Exposure to cerebrospinal fluid of sporadic amyotrophic lateral sclerosis patients alters Nav1.6 and Kv1.6 channel expression in rat spinal motor neurons.

Cerebro Spinal Fluid (CSF) from patients with ALS has been documented to have a toxic effect on motor neurons both in vivo and in vitro. Here we show that the CSF from Amyotrophic Lateral Sclerosis (ALS) patients (ALS-CSF) has the potential to perturb ion channel expression, specifically the Na(v)1.6, and K(v)1.6 channels in newborn rat spinal motor neurons both in vivo and in vitro. ALS-CSF and CSF from nonALS patients (nonALS-CSF) were intrathecally injected into 3-day-old rat pups at the rate of 1 microl/2.5 min using a microinjector. In addition, embryonic rat spinal cord cultures were also exposed to 10% ALS or nonALS-CSF on the 9th day in vitro (9DIV) in serum free DMEM medium. After 48 h of CSF exposure, the cultures and the spinal cord sections were processed for immunostaining of the above mentioned ion channels. We observed a decrease in the expression of Na(v)1.6 and K(v)1.6 channels in motor neurons in ALS-CSF treated group, and the presence of trophic factors like Brain Derived Neurotrophic Factor (BDNF) and Ciliary Neurotrophic Factor CNTF partially reversed the effects produced by ALS-CSF. Altered expression of these voltage-gated channels may interfere with the electrical activity of motor neurons, and thereby lead to the degeneration of neurons.

Altered in-vitro and in-vivo expression of glial glutamate transporter-1 following exposure to cerebrospinal fluid of amyotrophic lateral sclerosis patients.

Our earlier studies have shown that cerebrospinal fluid (CSF) of amyotrophic lateral sclerosis (ALS) patients causes death of motor neurons, both in in-vitro as well as in-vivo. There was an aberrant phosphorylation of neurofilaments in cultured spinal cord neurons of chick and rats following exposure to CSF of ALS patients (ALS-CSF). Other features of neurodegeneration, such as swollen neuronal soma and beading of neurites were also observed. In neonatal rat pups exposed to ALS-CSF, we observed phosphorylated neurofilaments in the soma of spinal motor neurons in addition to the increased lactate dehydrogenase activity and reactive astrogliosis. The present study examines the effect of ALS-CSF on the expression of glial glutamate transporter (GLT-1) in embryonic rat spinal cord cultures as well as in spinal astrocytes of neonatal rats. Immunostaining suggested a decrease in the expression of GLT-1 by astrocytes both in culture and in-vivo following exposure to ALS-CSF. Quantification of Western blots confirmed the decreased expression of GLT-1. Our results provide evidence that toxic factor(s) present in ALS-CSF depletes GLT-1 expression. This could lead to an increased level of glutamate in the synaptic pool causing excitotoxicity to motor neurons, possibly by triggering the 'glutamate-mediated toxicity-pathway'.

[Cytochemical study of dehydrogenase activity in the tissues of the pig female genital system after injection of xenogenic cerebrospinal fluid (a cytochemical study)].

The experimental study was performed to evaluate the biological effects of parenteral injection of cerebrospinal fluid (SCF) on the uterus, uterine tubes and the ovary of piglets aged 60 days. Lactating cows aged 4-6 years after first calving were taken as SCF donors, since their SCF contained maximal spectrum of biologically active substances. For injection the whole SCF, SCF after passage through bacterial filters and SCF supernatant were used. The effect of SCF was assessed using histological and histochemical (succinate- and lactate dehydrogenases demonstration) methods. It was found that SCF preparations studied had differential effect on oxidative-reduction processes in the tissues of reproductive system organs of immature pigs, that is associated with the broad spectrum of biologically active substances in mammalian SCF.

[Effect of fractions of the cerebrospinal fluid from patients with drug dependence on basic electrophysiological characteristics of the rat olfactory cortex slices]. / Issledovanie éffektov otdel'nykh fraktsii likvora bol'nykh narkoticheskoi zavisimost'iu na bazisnye élektrofiziologicheskie kharakteristiki perezhivaiushchikh srezov kory mozga krys.

Heroin addicts at the initial stage of abstinence syndrome were subjected to detoxication by liquorosorption technique. The fractions of their cerebrospinal fluid obtained by the thin layer chromatography technique were analyzed. The substances extracted from the cerebrospinal fluid of drug addicts, presumably peptides, negatively affected the conductive function and synaptic transmission in surviving slices of the olfactory cortex of rats. The conclusion was drawn about a possibility of application of surviving rat brain slices as test object for estimation of the extent of purification of the cerebrospinal fluid from toxic endogenous substances after the liquorosorption.

A novel substance associated with gallamine-induced myoclonus.

If gallamine or d-tubocurarine gains access to the central nervous system it produces a myoclonus, a synchronized jerking of many skeletal muscles. Each jerk is accompanied by a slow wave in the inferior olive. The jerking continues for 24 h or more after the gallamine or d-tubocurarine can no longer be detected in the CSF. We report here that a novel substance appears in the CSF and persists for a long period of time, possibly as long as the twitching. This substance is not corticotrophin-releasing factor (CRF) nor does CRF or harmaline (a substance causing a tremor by an action on the inferior olive) lead to the appearance of the novel substance. At present the nature of this substance is not known.

Purification of a factor from CSF in patient after SAH which induces the cytosolic free calcium elevation in vascular smooth muscle cells.

Cerebrospinal fluid from patients with aneurysmal subarachnoid hemorrhage induces the elevation of cytosolic free calcium [Ca2+]i in cultured vascular smooth muscle cells. We have purified a [Ca2+]i elevating factor from cerebrospinal fluid of a patient with subarachnoid hemorrhage due to aneurysm rupture. The calcium-elevating protein factor was purified to homogeneity by ammonium sulfate precipitation and a combination of Mono Q, Superose 12, and Mono S columns using liquid chromatography. Fifteen microgram of the purified protein was obtained from 340 mg of cerebrospinal fluid proteins and the molecular mass of the protein was estimated to be 81 kDa by sodium dodecyl sulfate polyacrylamide gel electrophoresis. It was also shown that the purified protein was cross-reactive with anti-human transferrin antibody. These results suggested that transferrin may be involved with the cerebral vasospasm after subarachnoid hemorrhage.

Cerebrospinal fluid of Parkinson's disease patients inhibits the growth and function of dopaminergic neurons in culture.

We report the possible existence of an inhibitory factor in the CSF of Parkinson's disease patients that inhibits the function and growth of dopaminergic neurons in rat mesencephalic culture. After 40 hours' exposure to the < 10 kd fraction of CSF from PD patients, the high-affinity dopamine uptake was 66% of that of cultures exposed to CSF from controls. However, the number of dopaminergic neurons remained unchanged at this time. After 90 hours' exposure to the < 10 kd fraction of CSF from PD patients, the number of dopaminergic neurons decreased to 10% of that in cultures exposed to CSF from controls, and the size of the remaining dopaminergic neurons in the culture became smaller. This inhibitory factor did not affect the growth of other types of neurons. The chemical nature of this inhibitory factor is under investigation.

Facilins, a novel class of biological factors that facilitate the aortic response to dopamine and other biogenic amines.

Biological fluids and tissues extracts were shown to contain biological factors, termed facilins, that facilitate the dopamine-, adrenaline-, and serotonin-mediated aortic contraction at concentrations devoid of any direct effect. Cyproheptadine and phentolamine antagonized the direct contracting effect of biogenic amines, but not the facilitated component of the aortic response thus indicating that the mechanism of action of facilins was unlike that of biogenic amines. Fresh schizophrenics' CSF displayed a stronger facilitating effect than normal CSF on the dopamine-mediated aortic response. This finding, however, was not confirmed with samples kept frozen for prolonged periods of time. Multiple molecular forms of facilins were detected in rabbit serum. Those with a high apparent molecular weight were proteinous and were neither insulin nor other factors known for their contracting effects on the aorta such as epidermal growth factor, transforming growth factor-beta, and platelet-derived growth factor.

DNA synthesis and intracellular calcium elevation in porcine cerebral arterial smooth muscle cells by cerebrospinal fluid from patients with subarachnoid haemorrhage.

To understand the molecular mechanism of the pathogenesis of cerebral vasospasm following subarachnoid haemorrhage, we analysed the effect of cerebrospinal fluid from patients with subarachnoid haemorrhage on DNA synthesis and cytosolic-free calcium elevation in cultured porcine cerebral smooth muscle cells. Cerebrospinal fluid from patients on day 2 after subarachnoid haemorrhage induced transient elevation in cytosolic-free calcium levels. In contrast, the maximal elevation of cytosolic-free calcium levels induced by cerebrospinal fluid from control patients (without subarachnoid haemorrhage) was significantly lower than that induced by cerebrospinal fluid from patients with subarachnoid haemorrhage. In cultured porcine cerebral arterial smooth muscle cells, cerebrospinal fluid from patients with subarachnoid haemorrhage promoted levels of [3H]-thymidine incorporation (DNA synthesis) more than 2.5-fold higher than that promoted by cerebrospinal fluid from control patients without subarachnoid haemorrhage. However, in cultured aortic smooth muscle cells, there was no significant difference in [3H]-thymidine incorporation between cerebrospinal fluid from patients with subarachnoid haemorrhage and that by control cerebrospinal fluid. From these results in cerebral arterial smooth muscle cells, cerebrospinal fluid from patients following subarachnoid haemorrhage may play not only constrictive functions, evidenced by cytosolic-free calcium elevations, but also proliferative functions, demonstrated by promotion of [3H]-thymidine incorporation. The relevance of these factors to vasospasm will be discussed.

Human immunodeficiency virus type 1 (HIV-1) infection of the central nervous system: an evaluation of cytokines in cerebrospinal fluid.

Cytokines play an important role not only for initiation of immune reactivity but also for development of tissue injury. Of 38 patients infected with human immunodeficiency virus type 1 (HIV-1) interleukin-1 beta (IL-1 beta) and interleukin-6 (IL-6) were identified in cerebrospinal fluid (CSF) of 22 (58%) and 16 (42%) patients, respectively. Among the IL-1 beta- and IL-6-positive CSF were eight of 15 HIV-1 patients with no clinical signs of central nervous system involvement and four of five patients with acquired immunodeficiency syndrome (AIDS) dementia complex. The presence of IL-6 was often associated with IL-1 beta and soluble interleukin-2 receptor in CSF as well as with intrathecal IgG synthesis. In none of the CSF samples tumor necrosis factor-alpha or interleukin-2 was detected.