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Local circuit architecture facilitates the emergence of feature selectivity in the cerebral cortex1. In the hippocampus, it remains unknown whether local computations supported by specific connectivity motifs2 regulate the spatial receptive fields of pyramidal cells3. Here we developed an in vivo electroporation method for monosynaptic retrograde tracing4 and optogenetics manipulation at single-cell resolution to interrogate the dynamic interaction of place cells with their microcircuitry during navigation. We found a local circuit mechanism in CA1 whereby the spatial tuning of an individual place cell can propagate to a functionally recurrent subnetwork5 to which it belongs. The emergence of place fields in individual neurons led to the development of inverse selectivity in a subset of their presynaptic interneurons, and recruited functionally coupled place cells at that location. Thus, the spatial selectivity of single CA1 neurons is amplified through local circuit plasticity to enable effective multi-neuronal representations that can flexibly scale environmental features locally without degrading the feedforward input structure. Single-cell tracing and optogenetics manipulation in mice are used to show how spatial tuning of individual pyramidal cells in CA1 can propagate to and be amplified by their local subnetwork of neurons.

Hepatocellular carcinoma (HCC) is the leading primary liver tumor and a main indication for transplant. Transplant criteria are based on clinicopathologic features, meanwhile adequate downstaging and molecular mechanisms are getting more attention in evolving therapeutic algorithm of HCC. The aim of our study was to overview the results of the Hungarian Liver Transplant Program in the field of HCC and introduce new aspects of personalized treatment options.We performed retrospective analysis of survival and tumor recurrence of HCC-associated liver transplant recipients between October 2013 and December 2020. Patients were categorized in Milan criteria (MC), beyond MC but within University of California, San Francisco (UCSF), and beyond UCSF criteria groups after pathologic examination of the explanted liver. Demographic data and preoperative locoregional treatments were assessed.A total of 529 primer liver transplants were performed, 88 because of HCC. A total of 87 patients had underlying cirrhosis because of hepatitis C (54%), alcohol-related liver disease (33.7%), hepatitis B (4.5%), or unknown etiology. A total of 55.6% of the patients had at least one locoregional treatment. A total of 67.4% of the patients were within MC, 5.6% were within UCSF criteria, and 27% were beyond UCSF criteria. The 1-, 3-, and 5-year survival rates were 80%, 79%, and 75%. The outcome was better in early-stage tumors, but the difference was not significant (P = .745) CONCLUSIONS: The favorable survival in our department legitimates the strict transplant criteria of HCC. Adequate locoregional therapy as downstaging can expand recipient pool. Molecular tumor profiling may lead to personalized treatment of HCC.

SummaryCortical circuits are composed predominantly of pyramidal-to-pyramidal neuron connections, yet their assembly during embryonic development is not well understood. We show that embryonic layer 5 pyramidal neurons, identified through single cell transcriptomics, display two phases of circuit assembly in vivo. At E14.5, a multi-layered circuit motif, composed of a single layer 5 cell type, forms. This motif is transient, switching to a second circuit motif, involving all three types, by E17.5. In vivo targeted single cell recordings and two-photon calcium imaging of embryonic layer 5 neurons reveal that, in both phases, neurons have active somas and neurites, tetrodotoxin-sensitive voltage-gated conductances, and functional glutamatergic synapses. Embryonic layer 5 neurons strongly express autism-associated genes, and perturbing these genes disrupts the switch between the two motifs. Hence, layer 5 pyramidal neurons form transient active pyramidal-to-pyramidal circuits, at the inception of neocortex, and studying these circuits could yield insights into the etiology of autism.

SUMMARYReward and punishment powerfully inform ongoing behaviors and drive learning throughout the brain, including neocortex. Yet it remains elusive how these global signals are represented and impact local cortical computations. Previously we found that in auditory cortex, VIP-expressing interneurons are recruited by reinforcement feedback. Here, we used 3D random-access two-photon microscopy and fiber photometry to monitor VIP neural activity in dozens of cortical areas while mice learned an auditory decision task. We show that reward and punishment evoke a rapid, cortex-wide activation of most VIP interneurons. This global recruitment mode of VIP interneurons showed variations in temporal dynamics in individual neurons and across areas. Neither their weak sensory tuning in visual cortex, nor their arousal state modulation was predictive of reinforcer responses of VIP interneurons. We suggest that VIP-expressing cortical inhibitory neurons transduce global reinforcement signals to provide disinhibitory control over local circuit computations and their plasticity.

Psoriasis is a chronic inflammatory disease with unmet medical needs. To clarify potential therapeutic targets, different animal models have been developed. In the current study, imiquimod-induced psoriasiform dermatitis was used for monitoring the changes in skin thickness, transepidermal water loss, body weight, blood perfusion and drug permeability for a topical cream formulation of caffeine, both in wild type and in knock out mice. Morphological characterization of control and diseased tissues was performed by scanning electron microscopy and two-photon microscopy. The chemically induced psoriatic group showed increased skin permeability for the model drug during disease progression. In wild type and TRPA1 KO mice, however, enhanced skin thickness and hyperkeratosis blocked further increase of drug penetration at the late phase (96 h). These results indicate that topical drug therapy can be more effective in early phases of plaque development, when skin thickness is lower. Although paracellular connections (tight junctions) are looser in the advanced phase, hyperkeratosis blocks drug delivery through the transappendageal routes. Novel drug formulations may have the potency for effective drug delivery across the epidermal barrier even in the advanced phase. For development of more effective topical drugs, further research is proposed to explore drug penetration both in healthy and diseased conditions.

Psoriasis is a chronic inflammatory disease with unmet medical needs. To clarify potential therapeutic targets, different animal models have been developed. In the current study, imiquimod-induced psoriasiform dermatitis was used for monitoring the changes in skin thickness, transepidermal water loss, body weight, blood perfusion and drug permeability for a topical cream formulation of caffeine, both in wild type and in knock out mice. Morphological characterization of control and diseased tissues was performed by scanning electron microscopy and two-photon microscopy. The chemically induced psoriatic group showed increased skin permeability for the model drug during disease progression. In wild type and TRPA1 KO mice, however, enhanced skin thickness and hyperkeratosis blocked further increase of drug penetration at the late phase (96 h). These results indicate that topical drug therapy can be more effective in early phases of plaque development, when skin thickness is lower. Although paracellular connections (tight junctions) are looser in the advanced phase, hyperkeratosis blocks drug delivery through the transappendageal routes. Novel drug formulations may have the potency for effective drug delivery across the epidermal barrier even in the advanced phase. For development of more effective topical drugs, further research is proposed to explore drug penetration both in healthy and diseased conditions.

In this paper, we present an additional, new cage-GABA compound, called 4-amino-1-(4′-dimethylaminoisopropoxy-5′,7′-dinitro-2′,3′-dihydro-indol-1-yl)-1-oxobutane-γ-aminobutyric acid (iDMPO-DNI-GABA), and currently, this compound is the only photoreagent, which can be applied for GABA uncaging without experimental compromises. By a systematic theoretical design and successful synthesis of several compounds, the best reagent exhibits a high two-photon efficiency within the 700–760 nm range with excellent pharmacological behavior, which proved to be suitable for a complex epileptic study. Quantum chemical design showed that the optimal length of the cationic side chain enhances the two-photon absorption by 1 order of magnitude due to the cooperating internal hydrogen bonding to the extra nitro group on the core. This feature increased solubility while suppressing membrane permeability. The efficiency was demonstrated in a systematic, wide range of in vitro single-cell neurophysiological experiments by electrophysiological as well as calcium imaging techniques. Scalable inhibitory ion currents were elicited by iDMPO-DNI-GABA with appropriate spatial–temporal precision, blocking both spontaneous and evoked cell activity with excellent efficiency. Additionally, to demonstrate its applicability in a real neurobiological study, we could smoothly and selectively modulate neuronal activities during artificial epileptic rhythms first time in a neural network of GCaMP6f transgenic mouse brain slices.

To understand brain function, we have developed a multiphoton approach based on acousto-optics (AO) to simultaneously measure and optically stimulate the activity of cortical neurons, even during behavior, with millisecond precision and subcellular spatial resolution.

Infrared neural stimulation is a promising tool for stimulating the brain because it can be used to excite with high spatial precision without the need of delivering or inserting any exogenous agent into the tissue. Very few studies have explored its use in the brain, as most investigations have focused on sensory or motor nerve stimulation. Using intravital calcium imaging with the genetically encoded calcium indicator GCaMP6f, here we show that the application of infrared neural stimulation induces intracellular calcium signals in Layer 2/3 neurons in mouse cortex in vivo. The number of neurons exhibiting infrared-induced calcium response as well as the amplitude of those signals are shown to be both increasing with the energy density applied. By studying as well the spatial extent of the stimulation, we show that reproducibility of the stimulation is achieved mainly in the central part of the infrared beam path. Stimulating in vivo at such a degree of precision and without any exogenous chromophores enables multiple applications, from mapping the brain’s connectome to applications in systems neuroscience and the development of new therapeutic tools for investigating the pathological brain.

Dendritic hotspots have been shown to play a determining role as local activity foci during sensory information coding and memory formation in cortical neurons in vivo. Here we aim to reveal the characteristics of co-activating more dendritic hotspots themselves within a dynamic time window. Using electrical stimulation and patterned two-photon uncaging for the coincident activation of two neighboring dendritic hotspots in rat hippocampal CA1 interneurons, the evoked local dendritic Ca2+ and the associated excitatory postsynaptic potentials (EPSPs) summed supralinearly. Between active hotspots, supralinearity was present in a relatively broad temporal (∼10 ms) and spatial (∼30 µm) window, mediated by NMDA receptors. Though at larger inter-hotspot distances (>30 µm) supralinear summation of EPSPs was replaced by sublinear integration, the summation of dendritic Ca2+ responses remained supralinear. The inter-hotspot integration represents a novel signal integration state, extending the interaction distance between dendritic inputs, and allowing long-range coincidence detection.

International audience; Functional mapping of the connectivity of the brain is an essential procedure for neurosurgeons aiming for maximal resection of a brain tumor, while minimizing damage to the eloquent cortex. Being able to delineate the location of critical cortical areas in individual patients allows the surgeon to preserve sensorimotor and cognitive functions. Several methods exist for functional mapping, however, none are capable of delivering fast, label free mapping of the cortex with millimeter precision. In this work, we investigate the potential use of infrared neural stimulation (λ=1470nm) as a means of focal stimulation of the mouse cortex in vivo. Here, we show that the application of infrared neural stimulation induces in vivo cortical intracellular calcium signals in Layer II/III mouse neurons, using intravital calcium imaging with the genetically encoded calcium indicator GCaMP6f.

Abstract: Machine perfusion of marginal grafts might be a possible solution to organ shortage and a promising tool for reducing waiting list morbidity and mortality. In recent years, optimizing the circumstances of organ preservation prior to implantation via machine perfusion has become a hot topic of research. Machine perfusion offers a platform for organ reconditioning, assessment of cell viability and function, pharmacological preconditioning, prolongation of preservation time (ischemia time) and finally reducing graft injury. The objective of the new technology is to increase the pool of transplantable organs safely. Multicentric prospective studies have been evaluating the short and long term outcomes of different methods, however, several questions still remain unanswered. This review summarizes the recent advances in the field of machine perfusion, focusing on preclinical and clinical results. Machine perfusion seems to be a new milestone in the modern era of solid organ transplantation. Orv Hetil. 2018; 159(46): 1882–1890.

How neuronal computations in the sensory periphery contribute to computations in the cortex is not well understood. We examined this question in the context of visual-motion processing in the retina and primary visual cortex (V1) of mice. We disrupted retinal direction selectivity, either exclusively along the horizontal axis using FRMD7 mutants or along all directions by ablating starburst amacrine cells, and monitored neuronal activity in layer 2/3 of V1 during stimulation with visual motion. In control mice, we found an over-representation of cortical cells preferring posterior visual motion, the dominant motion direction an animal experiences when it moves forward. In mice with disrupted retinal direction selectivity, the over-representation of posterior-motion-preferring cortical cells disappeared, and their responses at higher stimulus speeds were reduced. This work reveals the existence of two functionally distinct, sensory-periphery-dependent and -independent computations of visual motion in the cortex.

The simultaneous utilization of electrophysiological recordings and two-photon imaging allows the observation of neural activity in a high temporal and spatial resolution at the same time. The three dimensional monitoring of morphological features near the microelectrode array makes the observation more precise and complex. In vitro experiments were performed on mice neocortical slices expressing the GCaMP6 genetically encoded calcium indicator for monitoring the neural activity with two-photon microscopy around the implanted microelectrodes. A special filtering algorithm was used for data analysis to eliminate the artefacts caused by the imaging laser. Utilization of a special filtering algorithm allowed us to detect and sort single unit activities from simultaneous two-photon imaging and electrophysiological measurement.

Visual Abstract, Transparent and flexible materials are attractive for a wide range of emerging bioelectronic applications. These include neural interfacing devices for both recording and stimulation, where low electrochemical electrode impedance is valuable. Here the conducting polymer poly(3,4-ethylenedioxythiophene):poly(styrenesulfonate) (PEDOT:PSS) is used to fabricate electrodes that are small enough to allow unencumbered optical access for imaging a large cell population with two-photon (2P) microscopy, yet provide low impedance for simultaneous high quality recordings of neural activity in vivo. To demonstrate this, pathophysiological activity was induced in the mouse cortex using 4-aminopyridine (4AP), and the resulting electrical activity was detected with the PEDOT:PSS-based probe while imaging calcium activity directly below the probe area. The induced calcium activity of the neuronal network as measured by the fluorescence change in the cells correlated well with the electrophysiological recordings from the cortical grid of PEDOT:PSS microelectrodes. Our approach provides a valuable vehicle for complementing classical high temporal resolution electrophysiological analysis with optical imaging.

Simultaneously with fast 3D imaging, we generated holograms from multiple spots by introducing sine waves with non-linear frequency chirps into the acousto-optical deflectors and activated over 40cells with high resolution within 16ms in large volumes.

In order to improve the fluorescence properties of the green fluorescent protein chromophore, p‑HOBDI ((5‑(4‑hydroxybenzylidene)‑2,3‑dimethyl‑3,5‑dihydro‑4H‑imidazol‑4‑one), sixteen dihydroimidazolone derivates were synthesized from thiohydantoin and arylaldehydes. The synthesis developed is an efficient, novel, one-pot procedure. The study provides a detailed description of the spectroscopic characteristics of the newly synthesized compounds, using p‑HOBDI as a reference. The new compounds all exhibited significantly stronger fluorescence than p‑HOBDI, up to 28 times higher quantum yields. An experimental and theoretical investigation of the relationship of the fluorescence properties with the molecular structure was also carried out. A good correlation was found between the emission wavenumber and the Hammett constant of the functional group, which suggests the intermolecular charge transfer (ICT) mechanism between the aromatic groups.

Communication between individual GABAergic cells and their target neurons is mediated by synapses and, in the case of neurogliaform cells (NGFCs), by unitary volume transmission. Effects of non-synaptic volume transmission might involve non-neuronal targets, and astrocytes not receiving GABAergic synapses but expressing GABA receptors are suitable for evaluating this hypothesis. Testing several cortical interneuron types in slices of the rat cerebral cortex, we show selective unitary transmission from NGFCs to astrocytes with an early, GABAA receptor and GABA transporter-mediated component and a late component that results from the activation of GABA transporters and neuronal GABAB receptors. We could not detect Ca2+ influx in astrocytes associated with unitary GABAergic responses. Our experiments identify a presynaptic cell-type-specific, GABA-mediated communication pathway from individual neurons to astrocytes, assigning a role for unitary volume transmission in the control of ionic and neurotransmitter homeostasis.

Machine perfusion of marginal grafts might be a possible solution to organ shortage and a promising tool for reducing waiting list morbidity and mortality. In recent years, optimizing the circumstances of organ preservation prior to implantation via machine perfusion has become a hot topic of research. Machine perfusion offers a platform for organ reconditioning, assessment of cell viability and function, pharmacological preconditioning, prolongation of preservation time (ischemia time) and finally reducing graft injury. The objective of the new technology is to increase the pool of transplantable organs safely. Multicentric prospective studies have been evaluating the short and long term outcomes of different methods, however, several questions still remain unanswered. This review summarizes the recent advances in the field of machine perfusion, focusing on preclinical and clinical results. Machine perfusion seems to be a new milestone in the modern era of solid organ transplantation. Orv Hetil. 2018; 159(46): 1882-1890.Absztrakt: A modern transzplantációban a marginális szervek gépi perfúziója jelenthet egy lehetséges választ a várólistán lévő betegek növekvő halálozása és morbiditása miatt világszerte fokozódó szervigényre. A szervek beültetését megelőző aktív szervkonzerválás, a tárolás körülményeinek optimalizálása az utóbbi évek transzplantációs kutatásainak középpontjába került. A gépi perfúzió lehetőséget teremthet a szervek prezervációs károsodásának csökkentésére, rekondicionálására, a funkcionális paraméterek és biomarkerek beültetést megelőző értékelésére, a konzerválás időtartamának növelésére, valamint további terápiás eljárások egyidejű alkalmazására. Az új technológiák célja a transzplantációt követő szervfunkció javítása és a biztonsággal beültethető donorszervek számának növelése. A gépi perfúzió rövid és hosszú távú eredményeit multicentrikus vizsgálatok kutatják, a különböző módszerek előnyeiről számos kérdés maradt még megválaszolatlan. Közleményünkben összegezzük a gépi szervkonzerválás eddigi vívmányait, a közelmúlt legfontosabb preklinikai és klinikai kutatási eredményeit, melyek alapján a gépi perfúziót a transzplantáció újabb mérföldkövének tekinthetjük. Orv Hetil. 2018; 159(46): 1882–1890.

In the rodent olfactory bulb the smooth dendrites of the principal glutamatergic mitral cells (MCs) form reciprocal dendrodendritic synapses with large spines on GABAergic granule cells (GC), where unitary release of glutamate can trigger postsynaptic local activation of voltage-gated Na+-channels (Navs), i.e. a spine spike. Can such single MC input evoke reciprocal release? We find that unitary-like activation via two-photon uncaging of glutamate causes GC spines to release GABA both synchronously and asynchronously onto MC dendrites. This release indeed requires activation of Navs and high-voltage-activated Ca2+-channels (HVACCs), but also of NMDA receptors (NMDAR). Simulations show temporally overlapping HVACC- and NMDAR-mediated Ca2+-currents during the spine spike, and ultrastructural data prove NMDAR presence within the GABAergic presynapse. This cooperative action of presynaptic NMDARs allows to implement synapse-specific, activity-dependent lateral inhibition and thus could provide an efficient solution to combinatorial percept synthesis in a sensory system with many receptor channels.