A Bioszignál Metrológiai Eljárás alkalmazása az oszcillometriás vérnyomásmérési módszerek mérési bizonytalanságának meghatározása és csökkentése érdekében = Application of the Biosignal Metrology Method to determine and decrease the measurement uncertainity of the oscillometric blood pressure measurement solutions

A klasszikus auszkultatorikus sphygmomanometriás vérnyomásmérés és az automatizált oszcillometriás vérnyomásmérés technológiai alapelvei több évtizede kidolgozásra kerültek és majdnem változatlanok mind az egészségügyi rutin alkalmazások, mind a validációs standardok szintjén. Míg ezek a klasszikus módszerek elfogadhatóak az általános klinikai rutinban, ahol drasztikus patológiás hemodinamikai változásokat vagy azok induló fázisát képesek detektálni, addig a folytonos teljesítmény monitoring alkalmazások során pontosságuk megfelelősége megkérdőjelezhető. A finomabb szabályzás szintű hemodinamikai nyomon követés területén a szerző egy méréstechnológiai értelemben a klaszszikus megoldásra épülő, de méréstudományi és bioinformatikai értelemben újkeletű eljárást javasol, amellyel kontrollálhatóak a mérési bizonytalanság fő forrásai, annak érdekében, hogy a speciális repülésélettani és űrorvostani vizsgálatok során pontosabban, egyénre szabottan lehessen értékelni a hemodinamikai változásokat, támogatva ezzel a döntési mechanizmusokat. = In the field of blood pressure management, the estimated ratio of false positive and false negative diagnoses caused by inappropriate measurement technology is remarkable. Due to the above reason, several attempts have recently been made to harmonize the related ISO/IEC and AAMI protocols, in order to support the development of more accurate evaluation and classification methods. The fundamental standards of the classical auscultatoric sphygmomanometric and oscillometric blood pressure measurement methods have been in use for decades regarding both the clinical routine applications and the validation standards. While these classical methods can detect prominent pathological changes under clinical circumstances, their accuracy is questionable under continuous performance monitoring applications. The author proposes a new approach based on the classical measurement technologies but applying newly developed bioinformatics algorithms. This approach provides the possibility of monitoring minor differences of the haemodynamic regulation and controlling of the major components of the measurement uncertainty so as to support decision making during aviation physiological and aerospace medical examinations

Különböző típusú GABAerg interneuronok szerepe a hippokampális gamma oszcillációkban = The role of distinct types of GABAergic interneurons in hippocampal network oscillations at gamma frequency

A kérgi neuronhálózatokban megfigyelt gamma (30-100 Hz) oszcillációk alapvető szerepet játszanak olyan kognitív folyamatokban, mint pl. a szenzoros információfeldolgozás. Funkciójuk megértéséhez ismernünk kell a neuronhálózatokat alkotó serkentő és gátlósejtek viselkedését ill. szerepét az oszcillációk kialakításában. Pályázatunk célja a hippokampális ideghálózatok szinkronizált működését kialakító sejtszintű mechanizmusok felderítése volt. In vitro farmakológiailag indukált oszcillációk során vizualizált patch-clamp méréstechnika segítségével megállapítottuk, hogy a hippokampusz CA3 régiójában keletkező gamma oszcillációkat a gyorsan tüzelő kosársejtek és a piramissejtek időben összehangolt kisülése generálja szinaptikus visszacsatolás révén. A hippokampusz CA1 régiójába a gamma oszcilláció előrecsatoló gátlással terjed át a CA3 régióból. Mindkét régióban a gátlósejtek oszcillációhoz viszonyított fáziskapcsolt tüzelését a rájuk érkező szinaptikus serkentés, míg a piramissejtek kisülését a szinaptikus gátlás határozta meg. Kifejlesztettünk egy szabadalmi bejelentéssel védett szeletkamrát in vitro mérésekhez, melyben az agyszeletek oxigénellátása megközelíti az in vivo körülményeket. Az eredményeinknek klinikai vonatkozása is elképzelhető, hiszen az epilepszia tünetcsoportban tapasztalt hiperszinkonitás kialakulásában is kulcsfontosságú szerepet játszhatnak a gyorsan tüzelő kosársejtek, amely gátlósejtek működésének célzott szabályozása egy potenciális gyógyszercélpont lehet. | Cortical network oscillations at gamma (30-100 Hz) frequencies were suggested to be linked to several cognitive tasks including sensory processing. To understand the role of oscillations in neuronal operation, the behavior and the function of different neuronal types during oscillatory activities need to be revealed. The aim of our project was to uncover the basic cellular mechanisms generating synchronous network activities in hippocampal neuronal circuitries. The combination of visualized patch-clamp recordings with pharmacologically-induced in vitro oscillations allowed us to determine that in CA3 hippocampal region the precisely timed discharge of fast spiking basket cells and pyramidal cells could generate the gamma oscillations via a synaptic feed-back loop. The gamma oscillation emerged intrinsically in CA3 propagates to CA1 via feed-forward inhibition. In both regions, the phase-coupled firing of inhibitory cells was controlled by synaptic excitation, whereas the discharge of pyramidal cells was primarily determined by synaptic inhibition. For in vitro recordings we developed a new type of slice chamber protected by a patent, where the oxygen supply of brain slices approaches the in vivo circumstances. Our results also have clinical relevance implying the pivotal role of fast spiking basket cells in hypersynchrony during epileptic discharges, therefore the modulation of the fast spiking basket cell operation might be a novel target for drug development

Proteolízis az idegrendszerben: a humán agyi tripszin szerkezetének és funkciójának vizsgálata = Proteolysis in central nervous system: investigations of the structure and function of human brain trypsin

Klónoztunk a humán tripszinogén 4 teljes kódoló szekvenciáját. Bakterális expressziós rendszerben kifejeztük a két alternatív iniciációval keletkező izoformáját. Expresszáltuk és enzimatikusan karakterizáltuk az a tripszin 4-et. Meghatároztuk az aktív enzim benzamidin komplexének térszerkezetét. Feltérképeztük a mRNS és a fehérje eloszlását a humán agy 17 különböző régiójában. Vizsgáltuk az agyban előforduló potenciális szubsztrátjait. Megállapítottuk, hogy számos, a citoszkeletális váz felépítésében és regulálásában szereplő fehérje és a mielin bázikus fehérje szubsztrátja lehet a tripszin 4-nek. Izoláltuk a tripszinogén 4-et emberi agyszövetből és meghatároztuk amino terminális szekvenciáját. Tranziens szövettenyészetben megfelelően tervezett expressziós vektorok segítségével megállapítottuk, hogy az amino terminális leucin beépülését nem-konvencionális CTG iniciátor kodon irányítja. Meghatároztuk a tripszin - metilubellireril guanidinobenzoáttal történő reakciójának elemi sebességi állandóit. Vizsgáltuk számos természetes és szintetikus inhibitor gátló képességét, és valószínű magyarázatot adtunk arra, miért lehetetlen kanonikus mechanizmus alapján gátolni a tripszin 4-et. | We have isolated the complete coding sequence of human trypsinogen 4. In bacterial expression system we expressed and purified the two putative isoforms of the enzyme. We isolated the active enzyme as well, and characterized its enzymatic properties. We determined the X-ray structure of the trypsin 4 ? benzamidine complex. By quantitative real time PCR and sandwich ELISA we determined the determined the distribution of the mRNA and protein in 17 different regions of the human brain. By sequencing the zymogen isolated from human brain we concluded that predominant form in the human brain possesses leucine amino terminus. Using human tissue cultures transiently transfected with appropriately designed expression vectors we proved that the incorporation of the amino terminal leucine is directed by a non-conventional CTG initiation codon. We determined the elementary kinetic constants of the trypsin 4 - methylumbelliferyl guanidine benzoate reaction. We studied the inhibitory potential of several natural and synthetic polypeptides, and we reasoned why is it impossible to inhibit trypsin 4 by a canonical inhibitor

Maintaining network activity in submerged hippocampal slices: importance of oxygen supply

Studies in brain slices have provided a wealth of data on the basic features of neurons and synapses. In the intact brain, these properties may be strongly influenced by ongoing network activity. Although physiologically realistic patterns of network activity have been successfully induced in brain slices maintained in interface-type recording chambers, they have been harder to obtain in submerged-type chambers, which offer significant experimental advantages, including fast exchange of pharmacological agents, visually guided patch-clamp recordings, and imaging techniques. Here, we investigated conditions for the emergence of network oscillations in submerged slices prepared from the hippocampus of rats and mice. We found that the local oxygen level is critical for generation and propagation of both spontaneously occurring sharp wave–ripple oscillations and cholinergically induced fast oscillations. We suggest three ways to improve the oxygen supply to slices under submerged conditions: (i) optimizing chamber design for laminar flow of superfusion fluid; (ii) increasing the flow rate of superfusion fluid; and (iii) superfusing both surfaces of the slice. These improvements to the recording conditions enable detailed studies of neurons under more realistic conditions of network activity, which are essential for a better understanding of neuronal network operation

Integrated Systems Biology Approach Identifies Novel Maternal and Placental Pathways of Preeclampsia

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Phase-amplitude coupled persistent theta and gamma oscillations in rat primary motor cortex in vitro

In vivo, theta (4-7 Hz) and gamma (30-80 Hz) neuronal network oscillations are known to coexist and display phase-amplitude coupling (PAC). However, in vitro, these oscillations have for many years been studied in isolation. Using an improved brain slice preparation technique we have, using co-application of carbachol (10 μM) and kainic acid (150 nM), elicited simultaneous theta (6.6 ± 0.1 Hz) and gamma (36.6 ± 0.4 Hz) oscillations in rodent primary motor cortex (M1). Each oscillation showed greatest power in layer V. Using a variety of time series analyses we detected significant cross-frequency coupling 74% of slice preparations. Differences were observed in the pharmacological profile of each oscillation. Thus, gamma oscillations were reduced by the GABAA receptor antagonists, gabazine (250 nM and 2 μM), and picrotoxin (50 μM) and augmented by AMPA receptor antagonism with SYM2206 (20 μM). In contrast, theta oscillatory power was increased by gabazine, picrotoxin and SYM2206. GABAB receptor blockade with CGP55845 (5 μM) increased both theta and gamma power, and similar effects were seen with diazepam, zolpidem, MK801 and a series of metabotropic glutamate receptor antagonists. Oscillatory activity at both frequencies was reduced by the gap junction blocker carbenoxolone (200 μM) and by atropine (5 μM). These data show theta and gamma oscillations in layer V of rat M1 in vitro are cross-frequency coupled, and are mechanistically distinct. The development of an in vitro model of phase-amplitude coupled oscillations will facilitate further mechanistic investigation of the generation and modulation of coupled activity in mammalian cortex

Reduced Gamma Oscillations in a Mouse Model of Intellectual Disability: A Role for Impaired Repetitive Neurotransmission?

Intellectual disability affects 2-3% of the population; mutations of the X-chromosome are a major cause of moderate to severe cases. The link between the molecular consequences of the mutation and impaired cognitive function remains unclear. Loss of function mutations of oligophrenin-1 (OPHN1) disrupt Rho-GTPase signalling. Here we demonstrate abnormal neurotransmission at CA3 synapses in hippocampal slices from Ophn1-/y mice, resulting from a substantial decrease in the readily releasable pool of vesicles. As a result, synaptic transmission fails at high frequencies required for oscillations associated with cognitive functions. Both spontaneous and KA-induced gamma oscillations were reduced in Ophn1-/y hippocampal slices. Spontaneous oscillations were rapidly rescued by inhibition of the downstream signalling pathway of oligophrenin-1. These findings suggest that the intellectual disability due to mutations of oligophrenin-1 results from a synaptopathy and consequent network malfunction, providing a plausible mechanism for the learning disabilities. Furthermore, they raise the prospect of drug treatments for affected individuals

Bimoclomol improves early electrophysiological signs of retinopathy in diabetic rats

A silent process involving both neural and vascular structures in diabetic retina persists for several years before clinically detectable retinopathy. Recordings of the electroretinogram (ERG) and visual evoked potential (VEP) provide early warning of abnormalities in the visual pathway of diabetic patients and animal models. Treatment of streptozotocin-diabetic rats for 1 or 2 months with the heat-shock protein coinducer bimoclomol, a drug ameliorating experimental neuropathy, prevented and corrected the abnormal increase in latency and reduction of amplitude of ERG and VEP waves both in acute and chronic experiments. Improvements may be explained by cytoprotective effect of bimoclomol on retinai glia and/or neurons against diabetes-related ischemic cell damages. These findings suggest that bimoclomol may have future therapeutic use in diabetic retinopathy

Spike timing of distinct types of GABAergic interneuron during hippocampal gamma oscillations in vitro.

Gamma frequency (30-100 Hz) network oscillations occur in the intact hippocampus during awake, attentive behavior. Here, we explored the underlying cellular mechanisms in an in vitro model of persistent gamma-frequency oscillations, induced by bath application of 20 microm carbachol in submerged hippocampal slices at 30 +/- 1 degrees C. Current-source density analysis of the field oscillation revealed a prominent alternating sink-source pair in the perisomatic and apical dendritic regions of CA3. To elucidate the active events generating these extracellular dipoles, we examined the firing properties of distinct neuron types. Visually guided unit recordings were obtained from individual CA3 neurons followed by intracellular labeling for anatomical identification. Pyramidal cells fired at 2.82 +/- 0.7 Hz, close to the negative peak of the oscillation (0.03 +/- 0.65 msec), and often in conjunction with a negative spike-like component of the field potential. In contrast, all phase-coupled interneurons fired after this negative peak. Perisomatic inhibitory interneurons fired at high frequency (18.1 +/- 2.7 Hz), shortly after the negative peak (1.97 +/- 0.95 msec) and were strongly phase-coupled. Dendritic inhibitory interneurons fired at lower frequency (8.4 +/- 2.4 Hz) and with less fidelity and a longer delay after the negative peak (4.3 +/- 1.1 msec), whereas interneurons with cell body in the stratum radiatum often showed no phase relationship with the field oscillation. The phase and spike time data of individual neurons, together with the current-source density analysis, support a synaptic feedback model of gamma oscillations primarily involving pyramidal cells and inhibitory cells targeting their perisomatic region