A new insight into mechanisms of interferon alpha neurotoxicity: Expression of GRIN3A subunit of NMDA receptors and NMDA-evoked exocytosis.

Neurological and psychiatric side effects accompany the high-dose interferon-alpha (IFNA) therapy. The primary genes responsible for these complications are mostly unknown. Our genome-wide search in mouse and rat genomes for the conservative genes containing IFN-stimulated response elements (ISRE) in their promoters revealed a new potential target gene of IFNA, Grin3α, which encodes the 3A subunit of NMDA receptor. This study aimed to explore the impact of IFNA on the expression of Grin3α and Ifnα genes and neurotransmitters endo/exocytosis in the mouse brain. We administered recombinant human IFN-alpha 2b (rhIFN-α2b) intracranially, and 24 h later, we isolated six brain regions and used the samples for RT-qPCR and western blot analysis. Synaptosomes were isolated from the cortex to analyze endo/exocytosis with acridine orange and L-[14C]glutamate. IFNA induced an increase in Grin3α mRNA and GRIN3A protein, but a decrease in Ifnα mRNA and protein. IFNA did not affect the accumulation and distribution of L-[14C]glutamate and acridine orange between synaptosomes and the extra-synaptosomal space. It caused the more significant acridine orange release activated by NMDA or glutamate than from control mice's synaptosomes. In response to IFNA, the newly discovered association between elevated Grin3α expression and NMDA- and glutamate-evoked neurotransmitters release from synaptosomes implies a new molecular mechanism of IFNA neurotoxicity.

Dual benefit of combined neuroprotection: Cholesterol depletion restores membrane microviscosity but not lipid order and enhances neuroprotective action of hypothermia in rat cortex nerve terminals.

Here, both neuroprotectants, i.e. cholesterol depletion of the plasma membrane of rat brain nerve terminals (synaptosomes) using methyl-ß-cyclodextrin (MßCD) and deep/propound hypothermia, were analyzed during their combined administration and regarding additive neuroprotective effect. The extracellular synaptosomal level of L-[14C]glutamate significantly increased after treatment with MßCD in both deep and profound hypothermia. Cholesterol depletion gradually enhanced inhibiting effect of deep and profound hypothermia on glutamate uptake and "excitotoxic" transporter-mediated release of L-[14C]glutamate. A decrease in L-[14C]glutamate release via heteroexchange from nerve terminals in deep and profound hypothermia was enhanced by cholesterol deficiency that confirmed previous result. Fluorometric studies with probes NR12S and DCVJ revealed oppositely directed effects of cholesterol depletion and hypothermia on synaptosomal membrane lipid order and microviscosity showing that cholesterol depletion can normalise up to the control hypothermia-induced increase in microviscosity, but not the lipid order of the synaptosomal membrane. Dynamics of changes in exocytosis in nerve terminals, which involved membrane fusion stage, was different from transporter-dependent ones. Hypothermia did not augment effects of cholesterol depletion on exocytotic L-[14C]glutamate release and lowering cholesterol enhanced the impact of deep, but not profound hypothermia on this parameter. Therefore, dual benefit of combined neuroprotection was demonstrated. Cholesterol depletion enhanced neuroprotective effects of hypothermia intensifying inhibition of "excitotoxic" transporter-mediated glutamate release and can normalise a hypothermia-induced increase in microviscosity of the synaptosomal membrane. This feature is prospective in mitigation of side effects of therapeutic hypothermia, and also for brain conservation preserving normal physical and chemical properties of the cellular membranes.

Levetiracetam-mediated improvement of decreased NMDA-induced glutamate release from nerve terminals during hypothermia.

A combination of a beneficial neuroprotectant, hypothermia, with targeted medication is a perspective therapeutic approach. Here, we analyzed both non-specific (deep and profound hypothermia, 27 °C and 17 °C, respectively) and targeted (anticonvulsant drug levetiracetam) modulation of l-[14C]glutamate release induced by activation of presynaptic NMDA, AMPA, and kainate receptors in rat brain nerve terminals (synaptosomes). Gradual dynamics of hypothermia-mediated decrease in synaptosomal l-[14C]glutamate release evoked by the receptor agonists NMDA-, AMPA-, and kainate (250 µM) has been demonstrated that can be of value for the justification of optimal temperature regimes in therapeutic hypothermia. 250 µM NMDA-induced l-[14C]glutamate release from nerve terminals was higher in the presence of levetiracetam (100 µM) as compared to that without the drug. Despite levetiracetam effects decreased in hypothermia, combined application of hypothermia and levetiracetam resulted in higher NMDA-induced l-[14C]glutamate release from nerve terminals as compared to that without the drug. These effects were not revealed for synaptosomal AMPA- and kainate-induced l-[14C]glutamate release in the presence of levetiracetam at the similar concentration. Therefore, levetiracetam administration significantly mitigated a hypothermia-induced decrease in NMDA receptor response at the presynaptic level and can be used for the targeted neurocorrection to reduce side effects of hypothermia in cardiac surgery. However, levetiracetam-mediated improvement of NMDA receptor response is not applicable in stroke, brain trauma and neonatal asphyxia therapies, where the main neuroprotective action of hypothermia is associated with prevention of damaging consequence of pre-existing acute glutamate exitotoxicity.

An amperometric glutamate biosensor for monitoring glutamate release from brain nerve terminals and in blood plasma.

An excess of the excitatory neurotransmitter, glutamate, in the synaptic cleft during hypoxia/ischemia provokes development of neurotoxicity and originates from the reversal of Na+-dependent glutamate transporters located in the plasma membrane of presynaptic brain nerve terminals. Here, we have optimized an electrochemical glutamate biosensor using glutamate oxidase and developed a biosensor-based methodological approach for analysis of rates of tonic, exocytotic and transporter-mediated glutamate release from isolated rat brain nerve terminals (synaptosomes). Changes in the extracellular glutamate concentrations from 11.5 ±â€¯0.9 to 11.7 ±â€¯0.9 µΜ for 6 min reflected a low tonic release of endogenous glutamate from nerve terminals. Depolarization-induced exocytotic release of endogenous glutamate was equal to 7.5 ±â€¯1.0 µΜ and transporter reversal was 8.0 ±â€¯1.0 µΜ for 6 min. The biosensor data correlated well with the results obtained using radiolabelled L-[14C]glutamate, spectrofluorimetric glutamate dehydrogenase and amino acid analyzer assays. The blood plasma glutamate concentration was also tested, and reliability of the biosensor measurements was confirmed by glutamate dehydrogenase assay. Therefore, the biosensor-based approach for accurate monitoring rates of tonic, exocytotic and transporter-mediated release of glutamate in nerve terminals was developed and its adequacy was confirmed by independent analytical methods. The biosensor measurements provided precise data on changes in the concentrations of endogenous glutamate in nerve terminals in response to stimulation. We consider that the glutamate biosensor-based approach can be applied in clinics for neuromonitoring glutamate-related parameters in brain samples, liquids and blood plasma in stroke, brain trauma, therapeutic hypothermia treatment, etc., and also in laboratory work to record glutamate release and uptake kinetics in nerve terminals.

Dynamic Gradient of Glutamate Across the Membrane: Glutamate/Aspartate-Induced Changes in the Ambient Level of L-[14C]glutamate and D-[3H]aspartate in Rat Brain Nerve Terminals.

Extracellular/intracellular L-[14C]glutamate exchange and conservativeness of the extracellular level of L-[14C]glutamate was analyzed in isolated rat brain nerve terminals. L-Glutamate-, DL-threo-ß-hydroxyaspartate (DL-THA)-, and D-aspartate-induced increase in the ambient level of L-[14C]glutamate or D-[3H]aspartate was evaluated comparatively. 100 µM "cold" nonradiolabeled L-glutamate, DL-THA, D-aspartate extruded a quarter of radioactivity from L-[14C]glutamate-preloaded synaptosomes for 6 min. The similar results were obtained with L-glutamate-evoked extracellular/intracellular redistribution of D-[3H]aspartate. Contribution of presynaptic glutamate receptors to an increase in the extracellular L-[14C]glutamate level was evaluated using receptor agonists NMDA, AMPA, and kainate (100 µM), and it consisted of less than 5 % of total accumulated label. The existence of the efficient extracellular/intracellular glutamate exchange, and so dynamic glutamate gradient across the plasma membrane of nerve terminals was demonstrated. A two-substrate kinetic algorithm that included transporter reversal was considered. The extracellular level of L-[14C]glutamate and D-[3H]aspartate in nerve terminals depended on the amount of exogenous substrates of glutamate transporter available. Taking into account that L-glutamate, DL-THA, and D-aspartate are the substrates of glutamate transporters, and also the similarity in their effectiveness in the establishment of new extracellular level of the neurotransmitters, the central role of glutamate transporters in permanent glutamate turnover in nerve terminals was demonstrated.

[Organisational aspects of medical support for civilians employed in the Armed Forces in the military-medical institutions of the Ministry of Defence, deployed in Moscow].

Organisational aspects of medical support for civilians employed in the Armed Forces in the military-medical institutions of the Ministry of Defence, deployed in Moscow. To ensure social protection of the civilian personnel of the Armed Forces is one of the main tasks of the Ministry of Defence of the Russian Federation. In Moscow formed a territorial system of medical support of citizens who have the right for medical care in military medical institutions of the Ministry of Defence of the Russian Federation. Russian legislation does not provide the right for medical assistance provision to the civilian personnel of the Armed Forces in military medical institutions at the expense of funds allocated from the federal budget for the maintenance of the Ministry of Defence of the Russian Federation. The function of the physician in providing primary medical care performs primary care physician. Providing medical assistance to the civilian personnel of the Armed Forces of the Russian Federation in military medical institutions on the basis of their attachment to the clinics onlv the Russian Defence Ministrv. or in the direction of the clinics of Moscow.

[Non-infectious morbidity in the Armed Forces (a review on results of the round table in the State Duma)].

The article presents the results of the medical service of the Armed Forces of the Russian Federation on prevention of non-infectious diseases among the military, the results reported on May 12, 2016 at the meeting of the Committee for the Protection of Health of the State Duma of the Federal Assembly of the Russian Federation. Attention paid to the fact that the Armed Forces are an integral part of society and the state, and therefore the negative trend in the health status of the population are also reflected in the troops. In the common-ness of non-communicable diseases in the Armed Forces have an influence, especially of military service. Consider an organization and legal framework to combat non-communicable diseases in the Armed Forces successes in this direction, problematic issues and their solution.

[Organisation of medical care delivery to citizens, enjoying a right to get medical care at military-medical organisations of the Ministry of Defence of the Russian Federation].

One of the main priorities of the medical service of the armed forces of the Russian federation is a realization of rights for military retirees and members of their families to free medical care. For this purpose was founded a system of organization of medical care delivery at military-medical subdivisions, units and organizations of the ministry of defence of the Russian federation, based on territorial principle of medical support. In order to improve availability and quality of medical care was determined the order of free medical care delivery to military servicemen and military retirees in medical organizations of state and municipal systems of the health care.

[Hi-tech health care: modern status and prospects of development in medical facilities of the Ministry of Defence].

The article is devoted to the current issues of providing hi-tech medical care in hospitals of the Ministry of Defence. Since the beginning of 2013 the executive body of the Russian Ministry of Defense pays special attention to improvement of the quality and accessibility of health care contingent of the Ministry of Defence. Thus, according to decision of the Minister of Defense of the Russian Federation, General of the Army Sergei Shoigu in 2013 more than 1.1 billion rubles (in 2012, targeted funding of high-tech medical care in the Ministry of Defence did not materialize) was allocated for military medical institutions of the Ministry of Defense of the Russian Federation to provide high-tech medical care. As a result, in 7 months in 2013 the volume of medical care has increased by 32% in comparison with the same period in 2012. Currently the main military medical department of the Ministry of Defense is working to resolve the order of delivery and financing hi-tech medical care in the Armed Forces in the following areas: inclusion of military medical institutions of the Ministry of Defence in the list of health organizations, providing high-tech medical care, approved by Order of the Ministry of Health of the Russian Federation, legal regulation of the provision of high-tech medical care in military medical establishments of the Ministry of defense of the Russian Federation within the budget appropriation allocated to the Ministry of Defence.

[Improvement of pharmaceutical benefits for military retirees in outpatient care].

For the purpose of improvement of pharmaceutical benefits for military retirees was designed the drug usage standard. This standard consists of 216 drug titles and is utilized in outpatient care in case of consistency. Monthly outpatient care costs for one military retiree are more than 900 rubles. New procedural and institutional mechanisms, ways of interdepartmental interactions, rational forms of its organization are necessary for the improvement of the effectiveness. These measures correspond to main principals of reorganization of the system of health care.

[Dental orthopedic service for retirees from MD RF in medical institutions of state and municipal system of healthcare].

Financial feasibility study of dental orthopedic service for retirees of Ministry of Defence is analyzed, suggestions about the organization of dental orthopedic service for contingent in medical institution of state and municipal system of healthcare. Information about the number of retirees, index of needs in dental orthopedic service which was 40,29%, information about 2806 prosthodontics oders for retirees in military medical institution, data about the average price of production of dental in different subjects of Russian Federation is the basis of the given research. Algorithm of dental orthopedic service for retirees by stomatologies is suggested.

Enzyme-like effect of metmyoglobin on the thermal cis-trans isomerization of stilbazolium betaine.

A photochromic compound, stilbazolium betaine M, when associated with metmyoglobin undergoes an accelerated thermal cis-trans isomerization. A study of the pH and ionic strength dependence of the isomerization reaction rate of the photochrome associated with metmyoglobin was performed. A comparative investigation of the reaction carried out in the presence of three proteins, metmyoglobin, apomyoglobin, and human albumin, indicates a specific influence of the heme pocket environment on the reaction. Possible mechanisms of the reaction acceleration are considered.

[Catalysis of 'dark' stage of cis-trans-isomerization of betaine stilbazole by myoglobin]. / Kataliz "temnovoi" stadii reaktsii tsis-trans-izomerizatsii betaina stil'bazola mioglobinom.

The back thermal cis-trans isomerization reaction of stilbazole betaine in the presence of metmyoglobin was studied. The catalytic effect of metmyoglobin heme on the back thermal cis-trans isomerization reaction of stilbazole betaine was observed.

[Effect of molecular dynamics on the photoinduced electron transfer in eosin-myoglobin complex]. / Vliianie molekuliarnoi dinamiki na fotoindutsirovannyi perenos électrona v komplekse éosin-mioglobin.

The temperature dependence of the rate constant of photoinduced electron transfer in the modified eosin-myoglobin complex by monitoring of the phosphorescence quenching of eosin is measured. The values of electron transfer rate constants are equal 10(2) + 10(3) s-1 in the temperature region 150-200 K. The kinetics of relaxation of the maximum of the time-resolved phosphorescence spectra of eosin on apomyoglobin is measured in the same temperature range. The solvation relaxation of the time-resolved phosphorescence spectra is nonexponential. The characteristic times of the solvation relaxation are given 10(-2) + 10(-4) s-1, that correlate with the time of electron transfer in this system. It was observed the "acceleration" of the relaxation rate of the time-resolved phosphorescence spectra of eosin in metmyoglobin due to nonequilibrium photoinduced electron transfer. The role of the matrix dynamics in photoinduced electron transfer in proteins is discussed.