Optogenetics: Perspectives in biomedical research

© 2016, Nizhny Novgorod State Medical Academy. All rights reserved.Optogenetic tools, photochromic switches and genetically encoded biosensors revolutionized contemporary neuroscience research. These approaches provided unprecedented opportunities for monitoring and modulating the function of specific neurons and have literally shed light on the mechanisms of neuronal networks function in the brain. A number of light-sensitive biosensors for non-invasive monitoring of ions and enzymes have been developed. These molecular designs expand extremely rapidly and a number of new approaches for image analysis of various proteins in living cells have being proposed. In this review we discuss new tools for molecular imaging and remote activation of receptors, ionic channels and synaptic networks, as well as its potential for biomedical research

Modified Recording Chamber for Sustained Kainate-Induced Gamma Oscillations in Submerged Rat Hippocampal Slices

© 2016, Springer Science+Business Media New York.Fast neuronal network oscillations in an in vitro model are a challenging task. Here, we report that sustained gamma (31–35 Hz) oscillations can be induced by kainate (50 nM) in submerged rat hippocampal slices using modified Hájos’s type recording chamber with a superfusion inlet positioned close to the CA3 pyramidal cell layer. The general features of these kainate-induced gamma oscillations were similar to those previously reported in the hippocampal slices using the interface-type chamber and superfused hippocampus in vivo. We suggest that close positioning of the superfusion inlet improves oxygen supply and temperature control of the oscillation-generating network and that this modification could be useful in studies of the gamma rhythmogenesis in the submerged slices

Kratkoročni učinci onečišćenja zraka na broj bolničkih prijama zbog bolesti srca i krvožilja i šećerne bolesti u Sofiji u Bugarskoj (2009. – 2018.)

Bulgaria has a very high incidence of cardiometabolic diseases and air pollution-related mortality rate. This study investigated the relationship between daily air pollution levels and hospital admissions for ischaemic heart diseases (IHD), cerebral infarction (CI), and type 2 diabetes mellitus (T2DM) in Sofia, Bulgaria. We obtained daily data on hospitals admissions and daily average air pollution levels from 2009 to 2018. Pollutants of interest were particulate matter (PM2.5 and PM10), nitrogen dioxide (NO2), sulphur dioxide (SO2), ozone (O3), and carbon monoxide (CO). Negative binomial regressions were fitted to study the effects of air pollution on hospital admission over the course of seven days prior to that event, accounting for autocorrelations and time trend in the data, day of the week, temperature, and relative humidity. Our findings confirm that higher air pollution levels generally increase the risk of hospital admissions for IHD and CI. For T2DM the association is less clear. Admissions often lagged several days behind and were more common in specific demographic subgroups or when pollution crossed a particular threshold. However, we did not expect to find the risk of hospital admissions increased in warmer rather than colder months of the year. Our findings are to be taken with reservation but do provide an idea about how air pollution could trigger acute episodes of related cardiovascular diseases, and our model may serve to investigate similar associations across the country.Bugarska ima izrazito visoku incidenciju kardiometaboličkih bolesti i mortaliteta koja je povezana s onečišćenjem zraka. Ovdje smo ispitivali povezanost dnevnih razina onečišćenja zraka s brojem bolničkih prijama zbog ishemijskih bolesti srca (I20 – I25), cerebralnog infarkta (I63) i šećerne bolesti neovisne o inzulinu (E11) u Sofiji u Bugarskoj. Na raspolaganju smo imali dnevne podatke o broju bolničkih prijama te o prosječnim razinama onečišćenja zraka za desetogodišnje razdoblje, tj. od početka 2009. do kraja 2018. Onečišćivala koja smo promatrali obuhvatila su lebdeće čestice (PM2.5 i PM10), dušikov dioksid (NO2), sumporov dioksid (SO2), ozon (O3) i ugljikov monoksid (CO). Kako bismo utvrdili učinke onečišćenja zraka na broj bolničkih prijama, oslonili smo se na modele binomijalne regresije, prilagođene za razdoblje do sedam dana uoči bolničkoga prijama, uzimajući pritom u obzir autokorelacije i vremenske trendove podataka, dan u tjednu te temperature i vlažnost zraka. Naši rezultati potvrđuju da veće onečišćenje načelno povećava rizik od bolničkoga prijama zbog ishemijskih bolesti srca i cerebralnog infarkta, a ta je povezanost sa šećernom bolesti nejasnija. Prijam u bolnicu obično je kasnio nekoliko dana za porastom onečišćenja te je bio učestaliji u pojedinim demografskim podskupinama odnosno nakon što bi onečišćenje prešlo određeni prag. Ono što, međutim, nismo očekivali jest da se broj bolničkih prijama (i povezani rizik) povećao za topla vremena, a ne za hladnih mjeseci. Dakako, naše rezultate treba uzeti s određenim oprezom, no i takvi daju dobru ideju kako onečišćenje zraka može potaknuti akutne epizode s njim povezanih bolesti srca i krvožilja, a naš model može poslužiti za istraživanje sličnih veza diljem Bugarske

Dynamic changes from depolarizing to hyperpolarizing GABAergic actions during giant depolarizing potentials in the neonatal rat hippocampus

© 2015 the authors. During development, GABA exerts depolarizing action on immature neurons and, acting in synergy with glutamate, drives giant depolarizing potentials (GDPs) in the hippocampal network. Yet, blockade of the GABA(A) receptors transforms GDPs to epileptiform discharges suggesting dual, both excitatory and inhibitory, actions of GABA in the immature hippocampal network. However, the nature of this dualism in early GABA actions is poorly understood. Here we characterized the dynamics of synaptic currents mediated by GABA(A) and glutamate receptors through an estimation of the changes in their conductance and driving forces in neonatal rat CA3 pyramidal cells during GDPs. We found that depolarizing GABAergic and glutamatergic currents act in synergy at the GDPs’ onset. However, during the peak of the population discharge, the inward synaptic current was essentially mediated by glutamate receptors whereas GABA currents transiently switched their direction from depolarizing to hyperpolarizing as a result of neuronal depolarization above the GABA(A) reversal potential. Thus, the action of GABA on CA3 pyramidal cells dynamically changes during GDPs from excitatory at the GDPs’ onset to inhibitory at the GDPs’ peak. We propose that the dynamic changes in GABA actions occurring during GDPs enable GABAergic interneurons not only to initiate the discharge of pyramidal cells but also to control excitation in the recurrent CA3 network preventing epileptiform synchronization

Generalized differential transformation method for solving two-interval Weber equation subject to transmission conditions

The main goal of this study is to adapt the classical differential transformation method to solve new types of boundary value problems. The advantage of this method lies in its simplicity, since there is no need for discretization, perturbation or linearization of the differential equation being solved. It is an efficient technique for obtaining series solution for both linear and nonlinear differential equations and differs from the classical Taylor’s series method, which requires the calculation of the values of higher derivatives of given function. It is known that the differential transformation method is designed for solving single interval problems and it is not clear how to apply it to many-interval problems. In this paper we have adapted the classical differential transformation method for solving boundary value problems for two-interval differential equations. To substantiate the proposed new technique, a boundary value problem was solved for the Weber equation given on two non-intersecting segments with a common end, on which the left and right solutions were connected by two additional transmission conditions

Intracellular blockade of GABA<inf>A</inf> receptors in the rat hippocampal neurons

The intracellular blockade of GABAA-receptor-mediated currents is a useful approach to suppress the GABAergic conductance in a single cell and to isolate the glutamatergic component of network-driven activities. Previously an approach has been described allowing intracellular blockade of GABAA receptors by means of intracellular dialysis of a neuron with the pipette-filling solution, in which fluoride ions that hardly pass through the GABAA receptor channels substitute for Cl- and in which Mg2+ and ATP are omitted to induce rundown of the GABAA receptors during whole-cell patch-clamp recordings. However, the kinetics of suppression of GABAergic conductance and the effect on the currents mediated by glutamate receptors remain unknown. Here, using whole-cell recordings with fluoride-based, Mg2+- and ATP-free solution on CA3 hippocampal neurons of neonatal rats, we show that after 1 h of such dialysis, both spontaneous and evoked GABAA-receptor-mediated synaptic currents and responses induced by the GABAA receptor agonist isoguvacine were completely suppressed. Inward GABAergic postsynaptic currents were suppressed prior to outward currents. Synaptic responses mediated by AM PA receptors were not affected by the dialysis, whereas the NM DA-receptor-mediated postsynaptic currents were reduced by approximately 20%. Dialysis with fluoride-based Mg2+, ATP-free solution either fully blocked giant depolarizing potentials (G DPs) in CA3 pyramidal cells (n = 2) or reduced the charge crossing the membrane during G DPs and shifted the G DP reversal potential to more positive values (n = 5). The dialysis-resistant component of G DPs was mediated by glutamate receptors, since: (i) it reversed around 0 mV; (ii) it demonstrated a negative slope conductance at negative membrane voltages, which is characteristic of NM DA receptor-mediated responses; (iii) kinetics of the individual events composing the dialysis-resistant component of G DPs at negative voltages were very similar to those of AM PA receptor-mediated synaptic currents. Thus, this procedure can be useful to isolate the glutamate receptor-mediated component of neuronal network-driven activities

Dynamics of the hypoxia—induced tissue edema in the rat barrel cortex in vitro

© 2018 Juzekaeva, Gainutdinov, Mukhtarov and Khazipov. Cerebral edema is a major, life threatening complication of ischemic brain damage. Previous studies using brain slices have revealed that cellular swelling and a concomitant increase in tissue transparency starts within minutes of the onset of metabolic insult in association with collective anoxic spreading depolarization (aSD). However, the dynamics of tissue swelling in brain slices under ischemia-like conditions remain elusive. Here, we explored the dynamics of brain tissue swelling induced by oxygen-glucose deprivation (OGD) in submerged rat barrel cortex slices. Video monitoring of the vertical and horizontal position of fluorescent dye-filled neurons and contrast slice surface imaging revealed elevation of the slice surface and a horizontal displacement of the cortical tissue during OGD. The OGD-induced tissue movement was also associated with an expansion of the slice borders. Tissue swelling started several minutes after aSD and continued during reperfusion with normal solution. Thirty minutes after aSD, slice borders had expanded by ~130 μm and the slice surface had moved up to attain a height of ~70 μm above control levels, which corresponded to a volume increase of ~30%. Hyperosmotic sucrose solution partially reduced the OGD-induced slice swelling. Thus, OGD-induced cortical slice tissue swelling in brain slices in vitro recapitulates many features of ischemic cerebral edema in vivo, its onset is tightly linked to aSD and it develops at a relatively slow pace after aSD. We propose that this model of cerebral edema in vitro could be useful for the exploration of the pathophysiological mechanisms underlying ischemic cerebral edema and in the search for an efficient treatment to this devastating condition

Microchannel avalanche photodiode with wide linearity range

Design and physical operation principles of new microchannel avalanche photodiode (MC APD) with gain up to 10^5 and linearity range improved an order of magnitude compared to known similar devices. A distinctive feature of the new device is a directly biased p-n junction under each pixel which plays role of an individual quenching resistor. This allows increasing pixel density up to 40000 per mm^2 and making entire device area sensitive.Comment: Submitted to Journal of Technical Physic

Intracellular blockade of GABAA receptors in the rat hippocampal neurons

The intracellular blockade of GABAA-receptor-mediated currents is a useful approach to suppress the GABAergic conductance in a single cell and to isolate the glutamatergic component of network-driven activities. Previously an approach has been described allowing intracellular blockade of GABA A receptors by means of intracellular dialysis of a neuron with the pipette-filling solution, in which fluoride ions that hardly pass through the GABAA receptor channels substitute for Cl- and in which Mg2+ and ATP are omitted to induce rundown of the GABAA receptors during whole-cell patch-clamp recordings. However, the kinetics of suppression of GABAergic conductance and the effect on the currents mediated by glutamate receptors remain unknown. Here, using whole-cell recordings with fluoride-based, Mg2+- and ATP-free solution on CA3 hippocampal neurons of neonatal rats, we show that after 1 h of such dialysis, both spontaneous and evoked GABAA-receptor-mediated synaptic currents and responses induced by the GABAA receptor agonist isoguvacine were completely suppressed. Inward GABAergic postsynaptic currents were suppressed prior to outward currents. Synaptic responses mediated by AMPA receptors were not affected by the dialysis, whereas the NMDA-receptor-mediated postsynaptic currents were reduced by approximately 20%. Dialysis with fluoride-based Mg 2+, ATP-free solution either fully blocked giant depolarizing potentials (GDPs) in CA3 pyramidal cells (n = 2) or reduced the charge crossing the membrane during GDPs and shifted the GDP reversal potential to more positive values (n = 5). The dialysis-resistant component of GDPs was mediated by glutamate receptors, since: (i) it reversed around 0 mV; (ii) it demonstrated a negative slope conductance at negative membrane voltages, which is characteristic of NMDA receptor-mediated responses; (iii) kinetics of the individual events composing the dialysis-resistant component of GDPs at negative voltages were very similar to those of AMPA receptor-mediated synaptic currents. Thus, this procedure can be useful to isolate the glutamate receptor-mediated component of neuronal network-driven activities. © 2014 Pleiades Publishing, Ltd

Postsynaptic GABA(B) receptors contribute to the termination of Giant Depolarizing Potentials in CA3 neonatal rat hippocampus

© 2017 Khalilov, Minlebaev, Mukhtarov, Juzekaeva and Khazipov. During development, hippocampal CA3 network generates recurrent population bursts, so-called Giant Depolarizing Potentials (GDPs). GDPs are characterized by synchronous depolarization and firing of CA3 pyramidal cells followed by afterhyperpolarization (GDP-AHP). Here, we explored the properties of GDP-AHP in CA3 pyramidal cells using gramicidin perforated patch clamp recordings from neonatal rat hippocampal slices. We found that GDP-AHP occurs independently of whether CA3 pyramidal cells fire action potentials (APs) or remain silent during GDPs. However, the amplitude of GDP-AHP increased with the number of APs the cells fired during GDPs. The reversal potential of the GDP-AHP was close to the potassium equilibrium potential. During voltage-clamp recordings, current-voltage relationships of the postsynaptic currents activated during GDP-AHP were characterized by reversal near the potassium equilibrium potential and inward rectification, similar to the responses evoked by the GABA(B) receptor agonists. Finally, the GABA(B) receptor antagonist CGP55845 strongly reduced GDP-AHP and prolonged GDPs, eventually transforming them to the interictal and ictal-like discharges. Together, our findings suggest that the GDP-AHP involves two mechanisms: (i) postsynaptic GABA(B) receptor activated potassium currents, which are activated independently on whether the cell fires or not during GDPs; and (ii) activity-dependent, likely calcium activated potassium currents, whose contribution to the GDP-AHP is dependent on the amount of firing during GDPs. We propose that these two complementary inhibitory postsynaptic mechanisms cooperate in the termination of GDP