Investigation of surface properties of rat spinal ganglion neuron by microelectrophoresis

The surface charge of isolated neurons in rat spinal ganglia was studied by microelectrophoresis. The surface potential of these neurons was shown to be caused by anionic groups which form complexes with calcium ions with a binding constant of between 10 and 50 liters/mole, and to titrate with hydrogen ions in accordance with pK=3.8. After treatment with trypsin under "mild" conditions many of these groups are removed from the surface. Tosyl chloride (a reagent for amino groups) leads to a small increase, and N-bromosuccinimide (a reagent for carboxyl groups) leads to a marked decrease in surface charge. It is suggested that the surface charge of neurons in rat spinal ganglia, determined by microelectrophoresis, is due to carboxyl groups of peripheral proteins. These groups are evidently located in the glycoprotein layer covering the outer side of the membrane. According to estimates the distance between the groups is about 2 nm and the thickness of the glycoprotein layer is 10 nm.A. A. Bogomolets Institute of Physiology, Academy of Sciences of the Ukrainian SSSR, Kiev. Translated from Neirofiziologiya, Vol. 16, No. 2, pp. 176–182, March–April, 1984.     

Effect of an epoxy derivative of 2′5′-trioligoadenylate on human neuroblastoma cells

We studied the effect of an epoxy derivative of dephosphorylated 2′,5′-trioligoadenylate (5′,5′ApApAepoxy) resistive to the action of cellular phosphodiesterase on cells of human neuroblastoma IMR 32 cultured in vitro. Twenty-two hours after the addition of 5·10⁻⁶ M 2′,5′ApApAepoxy to the culture medium, the number of cells decreased by 20% (P < 0.05), while the content of protein in these cells increased, on average, by 52% (P < 0.01), as compared with the control. The activities of Na⁺,K⁺-and Ca²⁺, Mg²⁺-ATPases in a microsomal fraction obtained from cells cultured in the presence of 2′, 5′ ApApAepoxy decreased by 50% (P < 0.001) as compared with those in the control cells. Our data indicate that 2′,5′ApApAepoxy possess antiproliferative activity. According to our findings, the antiproliferative effect of 2′,5′ ApApAepoxy can, to a great extent, be explained by the fact that this oligoadenylate derivative significantly modulates the activities of Na⁺,K⁺-and Ca²⁺,Mg²⁺-ATPases. Neirofiziologiya/Neurophysiology, Vol. 38, No. 2, pp. 97–102, March–April, 2006.     

Effect of dephosphorylated 2′,5′-trioligoadenylate on the entry of sodium ions into human neuroblastoma cells

Using a radioisotope technique, we studied the effect of dephosphorylated 2′,5′-trioligoadenylate (2′,5′ ApApA) on the entry of sodium ions into cultured human neuroblastoma cells (IMR 32 strain). Short-term (nearly 1 h) action of 2′,5′ ApApA did not influence the entry of sodium ions through voltage-operated sodium channels in the absence of neurotoxins modulating the characteristics of these channels (toxin of a scorpion, Leiurus quinquestriatus, + veratrine). At the same time, 2′,5′ ApApA weakened in a dose-dependent manner the entry of sodium ions through sodium channels opened upon the action of the above neurotoxins. In cells cultured for 22 h in a medium containing 5 · 10⁻⁶ M 2′,5′ ApApA, the entry of sodium ions in the absence of neurotoxins was 25% greater, while in the presence of neurotoxins it was 24% smaller than that in the control cells. Tetrodotoxin (TTX, 4 · 10⁻⁷ M) blocked completely sodium entry through sodium channels in cells cultured in the absence of 2′,5′ ApApA, while in cells cultured in the presence of this adenylate TTX decreased the entry by 64%. It is hypothesized that long-lasting action of 2′,5′ ApApA results in the appearance of voltage-operated TTX-insensitive sodium channels in the plasma membrane of IMR 32 cells. Our data show that 2′,5′ ApApA is capable of modulating to a considerable extent the functioning of mechanisms controlling transport of sodium ions in cells of human neuroblastoma cells of the IMR 32 strain. Neirofiziologiya/Neurophysiology, Vol. 40, No. 1, pp. 3–8, January–February, 2008.     

Modification to the Hodgkin-Huxley equation as applied to the somatic membrane of mollusk giant neurons

A modified system of Hodgkin-Huxley equations was used to describe transmembrane ionic currents during fixed changes of membrane potential and generation of action potentials in the soma of mollusk giant neurons. The effect of the axon was disregarded. The results of theoretical calculations are in satisfactory agreement with experimental results.A. A. Bogomolets Institute of Physiology, Academy of Sciences of the Ukrainian SSR, Kiev. Translated from Neirofiziologiya, Vol. 5, No. 3, pp. 315–322, May–June, 1973.     

Involvement of GDNF and LIMK1 and heat shock proteins in drosophila learning and memory formation

Molecular mechanisms of the synapse and dendrite integrity maintenance and their disruption in psychiatric and neurodegenerative diseases (NDD) are being studied intensively to identify target genes for therapeutic activities. It is suggested that synapse is a tripartite system in which glia, alongside with well-studied pre- and postsynaptic neurons, represents a third, poorly studied partner in synaptic transmission involved in a positive feedback loop between the other two partners. It is the glia cell-derived neurotrophic factor (GDNF) and the transmembrane proteins, neuregulins, that mediate bidirectional coupling between presynaptic neurons and their postsynaptic targets. Neuregulins are structurally related to the epidermal growth factor and have a cytoplasmic domain that interacts with intracellular LIM kinase 1 (LIMK1), the key enzyme of actin remodeling. Since neurons and axons that do not receive a sufficient GDNF supply are at risk of degeneration and synapse elimination, GDNF became a central target factor in human NDD therapy. The delivery of GDNF-producing stem cells to the nidus of neurodegeneration by transplantation surgery is an efficient tool for NDD treatment. A new approach is proposed based on the use of the Drosophila heat shock (hs) promoter that responds to the mammalian body temperature and ensures constant expression of the human GDNF gene. The Drosophila models facilitate studying the role of each component of the bidirectional signaling between pre- and postsynaptic neurons in the development of the key diagnostic NDD symptom—a defective memory formation resulted from synaptic atrophy. To assess the efficiency of memory formation depending on the level of GDNF and LIMK1 brain expression, we used the Drosophila strains simulating different nervous system disorders: GDNF, the transgenic flies that carry the human GDNF gene under hs-promoter, l(1)ts403, the mutants with disruption of heat shock proteins (HSPs) mRNA nuclear trafficking, and agn ^ts3 carrying a mutation in LIMK1 gene. We investigated at the behavioral (learning/memory) level the functional connections between GDNF, LIMK1 and HSP signaling transductions that might offer promising targets for complex approaches to NDD treatment.     

Effect of interferon-α 2b on cells from human embryonic nerve tissue in the early stages of neurogenesis

We studied the effects of interferon (IFN)- 2b on cells obtained from the brain of human embryos (4 to 12 weeks of gestation). It was demonstrated that IFN exerts modulatory effects on biochemical and physico-chemical properties of cells of embryonic nerve tissue in the early stages of embryonic development (from 4 weeks of gestation). IFN decreased the content of protein, inhibited the activity of Na⁺,K⁺-ATPase, and induced changes in the superficial charge of the plasma membrane. Based on the obtained experimental data, we suppose that IFN- 2b is involved in regulation of neurogenesis.Neirofiziologiya/Neurophysiology, Vol. 36, Nos. 5/6, pp. 363–369, September–December, 2004.This revised version was published online in April 2005 with a corrected cover date and copyright year.     

Effect of calcium ions on outward currents in the somatic membrane of mollusk giant neurons

Potassium currents through the somatic membrane of giant neurons ofHelix pomatia in normal (10 mM Ca) Ringer's solution and low-calcium (1 mM Ca) solution were studied by the voltage clamp method. With a decrease in the Ca concentration to 1 mM peak potassium conductance versus membrane, potential curves and inactivation curves were shifted along the voltage axis in the negative direction by about 10 mV. Inactivation of the delayed potassium current was slowed in low Ca solution. The effect of a decrease in external calcium concentration on volt-ampere and inactivation characteristics increased with a rise in external pH. These effects of a low Ca concentration on potassium mechanisms of the giant neuron somatic membrane can be attributed to changes in the negative surface potential in the region of the potassium channels.A. A. Bogomolets Institute of Physiology, Academy of Sciences of the Ukrainian SSR, Kiev. Institute of Biology, Hungarian Academy of Sciences, Tihany. Translated from Neirofiziologiya, Vol. 8, No. 4, pp. 400–409, July–August, 1976.     

Effect of Recombinant Interferon-α2b (Laferon) on Transport of Sodium Ions in Human Neuroblastoma Cells

To elucidate molecular mechanisms of neurotropic action of a recombinant interferon, IFN-2b (laferon), its effect on transport of ²²Na⁺ through the membrane of cultured human neuroblastoma cells (line IMR 32) was investigated. Within the first minutes after treatment with IFN-2b, the influx of ²²Na⁺ ions was reduced by 20%, as compared with the control. Depolarization of the plasma membrane by a mixture of veratrine and scorpion (Leiurus quinquestriatus) toxin (200 and 10 g/ml, respectively) increased this flux by 50% in the control and by 70% in the IFN-2b-treated cells. A blocker of voltage-operated sodium channels, tetrodotoxin (TTX, 4 · 10^-7 M), suppressed the inward flux of ²²Na⁺ ions (completely in the control cells and by 75% in the IFN-2b-treated cells). The influx of ²²Na⁺ ions into neuroblastoma cells depended on the concentration of IFN-2b in the incubation medium, reaching a maximum at concentrations of 600-1000 IU/ml. This allows us to suggest that entry of Na⁺ ions into neuroblastoma cells caused by IFN-2b is basically performed through voltage-operated TTX-sensitive sodium channels.     

Surface charge at the cytoplasmic membrane of murine C1300 neuroblastoma cells

Inside-out vesicles (IV, mainly with the cytoplasmic side outermost) were obtained from the plasma membranes of neuroblastoma cells from strain C1300 mice, clone N18. These served as a convenient model for investigating the surface charge of the cytoplasmic side of the cell membrane using microelectrophoretic techniques. Electrophoretic mobility (EM) at a neutral pH and with an external medium of the same ionic strength was found to be 2.7-fold less than that of right-side-out vesicles (RV). Processing vesicles with neuraminidase reduced EM in RV but not in IV, while trypsin did so in both. Treatment with phospholipase C produced the same effect in IV but none in RV. Phospholipase D increases the EM of both types of vesicles. Charged groups at the surface of IV are titrated in relation to a pK of 3.5. The EM of IV is dependent on the Ca²⁺ concentration of the external medium. On the cytoplasmic side of the membrane, Ca²⁺ forms a 11 complex with negatively charged protein and lipid molecules as well as those lipid molecules found with a neutral pH, in the form of a zwitterion with binding constants of 50, 12, and 25 liter/mole respectively.A. A. Bogomolets Institute of Physiology, Academy of Sciences of the Ukrainian SSR, Kiev. Translated from Neirofiziologiya, Vol. 20, No. 5, pp. 610–617, September–October, 1988.