A neokortikális és hippocampális epilepsziák komplex elektrofiziológiai rétegelvezetéses és szövettani vizsgálata emberben = Complex laminar electrophysiological and histological examination of the human neocortical and hippocampal epilepsies

Az OITI és a MTA PKI együttműködésében részletesen kidolgoztuk az intraoperatív és krónikus multielektróda beültetés technikai feltételeit és a vizsgálatok forgatókönyvét. Létrehoztunk egy kombinált multielektródás és konvencionális klinikai grid-sztrip elektródos elvezető rendszert, mely segítségével az operáció alatt, illetve krónikusan tudunk elvezetni intrahippokampális, intrakortikális, valamint szubdurális potenciálokat. Kimutattuk a szubikulumban generált különféle epilepsziás kisülések és a laterális temporális kéreg aktivitásának kapcsolatait, kimutattuk továbbá az alvásban és epilepsziában fontos K-komplexum generátorainak kérgi eredetét. Vizsgáltuk az alvási oszcilláció és az epilepsziás események kapcsolatát, kimutattuk, hogy epilepsziás emberben a felszínhez közeli kérgi rétegek igen erős szinaptikus és tüzelési aktivitást mutatnak az alvási oszcilláció aktív fázisában. Kimutattuk továbbá, hogy az aktív fázis csoportosítja az epilepsziás kisüléseket. A kérgi elektromos ingerlés hatását vizsgálva epilepsziás betegeken megállapítottuk, hogy a rövid elektromos ingerek inaktiválják a kérget, amit későbbiekben terápiás céllal lehet hasznosítani. | In collaboration with the OITI and MTA PKI we have worked out in details the intraoperative and chronic multielectrode implantation technique and the schedule of the investigation. We have established a combined system composed of the conventional clinical grid and strip based electrophysiology apparatus and the novel investigational multielectrode system to measure intrahippocampal, intracortical and subdural potentials chronically and intraoperatively. We have shown the relationship of the subicular and lateral temporal lobe epileptic discharges, we have also shown the cortical origin of the K-complex, an important brain wave in sleep and epilepsy. We have investigated the slow sleep oscillation and its relationship to epilepsy. The slow waves were originated in the superficial layers of the cortex and the epileptiform discharges were grouped by the up-states of the slow oscillation. Investigating the effect of electrical stimulation, we have shown that brief current pulses can inactivate the cortex, which effect can also be exploited in the therapy of epilepsy

Critical role of somatostatin receptor 2 in the vulnerability of the central noradrenergic system: new aspects on Alzheimer's disease

Alzheimer's disease and other age-related neurodegenerative disorders are associated with deterioration of the noradrenergic locus coeruleus (LC), a probable trigger for mood and memory dysfunction. LC noradrenergic neurons exhibit particularly high levels of somatostatin binding sites. This is noteworthy since cortical and hypothalamic somatostatin content is reduced in neurodegenerative pathologies. Yet a possible role of a somatostatin signal deficit in the maintenance of noradrenergic projections remains unknown. Here, we deployed tissue microarrays, immunohistochemistry, quantitative morphometry and mRNA profiling in a cohort of Alzheimer's and age-matched control brains in combination with genetic models of somatostatin receptor deficiency to establish causality between defunct somatostatin signalling and noradrenergic neurodegeneration. In Alzheimer's disease, we found significantly reduced somatostatin protein expression in the temporal cortex, with aberrant clustering and bulging of tyrosine hydroxylase-immunoreactive afferents. As such, somatostatin receptor 2 (SSTR2) mRNA was highly expressed in the human LC, with its levels significantly decreasing from Braak stages III/IV and onwards, i.e., a process preceding advanced Alzheimer's pathology. The loss of SSTR2 transcripts in the LC neurons appeared selective, since tyrosine hydroxylase, dopamine beta-hydroxylase, galanin or galanin receptor 3 mRNAs remained unchanged. We modeled these pathogenic changes in Sstr2 -/- mice and, unlike in Sstr1 -/- or Sstr4 -/- genotypes, they showed selective, global and progressive degeneration of their central noradrenergic projections. However, neuronal perikarya in the LC were found intact until late adulthood (<8 months) in Sstr2 -/- mice. In contrast, the noradrenergic neurons in the superior cervical ganglion lacked SSTR2 and, as expected, the sympathetic innervation of the head region did not show any signs of degeneration. Our results indicate that SSTR2-mediated signaling is integral to the maintenance of central noradrenergic projections at the system level, and that early loss of somatostatin receptor 2 function may be associated with the selective vulnerability of the noradrenergic system in Alzheimer's disease

Interhemispheric differences of pyramidal cells in the primary motor cortices of schizophrenia patients investigated postmortem

Motor disturbances are observed in schizophrenia patients, but the neuroanatomical background is unknown. Our aim was to investigate the pyramidal cells of the primary motor cortex (BA 4) in both hemispheres of postmortem control and schizophrenia subjects-8 subjects in each group-with 2.5-5.5 h postmortem interval. The density and size of the Sternberger monoclonal incorporated antibody 32 (SMI32)-immunostained pyramidal cells in layer 3 and 5 showed no change; however, the proportion of larger pyramidal cells is decreased in layer 5. Giant pyramidal neurons (Betz cells) were investigated distinctively with SMI32- and parvalbumin (PV) immunostainings. In the right hemisphere of schizophrenia subjects, the density of Betz cells was decreased and their PV-immunopositive perisomatic input showed impairment. Part of the Betz cells contained PV in both groups, but the proportion of PV-positive cells has declined with age. The rat model of antipsychotic treatment with haloperidol and olanzapine showed no differences in size and density of SMI32-immunopositive pyramidal cells. Our results suggest that motor impairment of schizophrenia patients may have a morphological basis involving the Betz cells in the right hemisphere. These alterations can have neurodevelopmental and neurodegenerative explanations, but antipsychotic treatment does not explain them

Spatiotemporal characterization of cellular tau pathology in the human locus coeruleus–pericoerulear complex by three-dimensional imaging

Tau pathology of the noradrenergic locus coeruleus (LC) is a hallmark of several age-related neurodegenerative disorders, including Alzheimer’s disease. However, a comprehensive neuropathological examination of the LC is difficult due to its small size and rod-like shape. To investigate the LC cytoarchitecture and tau cytoskeletal pathology in relation to possible propagation patterns of disease-associated tau in an unprecedented large-scale three-dimensional view, we utilized volume immunostaining and optical clearing technology combined with light sheet fluorescence microscopy. We examined AT8 + pathological tau in the LC/pericoerulear region of 20 brains from Braak neurofibrillary tangle (NFT) stage 0–6. We demonstrate an intriguing morphological complexity and heterogeneity of AT8 + cellular structures in the LC, representing various intracellular stages of NFT maturation and their diverse transition forms. We describe novel morphologies of neuronal tau pathology such as AT8 + cells with fine filamentous somatic protrusions or with disintegrating soma. We show that gradual dendritic atrophy is the first morphological sign of the degeneration of tangle-bearing neurons, even preceding axonal lesions. Interestingly, irrespective of the Braak NFT stage, tau pathology is more advanced in the dorsal LC that preferentially projects to vulnerable forebrain regions in Alzheimer’s disease, like the hippocampus or neocortical areas, compared to the ventral LC projecting to the cerebellum and medulla. Moreover, already in the precortical Braak 0 stage, 3D analysis reveals clustering tendency and dendro-dendritic close appositions of AT8 + LC neurons, AT8 + long axons of NFT-bearing cells that join the ascending dorsal noradrenergic bundle after leaving the LC, as well as AT8 + processes of NFT-bearing LC neurons that target the 4th ventricle wall. Our study suggests that the unique cytoarchitecture, comprised of a densely packed and dendritically extensively interconnected neuronal network with long projections, makes the human LC to be an ideal anatomical template for early accumulation and trans-neuronal spreading of hyperphosphorylated tau

Az endocannabinoid-mediált szignalizáció funkciója a hippocampus neuronhálózatainak normális és kóros működéseiben = The role of endocannabinoid-mediated signaling in normal and pathological operations os hippocampal circuits

Az OTKA pályázat támogatásával az elmúlt négy évben alapvető felismeréseket tettünk egy új kémiai szignálrendszer, az endokannabinoid rendszer molekuláris és anatómiai szerveződéséről, élettani és kórélettani jelentőségéről. Kimutattuk, hogy az endokannabinoid rendszer egy specializált jelátviteli rendszer, amelynek feladata, hogy a szinapszisok működését a preszinaptikus és a posztszinaptikus idegsejt aktivitásának függvényében szabályozza. Elsőként írtuk le egy endokannabinoid molekula, a 2-arachidonilglicerol kulcsszerepét ebben a folyamatban, és feltártuk, hogy milyen molekuláris mechanizmusok szabályozzák keletkezését és lebontását. Igazoltuk, hogy ez a kémiai szignálrendszer számos agyterületen (agykéregben, hippocampusban, nucleus accumbensben, a ventrális tegmentális areában) megtalálható. Élettani kísérleteink alapján az endokannabinoidok egy negatív visszacsatolási folyamat közvetítői a szinapszisokban. Ezzel párhuzamosan felfedeztük, hogy az endokannabinoid rendszer működési zavarai kulcsszerepet játszhatnak a temporális lebeny eredetű epilepsziában, valamint a szorongásos és poszttraumatikus stressz-okozta panaszok hátterében is megtalálhatók. Ezek az eredményeink egyben új molekuláris gyógyszercélpontokat jelölnek ki, amelyek számos idegrendszeri megbetegedésben vezethetnek hatékonyabb és szelektívebb farmakoterápia kidolgozásához. | During the last four years, our research group has reached fundamental milestones in the understanding of a new chemical messenger system, the so-called endocannabinoid system. We have uncovered the molecular and anatomical organization of endocannabinoid signaling and provided clues for its physiological and pathophysiological importance. We have shown that the endocannabinoid system is a specialized signaling machinery, which controls the efficacy synaptic transmission as a function of the activity of presynaptic and postsynaptic neurons. We have described for the first time that 2-arachidonoylglycerol is a key player in this process, and uncovered the basic mechanisms in its biosynthesis and degradation. We have provided evidence that this chemical signaling mechanism is a conserved feature of several types of synapses in various brain regions, for example in the neocortex, hippocampus, nucleus accumbens and the ventral tegmental area. We have shown that this system is involved in a negative feed-back regulation of transmitter release, and described its impairment in the human epileptic hippocampus, as well as its contribution to anxiety and post-traumatic stress disorder in animal models. These findings unravel new drug targets, whereby they could open novel therapeutic approaches for a more efficient and selective treatment of several brain disorders

Enhanced expression of potassium-chloride cotransporter KCC2 in human temporal lobe epilepsy

Synaptic reorganization in the epileptic hippocampus involves altered excitatory and inhibitory transmission besides the rearrangement of dendritic spines, resulting in altered excitability, ion homeostasis, and cell swelling. The potassium-chloride cotransporter-2 (KCC2) is the main chloride extruder in neurons and hence will play a prominent role in determining the polarity of GABAA receptor-mediated chloride currents. In addition, KCC2 also interacts with the actin cytoskeleton which is critical for dendritic spine morphogenesis, and for the maintenance of glutamatergic synapses and cell volume. Using immunocytochemistry, we examined the cellular and subcellular levels of KCC2 in surgically removed hippocampi of temporal lobe epilepsy (TLE) patients and compared them to control human tissue. We also studied the distribution of KCC2 in a pilocarpine mouse model of epilepsy. An overall increase in KCC2-expression was found in epilepsy and confirmed by Western blots. The cellular and subcellular distributions in control mouse and human samples were largely similar; moreover, changes affecting KCC2-expression were also alike in chronic epileptic human and mouse hippocampi. At the subcellular level, we determined the neuronal elements exhibiting enhanced KCC2 expression. In epileptic tissue, staining became more intense in the immunopositive elements detected in control tissue, and profiles with subthreshold expression of KCC2 in control samples became labelled. Positive interneuron somata and dendrites were more numerous in epileptic hippocampi, despite severe interneuron loss. Whether the elevation of KCC2-expression is ultimately a pro- or anticonvulsive change, or both-behaving differently during ictal and interictal states in a context-dependent manner-remains to be established

The neuropeptide landscape of human prefrontal cortex

Human prefrontal cortex (hPFC) is a complex brain region involved in cognitive and emotional processes and several psychiatric disorders. Here, we present an overview of the distribution of the peptidergic systems in 17 subregions of hPFC and three reference cortices obtained by microdissection and based on RNA sequencing and RNAscope methods integrated with published single-cell transcriptomics data. We detected expression of 60 neuropeptides and 60 neuropeptide receptors in at least one of the hPFC subregions. The results reveal that the peptidergic landscape in PFC consists of closely located and functionally different subregions with unique peptide/transmitter–related profiles. Neuropeptide-rich PFC subregions were identified, encompassing regions from anterior cingulate cortex/orbitofrontal gyrus. Furthermore, marked differences in gene expression exist between different PFC regions (>5-fold; cocaine and amphetamine–regulated transcript peptide) as well as between PFC regions and reference regions, for example, for somatostatin and several receptors. We suggest that the present approach allows definition of, still hypothetical, microcircuits exemplified by glutamatergic neurons expressing a peptide cotransmitter either as an agonist (hypocretin/orexin) or antagonist (galanin). Specific neuropeptide receptors have been identified as possible targets for neuronal afferents and, interestingly, peripheral blood-borne peptide hormones (leptin, adiponectin, gastric inhibitory peptide, glucagon-like peptides, and peptide YY). Together with other recent publications, our results support the view that neuropeptide systems may play an important role in hPFC and underpin the concept that neuropeptide signaling helps stabilize circuit connectivity and fine-tune/modulate PFC functions executed during health and disease

A conserved MTMR lipid phosphatase increasingly suppresses autophagy in brain neurons during aging

Ageing is driven by the progressive, lifelong accumulation of cellular damage. Autophagy (cellular self-eating) functions as a major cell clearance mechanism to degrade such damages, and its capacity declines with age. Despite its physiological and medical significance, it remains largely unknown why autophagy becomes incapable of effectively eliminating harmful cellular materials in many cells at advanced ages. Here we show that age-associated defects in autophagic degradation occur at both the early and late stages of the process. Furthermore, in the fruit fly Drosophila melanogaster , the myotubularin-related (MTMR) lipid phosphatase egg-derived tyrosine phosphatase (EDTP) known as an autophagy repressor gradually accumulates in brain neurons during the adult lifespan. The age-related increase in EDTP activity is associated with a growing DNA N 6 -adenine methylation at EDTP locus. MTMR14, the human counterpart of EDTP, also tends to accumulate with age in brain neurons. Thus, EDTP, and presumably MTMR14, promotes brain ageing by increasingly suppressing autophagy throughout adulthood. We propose that EDTP and MTMR14 phosphatases operate as endogenous pro-ageing factors setting the rate at which neurons age largely independently of environmental factors, and that autophagy is influenced by DNA N 6 -methyladenine levels in insects

HCN4 subunit expression in fast-spiking interneurons of the rat spinal cord and hippocampus

Hyperpolarisation-activated (Ih) currents are considered important for dendritic integration, synaptic transmission, setting membrane potential and rhythmic action potential (AP) discharge in neurons of the central nervous system. Hyperpolarisation-activated cyclic nucleotide-gated (HCN) channels underlie these currents and are composed of homo- and hetero-tetramers of HCN channel subunits (HCN1–4), which confer distinct biophysical properties on the channel. Despite understanding the structure–function relationships of HCN channels with different subunit stoichiometry, our knowledge of their expression in defined neuronal populations remains limited. Recently, we have shownthat HCN subunit expression is a feature of a specific population of dorsal horn interneurons that exhibit high-frequency AP discharge. Here we expand on this observation and use neuroanatomical markers to first identify well-characterised neuronal populations in the lumbar spinal cord and hippocampus and subsequently determine whether HCN4 expression correlates with high-frequency AP discharge in these populations. In the spinal cord, HCN4 is expressed in several putative inhibitory interneuron populations including parvalbumin (PV)-expressing islet cells (84.1%; SD: ±2.87), in addition to all putative Renshaw cells and Ia inhibitory interneurons. Similarly, virtually all PVexpressing cells in the hippocampal CA1 subfield (93.5%;±3.40) and the dentate gyrus (90.9%; ±6.38) also express HCN4. This HCN4 expression profile in inhibitory interneurons mirrors both the prevalence of Ih sub-threshold currents and high-frequency AP discharge. Our findings indicate that HCN4 subunits are expressed in several populations of spinal and hippocampal interneurons, which are known to express both Ih sub-threshold currents and exhibit high-frequency AP discharge. As HCN channel function plays a critical role in pain perception, learning and memory,and sleep as well as the pathogenesis of several neurologicaldiseases, these findings provide important insights into the identity and neurochemical status of cells that could underlie such conditions