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Understanding the routing of neuronal information requires the functional characterization of connections. Neuronal projections recruit large postsynaptic ensembles with distinct postsynaptic response types (PRTs). PRT is probed by low-throughput whole-cell electrophysiology and is not a selection criterion for single-cell RNA-sequencing (scRNA-seq). To overcome these limitations and target neurons based on specific PRTs for soma harvesting and subsequent scRNA-seq we created Voltage-Seq. To test our methodology, we established all-optical voltage imaging and recorded the PRT of 8347 periaqueductal gray (PAG) neurons evoked by the optogenetic activation of ventromedial hypothalamic (VMH) terminals. PRTs were classified and spatially resolved in the entire VMH-PAG connectome. We built an on-site analysis named VoltView to navigate soma harvesting towards target PRTs guided by a classifier which used the VMH-PAG connectome database as a reference. We demonstrated the agility of Voltage-Seq in locating VMH-driven GABAergic neurons in the PAG, solely guided by the on-site classification in VoltView.

Inhibitory interneurons expressing parvalbumin (PV) in the prefrontal cortex (PFC) are central to excitatory/inhibitory (E/I) balance, generation of gamma oscillations, and cognition. Dysfunction of PV interneurons disrupts information processing and cognitive behavior. Tyrosine receptor kinase B (trkB) signaling is known to regulate the differentiation and maturation of cortical PV interneurons during development, but is also suggested to be involved in the activity and network functions of PV interneurons in the adult brain. Using a novel viral strategy for cell-type and region-specific expression of a dominant negative trkB in adult mice, we show that reduced trkB signaling in PV interneurons in the PFC leads to pronounced morphological, physiological, and behavioral changes. Our results provide evidence for a critical role of trkB signaling in the function of PV interneurons in the adult brain, local network activities central to prefrontal circuit dynamics, and cognitive behavior.

Encoding and predicting aversive events are critical functions of circuits that support survival and emotional well-being. Maladaptive circuit changes in emotional valence processing can underlie the pathophysiology of affective disorders. The lateral habenula (LHb) has been linked to aversion and mood regulation through modulation of the dopamine and serotonin systems. We have defined the identity and function of glutamatergic (Vglut2) control of the LHb, comparing the role of inputs originating in the globus pallidus internal segment (GPi), and lateral hypothalamic area (LHA), respectively. We found that LHb-projecting LHA neurons, and not the proposed GABA/glutamate co-releasing GPi neurons, are responsible for encoding negative value. Monosynaptic rabies tracing of the presynaptic organization revealed a predominantly limbic input onto LHA Vglut2 neurons, while sensorimotor inputs were more prominent onto GABA/glutamate co-releasing GPi neurons. We further recorded the activity of LHA Vglut2 neurons, by imaging calcium dynamics in response to appetitive versus aversive events in conditioning paradigms. LHA Vglut2 neurons formed activity clusters representing distinct reward or aversion signals, including a population that responded to mild foot shocks and predicted aversive events. We found that the LHb-projecting LHA Vglut2 neurons encode negative valence and rapidly develop a prediction signal for negative events. These findings establish the glutamatergic LHA-LHb circuit as a critical node in value processing.

Sensory perception depends on neocortical computations that contextually adjust sensory signals in different internal and environmental contexts. Neocortical layer 1 (L1) is the main target of cortical and subcortical inputs that provide “top-down” information for context-dependent sensory processing. Although L1 is devoid of excitatory cells, it contains the distal “tuft” dendrites of pyramidal cells (PCs) located in deeper layers. L1 also contains a poorly characterized population of GABAergic interneurons (INs), which regulate the impact that different top-down inputs have on PCs. A poor comprehension of L1 IN subtypes and how they affect PC activity has hampered our understanding of the mechanisms that underlie contextual modulation of sensory processing. We used novel genetic strategies in male and female mice combined with electrophysiological and morphological methods to help resolve differences that were unclear when using only electrophysiological and/or morphological approaches. We discovered that L1 contains four distinct populations of INs, each with a unique molecular profile, morphology, and electrophysiology, including a previously overlooked IN population (named here “canopy cells”) representing 40% of L1 INs. In contrast to what is observed in other layers, most L1 neurons appear to be unique to the layer, highlighting the specialized character of the signal processing that takes place in L1. This new understanding of INs in L1, as well as the application of genetic methods based on the markers described here, will enable investigation of the cellular and circuit mechanisms of top-down processing in L1 with unprecedented detail.SIGNIFICANCE STATEMENTNeocortical layer 1 (L1) is the main target of corticocortical and subcortical projections that mediate top-down or context-dependent sensory perception. However, this unique layer is often referred to as “enigmatic” because its neuronal composition has been difficult to determine. Using a combination of genetic, electrophysiological, and morphological approaches that helped to resolve differences that were unclear when using a single approach, we were able to decipher the neuronal composition of L1. We identified markers that distinguish L1 neurons and found that the layer contains four populations of GABAergic interneurons, each with unique molecular profiles, morphologies, and electrophysiological properties. These findings provide a new framework for studying the circuit mechanisms underlying the processing of top-down inputs in neocortical L1.

Psychostimulant use is an ever-increasing socioeconomic burden, including a dramatic rise during pregnancy. Nevertheless, brain-wide effects of psychostimulant exposure are incompletely understood. Here, we performed Fos-CreERT2-based activity mapping, correlated for pregnant mouse dams and their fetuses with amphetamine, nicotine and caffeine applied acutely during mid-gestation. While light-sheet microscopy-assisted intact tissue imaging revealed drug- and age-specific neuronal activation, the indusium griseum (IG) appeared indiscriminately affected. By using GAD67gfp/+ mice we subdivided the IG into a dorsolateral domain populated by GABA interneurons and a ventromedial segment containing glutamatergic neurons, many showing drug-induced activation and sequentially expressing Pou3f3/Brn1 and secretagogin (Scgn) during differentiation. We then combined Patch-seq and circuit mapping to show that the ventromedial IG is a quasi-continuum of glutamatergic neurons (IG-Vglut1+) reminiscent to dentate granule cells in both rodents and humans, whose dendrites emanate perpendicularly towards, while their axons course parallel with the superior longitudinal fissure. IG-Vglut1+ neurons receive Vglut1+ and Vglut2+ excitatory afferents that topologically segregate along their somatodendritic axis. In turn, their efferents terminate in the olfactory bulb, thus being integral to a multi-synaptic circuit that could feed information antiparallel to the olfactory-cortical pathway. In IG-Vglut1+ neurons, prenatal psychostimulant exposure delayed the onset of Scgn expression. Genetic ablation of Scgn was then found to sensitize adult mice towards methamphetamine-induced epilepsy, suggesting a role for this Ca2+-binding protein in scaling IG-Vglut1+ neuronal excitability. Overall, our study identifies brain-wide targets of the most common psychostimulants, among which Scgn+/Vglut1+ neurons of the IG link limbic and olfactory circuits.Significance statementDrug abuse during pregnancy is a significant socioeconomic problem. The use of psychostimulants is particularly common during pregnancy even though a risk to the developing fetus is significant. Here, we show that short-lived exposure to amphetamine, nicotine and caffeine during pregnancy induces neuronal activation in the fetal brain with the indusium griseum (IG), a brain area situated parallel to the central surface of the cortical hemispheres, becoming indiscriminately activated. By using mouse genetics, we find that psychostimulants preferentially target glutamatergic IG neurons, and delay their differentiation postnatally. Notably, the expressional onset of secretagogin, a Ca2+-sensor amenable for synaptic integration, is deregulated. This is significant because these neurons are integral to a multi-synaptic neuronal pathway that links limbic and olfactory circuits. As such, genetic deletion of secretagogin brings about heightened sensitivity to psychostimulants, manifesting as epileptiform discharges. Cumulatively, we describe a novel psychostimulant-sensitive neuronal subtype and its circuit arrangement whose developmental delay seems critical for behavioral abnormalities in offspring prenatally exposed to the most common psychostimulants.

SUMMARYThe striatum is organized into two major outputs formed by striatal projection neuron (SPN) subtypes with distinct molecular identities. In addition, the histochemical division into patch and matrix compartments represents an additional spatial organization, proposed to mirror a functional specialization in a motor-motivation dimension. To map the molecular diversity of SPNs in the context of the patch and matrix division, we genetically labeled mu-opioid receptor (Oprm1) expressing striatal neurons and performed single-nucleus RNA sequencing (snRNA-seq). This allowed us to establish new molecular definitions of the patch-matrix compartments, resulting in a molecular code for mapping patch SPNs at the cellular level. In addition, Oprm1 expression labeled exopatch SPNs, which we found to be molecularly distinct from both patch as well as neighboring matrix SPNs, thereby forming a separate molecular entity. At the cell-type level, we found an unexpected SPN diversity, leading to the identification of a new Col11a1+ striatonigral SPN type. At the tissue level, we found that mapping the spatial expression of a number of markers revealed new definitions of spatial domains in the striatum, which were conserved in the non-human primate brain. Interestingly, the spatial markers were cell-type independent and instead represented a spatial code that was found across all SPNs within a spatially restricted domain. This spatiomolecular map establishes a formal system for targeting and studying the striatal subregions and SPNs subtypes, beyond the classical striatonigral and striatopallidal division.

Psychostimulant use is an ever-increasing socioeconomic burden, including a dramatic rise during pregnancy. Nevertheless, brain-wide effects of psychostimulant exposure are incompletely understood. Here, we performed Fos-CreER(T2)–based activity mapping, correlated for pregnant mouse dams and their fetuses with amphetamine, nicotine, and caffeine applied acutely during midgestation. While light-sheet microscopy-assisted intact tissue imaging revealed drug- and age-specific neuronal activation, the indusium griseum (IG) appeared indiscriminately affected. By using GAD67(gfp/+) mice we subdivided the IG into a dorsolateral domain populated by γ-aminobutyric acidergic interneurons and a ventromedial segment containing glutamatergic neurons, many showing drug-induced activation and sequentially expressing Pou3f3/Brn1 and secretagogin (Scgn) during differentiation. We then combined Patch-seq and circuit mapping to show that the ventromedial IG is a quasi-continuum of glutamatergic neurons (IG-Vglut1(+)) reminiscent of dentate granule cells in both rodents and humans, whose dendrites emanate perpendicularly toward while their axons course parallel with the superior longitudinal fissure. IG-Vglut1(+) neurons receive VGLUT1(+) and VGLUT2(+) excitatory afferents that topologically segregate along their somatodendritic axis. In turn, their efferents terminate in the olfactory bulb, thus being integral to a multisynaptic circuit that could feed information antiparallel to the olfactory–cortical pathway. In IG-Vglut1(+) neurons, prenatal psychostimulant exposure delayed the onset of Scgn expression. Genetic ablation of Scgn was then found to sensitize adult mice toward methamphetamine-induced epilepsy. Overall, our study identifies brain-wide targets of the most common psychostimulants, among which Scgn(+)/Vglut1(+) neurons of the IG link limbic and olfactory circuits.

Aims Brain ischaemia models are essential to study the pathomechanisms of stroke. Our aim was to investigate the reliability and reproducibility of our novel focal ischaemia-reperfusion model. Methods To induce a cortical transient ischaemic attack, we lifted the distal middle cerebral artery (MCA) with a special hook. The early changes after 2 × 15-min occlusion were observed in the somatosensory evoked responses (SERs). The histological responses to 2 × 15-min MCA occlusion and to 30-, 45- or 60-min ischaemia were examined after a 1-day survival period by 2,3,5-triphenyltetrazolium chloride (TTC) and Fluoro Jade C (FJC) staining. Another group, with 30-min ischaemia, was analysed histologically by FJC, S100 and CD11b labelling after a 5-day survival period. Results The amplitudes of the SERs decreased immediately at the beginning of the ischaemic period, and remained at a reduced level during the ischaemia. Reperfusion resulted in increasing SER amplitudes, but they never regained the control level. The short-lasting ischaemia did not lead to brain infarction when evaluated with TTC, but intense labelling was found with FJC. The 30-min ischaemia did not result in FJC labelling after 1 day, but marked labelling was observed after 5 days with FJC, S100 and CD11b in the cortical area supplied by the MCA. Conclusions We present here a novel, readily reproducible method to induce focal brain ischaemia. The ischaemia-reperfusion results in noteworthy changes in the SERs and the appearance of conventional tissue damage markers. This method involves possibilities for precise blood flow regulation, and the setting of the required level of perfusion.

Although extensively studied postnatally, the functional differentiation of cholecystokinin (CCK)-containing interneurons en route towards the cerebral cortex during fetal development is incompletely understood. Here, we used CCKBAC/DsRed mice encoding a CCK promoter-driven red fluorescent protein to analyze the temporal dynamics of DsRed expression, neuronal identity, and positioning through high-resolution developmental neuroanatomy. Additionally, we developed a dual reporter mouse line (CCKBAC/DsRed::GAD67gfp/+) to differentiate CCK-containing interneurons from DsRed+ principal cells during prenatal development. We show that DsRed is upregulated in interneurons once they exit their proliferative niche in the ganglionic eminence and remains stably expressed throughout their long-distance migration towards the cerebrum, particularly in the hippocampus. DsRed+ interneurons, including a cohort coexpressing calretinin, accumulated at the palliosubpallial boundary by embryonic day 12.5. Pioneer DsRed+ interneurons already reached deep hippocampal layers by embryonic day 14.5 and were morphologically differentiated by birth. Furthermore, we probed migrating interneurons entering and traversing the cortical plate, as well as stationary cells in the hippocampus by patch-clamp electrophysiology to show the first signs of Na+ and K+ channel activity by embryonic day 12.5 and reliable adult-like excitability by embryonic day 18.5. Cumulatively, this study defines key positional, molecular, and biophysical properties of CCK+ interneurons in the prenatal brain.

Global forebrain ischemia results in damage to the pyramids in the CA1 hippocampal subfield, which is particularly vulnerable to excitotoxic processes. Morphological and functional disintegration of this area leads to a cognitive dysfunction and neuropsychiatric disorders. Treatment with N-methyl-d-aspartate receptor antagonists is a widely accepted method with which to stop the advance of excitotoxic processes and concomitant neuronal death. From a clinical aspect, competitive glycine- and polyamine-site antagonists with relatively low affinity and moderate side-effects are taken into account. Endogenous kynurenic acid acts as an antagonist on the obligatory co-agonist glycine site, and has long been at the focus of neuroprotective trials. In the present study, we estimated the neuroprotective capability of a novel kynurenic acid analog in transient global forebrain ischemia, measuring the rate of hippocampal CA1 pyramidal cell loss and the preservation of long-term potentiation at Schaffer collateral-CA1 synapses. The neuroprotective potential was reflected by a significantly diminished hippocampal CA1 cell loss and preserved long-term potentiation expression. The neuroprotective effect was robust in the event of pretreatment, and also when the drug was administered at the time of reperfusion. This result is beneficial since a putative neuroprotectant proven to be effective as post-treatment is of much greater benefit.

Various acute brain pathological conditions are characterized by the presence of elevated glutamate concentrations in the brain interstitial fluids. It has been established that a decrease in the blood glutamate level enhances the brain-to-blood efflux of glutamate, removal of which from the brain may prevent glutamate excitotoxicity and its contribution to the long-lasting neurological deficits seen in stroke. A decrease in blood glutamate level can be achieved by exploiting the glutamate-scavenging properties of the blood-resident enzyme glutamate-oxaloacetate transaminase, which transforms glutamate into 2-ketoglutarate in the presence of the glutamate co-substrate oxaloacetate. The present study had the aim of an evaluation of the effects of the blood glutamate scavenger oxaloacetate on the impaired long-term potentiation (LTP) induced in the 2-vessel occlusion ischaemic model in rat. Transient (30-min) incomplete forebrain ischaemia was produced 72 h before LTP induction. Although the short transient brain hypoperfusion did not induce histologically identifiable injuries in the CA1 region (Fluoro-Jade B, S-100 and cresyl violet), it resulted in an impaired LTP function in the hippocampal CA1 region without damaging the basal synaptic transmission between the Schaffer collaterals and the pyramidal neurons. This impairment could be fended off in a dose-dependent manner by the intravenous administration of oxaloacetate in saline (at doses between 1.5 mmol and 0.1 μmol) immediately after the transient hypoperfusion. Our results suggest that oxaloacetate-mediated blood and brain glutamate scavenging contributes to the restoration of the LTP after its impairment by brain ischaemia.

Extreme longevity requires the continuous and large-scale adaptation of organ systems to delay senescence. Naked mole rats are the longest-living rodents, whose nervous system likely undergoes life-long adaptive reorganization. Nevertheless, neither the cellular organization of their cerebral cortex nor indices of structural neuronal plasticity along extreme time-scales have been established. We find that adult neurogenesis and neuronal migration are not unusual in naked mole rat brains. Instead, we show the prolonged expression of structural plasticity markers, many recognized as being developmentally controlled and multi-year-long postnatal neuromorphogenesis and spatial synapse refinement in hippocampal and olfactory structures of the naked mole rat brain. Neurophysiological studies on identified hippocampal neurons demonstrated that morphological differentiation is disconnected from the control of excitability in all neuronal contingents regardless of their ability to self-renew. Overall, we conclude that naked mole rats show an extremely protracted period of brain maturation that may permit plasticity and resilience to neurodegenerative processes over their decades-long life span. This conclusion is consistent with the hypothesis that naked mole rats are neotenous, with retention of juvenile characteristics to permit survival in a hypoxic environment, with extreme longevity a consequence of greatly retarded development.

Traditionally, neuroscientists have defined the identity of neurons by the cells' location, morphology, connectivity and excitability. However, the direct relationship between these parameters and the molecular phenotypes has remained largely unexplored. Here, we present a method for obtaining full transcriptome data from single neocortical pyramidal cells and interneurons after whole-cell patch-clamp recordings in mouse brain slices. In our approach, termed Patch-seq, a patch-clamp stimulus protocol is followed by the aspiration of the entire somatic compartment into the recording pipette, reverse transcription of RNA including addition of unique molecular identifiers, cDNA amplification, Illumina library preparation and sequencing. We show that Patch-seq reveals a close link between electrophysiological characteristics, responses to acute chemical challenges and RNA expression of neurotransmitter receptors and channels. Moreover, it distinguishes neuronal subpopulations that correspond to both well-established and, to our knowledge, hitherto undescribed neuronal subtypes. Our findings demonstrate the ability of Patch-seq to precisely map neuronal subtypes and predict their network contributions in the brain.

Local environmental cues are indispensable for axonal growth and guidance during brain circuit formation. Here, we combine genetic and pharmacological tools, as well as systems neuroanatomy in human fetuses and mouse models, to study the role of endocannabinoid and Slit/Robo signaling in axonal growth. We show that excess 2-arachidonoylglycerol, an endocannabinoid affecting directional axonal growth, triggers corpus callosum enlargement due to errant CB1 cannabinoid receptor (CB1R)-containing corticofugal axon spreading. This phenotype mechanistically relies on the premature differentiation and end-feet proliferation of CB2R-expressing oligodendrocytes. We further show the dependence of both axonal Robo1 positioning and oligodendroglial Slit2 production on cell-type specific cannabinoid receptor activation. Accordingly, Robo1 and/or Slit2 manipulation limits endocannabinoid modulation of axon guidance. We conclude that endocannabinoids can configure focal Slit2/Robo1 signaling to modulate directional axonal growth, which may provide a basis for understanding impaired brain wiring associated with metabolic deficits and prenatal drug exposure.

Children exposed in utero to cannabis present permanent neurobehavioral and cognitive impairments. Psychoactive constituents from Cannabis spp., particularly Δ(9)-tetrahydrocannabinol (THC), bind to cannabinoid receptors in the fetal brain. However, it is unknown whether THC can trigger a cannabinoid receptor-driven molecular cascade to disrupt neuronal specification. Here, we show that repeated THC exposure disrupts endocannabinoid signaling, particularly the temporal dynamics of CB1 cannabinoid receptor, to rewire the fetal cortical circuitry. By interrogating the THC-sensitive neuronal proteome we identify Superior Cervical Ganglion 10 (SCG10)/stathmin-2, a microtubule-binding protein in axons, as a substrate of altered neuronal connectivity. We find SCG10 mRNA and protein reduced in the hippocampus of midgestational human cannabis-exposed fetuses, defining SCG10 as the first cannabis-driven molecular effector in the developing cerebrum. CB1 cannabinoid receptor activation recruits c-Jun N-terminal kinases to phosphorylate SCG10, promoting its rapid degradation in situ in motile axons and microtubule stabilization. Thus, THC enables ectopic formation of filopodia and alters axon morphology. These data highlight the maintenance of cytoskeletal dynamics as a molecular target for cannabis, whose imbalance can limit the computational power of neuronal circuitries in affected offspring.

Brain ischaemia models are essential to study the pathomechanisms of stroke. Our aim was to investigate the reliability and reproducibility of our novel focal ischaemia-reperfusion model.To induce a cortical transient ischaemic attack, we lifted the distal middle cerebral artery (MCA) with a special hook. The early changes after 2 × 15-min occlusion were observed in the somatosensory evoked responses (SERs). The histological responses to 2 × 15-min MCA occlusion and to 30-, 45- or 60-min ischaemia were examined after a 1-day survival period by 2,3,5-triphenyltetrazolium chloride (TTC) and Fluoro Jade C (FJC) staining. Another group, with 30-min ischaemia, was analysed histologically by FJC, S100 and CD11b labelling after a 5-day survival period.The amplitudes of the SERs decreased immediately at the beginning of the ischaemic period, and remained at a reduced level during the ischaemia. Reperfusion resulted in increasing SER amplitudes, but they never regained the control level. The short-lasting ischaemia did not lead to brain infarction when evaluated with TTC, but intense labelling was found with FJC. The 30-min ischaemia did not result in FJC labelling after 1 day, but marked labelling was observed after 5 days with FJC, S100 and CD11b in the cortical area supplied by the MCA.We present here a novel, readily reproducible method to induce focal brain ischaemia. The ischaemia-reperfusion results in noteworthy changes in the SERs and the appearance of conventional tissue damage markers. This method involves possibilities for precise blood flow regulation, and the setting of the required level of perfusion.

Four-vessel occlusion (4VO), a frequently used model of global cerebral ischemia in rats, results in a dysfunction in wide brain areas, including the cerebral cortex and hippocampus. However, there are pronounced differences in response to global ischemia between the laboratory rat strains used in these studies. In the present work, the immediate acute effects of 4VO-induced global ischemia on the spontaneous electrocorticogram (ECoG) signals were analyzed in Wistar and Sprague–Dawley rats. The ECoG was isoelectric during the 10 min of global cerebral ischemia in Wistar rats and the first burst (FB) was seen 10–13 min after the start of reperfusion. In Sprague–Dawley rats, the FB was detected immediately after the start of 4VO or a few seconds later. The burst suppression ratio (BSR) in Wistar rats decreased to 45% in 5 min after FB, and after 25 min it was approximately 40%. In Sprague–Dawley rats, the BSR was 55% immediately after the FB and it decreased steeply to reach 0% by 10 min. There was also a significant difference between the two strains in the frequency composition of the ECoG pattern. The power spectral densities of the two strains differed virtually throughout the post-ischemic state. The histological results (Evans Blue, Cresyl Violet and Fluoro Jade C stainings) supplemented the electrophysiological data: the neuronal damage in the CA1 pyramids in Wistar rats was severe, whereas in the Sprague–Dawley animals it was only partial. These observations clearly demonstrate that the use of different rat strains (e.g. Wistar vs. Sprague–Dawley) can be a source of considerable variability in the results of acute experiments on global ischemia and it is important that the laboratory rats used in such experiments should be carefully chosen.

Postconditioning can be induced by a broad range of stimuli within minutes to days after an ischemic cerebral insult. A special form is elicited by pharmacological intervention called second pathophysiological stress. The present study aimed to evaluate the effects of low-dose (5 mg/kg) kainate postconditioning with onsets 0, 24 and 48 h after the ischemic insult on the hippocampal synaptic plasticity in a 2-vessel occlusion model in rat. The hippocampal function was tested by LTP measurements of Schaffer collateral-CA1 pyramidal cell synapses in acute slices and the changes in density of Golgi-Cox-stained apical dendritic spines. Postconditioning 0 and 24 h after ischemia was not protective, whereas 48-h-onset postconditioning resulted in the reappearance of a normal spine density (>100,000 spines) 3 days after ischemia, in parallel with the long-term restoration of the damaged LTP function. Similar, but somewhat less effects were observed after 10 days. Our data clearly demonstrate the onset dependence of postconditioning elicited by a subconvulsant dose of kainate treatment in global ischemia, with restoration of the structural plasticity and hippocampal function.

Gáspár Oláh,1 Judit Herédi,1 Ákos Menyhárt,1 Zsolt Czinege,2 Dávid Nagy,1 János Fuzik,1 Kitti Kocsis,1 Levente Knapp,1 Erika Krucsó,1 Levente Gellért,1 Zsolt Kis,1 Tamás Farkas,1 Ferenc Fülöp,3 Árpád Párdutz,4 János Tajti,4 László Vécsei,4 József Toldi1 1Department of Physiology, Anatomy and Neuroscience, 2Department of Software Engineering, 3Institute of Pharmaceutical Chemistry and MTA-SZTE Research Group for Stereochemistry, 4Department of Neurology and MTA-SZTE Neuroscience Research Group, University of Szeged, Szeged, Hungary Abstract: Cortical spreading depression (CSD) involves a slowly-propagating depolarization wave in the cortex, which can appear in numerous pathophysiological conditions, such as migraine with aura, stroke, and traumatic brain injury. Neurons and glial cells are also depolarized transiently during the phenomena. CSD is followed by a massive increase in glutamate release and by changes in the brain microcirculation. The aim of this study was to investigate the effects of two N-methyl-D-aspartate receptor antagonists, endogenous kynurenic acid (KYNA) and dizocilpine, on CSD and the related blood–brain barrier (BBB) permeability in rats. In intact animals, KYNA hardly crosses the BBB but has some positive features as compared with its precursor L-Kynurenine, which is frequently used in animal studies (KYNA cannot be metabolized to excitotoxic agents such as 3-hydroxy-L-kynurenine and quinolinic acid). We therefore investigated the possible effects of peripherally administered KYNA. Repetitive CSD waves were elicited by the application of 1 M KCl solution to the cortex. Direct current-electrocorticograms were measured for 1 hour. Four parameters of the waves were compared. Evans blue dye and fluorescent microscopy were used to study the possible changes in the permeability of the BBB. The results demonstrated that N-methyl-D-aspartate receptor antagonists can reduce the number of CSD waves and decrease the permeability of the BBB during CSD. These results suggest that KYNA itself or its derivatives may offer a new approach in the therapy of migraines. Keywords: blood–brain barrier, cortical spreading depression, glutamate receptors, kynurenic acid, kynurenines, NMDAR

The quantity of cells with paracrine effects for use in myocardial regeneration therapy is limited. This study investigated the effects of catheter-based endomyocardial delivery of secretome of 2.5 × 109 apoptotic peripheral blood mononuclear cells (APOSEC) on porcine chronic post-myocardial infarction (MI) left ventricular (LV) dysfunction and on gene expression. Closed-chest reperfused MI was induced in pigs by 90-min occlusion followed by reperfusion of the mid-LAD (day 0). At day 30, animals were randomized to receive porcine APOSEC (n = 8) or medium solution (control; n = 8) injected intramyocardially into the MI border zone using 3D NOGA guidance. At day 60, cardiac MRI with late enhancement and diagnostic NOGA (myocardial viability) were performed. Gene expression profiling of the infarct core, border zone, and normal myocardium was performed using microarray analysis and confirmed by quantitative real-time PCR. Injection of APOSEC significantly decreased infarct size (p