retrograde axoplasmic transport. Certain differences in these aspects between alpha motor neurons and other neuronal types exist. Some differences concern the neuronal turnover of acetylcholinesterase (AChE), which could be of special interest in view of the recent demonstration of regular occurrence of autoantibodies towards this enzyme in ALS patients.

CYTOTOXIC FACTOR IN PLASMA FROM ALS PATIENTS PROVOKES HAEMOLYSIS OF NORMAL ERYTHROCYTES.

During our studies of the metabolism of lead in ALS-patients. the uptake of lead by the erythrocytes from plasma was measured in incubated whole-blood It was observed that haemolysis occurred significantly (p < 0.01) earlier in the incubated blood samples from ALS-patients than in those from controls. The time of incubation until haemolysis showed a weak correlation to pre-incubation plasma lead levels in the ALS-cases. This indicates that the abnormal fragility of erythrocytes could be responsible for the elevated plasma lead levels earlier found by us to occur in ALS-patients. The observation is however also of interest as a possible manifestation of a more wide-spread membrane defect in the disease. The present study was undertaken in order to study whether plasma from ALS-patients can induce abnormal fragility of erythrocytes.

Immunoglobulin-mediated activity against red blood cells in the saliva of amyotrophic lateral sclerosis (ALS) patients

Immunoglobulin (Ig)‐mediated activity in plasma directed towards normal blood type matched red blood cells (RBC) inducing haemolysis in vitro has earlier been demonstrated to be a characteristic feature in ALS‐patients. In this study, saliva of ALS‐patients, normal and diseased controls was tested with the same in vitro test. An increased degree of haemolysis was induced by the ALS‐patient as compared with control samples. The activity thus found in saliva had the same basic characteristics as that earlier described for plasma; it reacted similarly to serial dilution and was retained in salivary Ig. The effect on red blood cells of saliva from patients with bulbar paralysis was larger than that of saliva from ALS‐patients lacking bulbar symptoms. It is discussed whether cytotoxic Ig in saliva could be pathophysiologically active in bulbar paralysis by means of passage through the oral mucosa and local action on motor end plates in perioral muscles.