Larger Connection Radius Increases Hub Astrocyte Number in a 3D Neuron-Astrocyte Network Model

Astrocytes – a prominent glial cell type in the brain – form networks that tightly interact with the brain’s neuronal circuits. Thus, it is essential to study the modes of such interaction if we aim to understand how neural circuits process information. Thereby, calcium elevations, the primary signal in astrocytes, propagate to the adjacent neighboring cells and directly regulate neuronal communication. It is mostly unknown how the astrocyte network topology influences neuronal activity. Here, we used a computational model to simulate planar and 3D neuron-astrocyte networks with varying topologies. We investigated the number of active nodes, the shortest path, and the mean degree. Furthermore, we applied a graph coloring analysis that highlights the network organization between different network structures. With the increase of the maximum distance between two connected astrocytes, the information flow is more centralized to the most connected cells. Our results suggest that activity-dependent plasticity and the topology of brain areas might alter the amount of astrocyte controlled synapses

Astrocytic control in in vitro and simulated neuron-astrocyte networks

acceptedVersionPeer reviewe

Astrocytes in modulating subcellular, cellular and intercellular molecular neuronal communication

Astrocytes are one of the most abundant cell types in our brain. They modulate the brain homeostasis and play a role in the synaptic signalling and thus the molecular propagation inside the brain. Moreover, they form communication networks that co-localise with the neuronal networks with comparable topological complexity. There is an increasing piece of evidence that astrocytes are important in plasticity and learning from the level of the single synapse to the entire network. Moreover, several diseases are molecular communications on different scales from the synaptic to network level.acceptedVersionPeer reviewe

Astrocytes Exhibit a Protective Role in Neuronal Firing Patterns under Chemically Induced Seizures in Neuron-Astrocyte Co-Cultures.

Astrocytes and neurons respond to each other by releasing transmitters, such as γ-aminobutyric acid (GABA) and glutamate, that modulate the synaptic transmission and electrochemical behavior of both cell types. Astrocytes also maintain neuronal homeostasis by clearing neurotransmitters from the extracellular space. These astrocytic actions are altered in diseases involving malfunction of neurons, e.g., in epilepsy, Alzheimer's disease, and Parkinson's disease. Convulsant drugs such as 4-aminopyridine (4-AP) and gabazine are commonly used to study epilepsy in vitro. In this study, we aim to assess the modulatory roles of astrocytes during epileptic-like conditions and in compensating drug-elicited hyperactivity. We plated rat cortical neurons and astrocytes with different ratios on microelectrode arrays, induced seizures with 4-AP and gabazine, and recorded the evoked neuronal activity. Our results indicated that astrocytes effectively counteracted the effect of 4-AP during stimulation. Gabazine, instead, induced neuronal hyperactivity and synchronicity in all cultures. Furthermore, our results showed that the response time to the drugs increased with an increasing number of astrocytes in the co-cultures. To the best of our knowledge, our study is the first that shows the critical modulatory role of astrocytes in 4-AP and gabazine-induced discharges and highlights the importance of considering different proportions of cells in the cultures

Investigation of the input-output relationship of engineered neural networks using high-density microelectrode arrays

Bottom-up neuroscience utilizes small, engineered biological neural networks to study neuronal activity in systems of reduced complexity. We present a platform that establishes up to six independent networks formed by primary rat neurons on planar complementary metal–oxide–semiconductor (CMOS) microelectrode arrays (MEAs). We introduce an approach that allows repetitive stimulation and recording of network activity at any of the over 700 electrodes underlying a network. We demonstrate that the continuous application of a repetitive super-threshold stimulus yields a reproducible network answer within a 15 ms post-stimulus window. This response can be tracked with high spatiotemporal resolution across the whole extent of the network. Moreover, we show that the location of the stimulation plays a significant role in the networks' early response to the stimulus. By applying a stimulation pattern to all network-underlying electrodes in sequence, the sensitivity of the whole network to the stimulus can be visualized. We demonstrate that microchannels reduce the voltage stimulation threshold and induce the strongest network response. By varying the stimulation amplitude and frequency we reveal discrete network transition points. Finally, we introduce vector fields to follow stimulation-induced spike propagation pathways within the network. Overall we show that our defined neural networks on CMOS MEAs enable us to elicit highly reproducible activity patterns that can be precisely modulated by stimulation amplitude, stimulation frequency and the site of stimulation.Peer reviewe

Astrocyte regulation of neuronal activity in vitro and in silico : From computation to data analysis

Yli 86 miljoonaa synapsein yhdistettyä hermosolua muodostaa ihmisaivot jatkuvassa vuorovaikutuksessa muiden solutyyppien, kuten astrosyyttien kanssa. Astrosyytit ovat hermotukisoluja, jotka muovaavat aivojen solurakennetta, tarjoavat aineenvaihdunnallista tukea hermosoluille sekä edistävät synapsien muodostumista, kypsymistä ja poistoa välittäjäaineiden avulla. Täten astrosyytit vaikuttavat merkittävästi hermosolujen sähköiseen aktiivisuuteen. Astrosyyttien toimintahäiriöt voivat johtaa yli- aktiivisuuteen (hyperaktiivisuus) tai aliaktiivisuuteen (hypoaktiivisuus) ja siten myös hermostollisiin sairauksiin, kuten Alzheimerin tautiin ja epilepsiaan. Laskennallisia malleja hyödynnetään yhä enemmän tutkittaessa astrosyyttien vaikutuksia hermosolujen sähköiseen aktiivisuuteen. Tässä väitöskirjatyössä yhdistyvät laskennalliset menetelmät ja in vitro -laboratoriokokeet. Ensin hyödynsimme laskennallisia menetelmiä tutkiaksemme, miten astrosyyttien verkoston topologia vaikuttaa hermosolujen sähköiseen aktiivisuuteen ja sen ja sen synkronisuuteen hermosolu-astrosyyttiverkostossa. Seuraavaksi tutkimme in vitro laboratoriokokeilla, miten astrosyyttien ioni- ja välittäjäainesäätely ohjaa hermosolujen aktiivisuutta. Laajensimme jo aiemmin ryhmässämme kehitettyä laskennallista mallia tutkiaksemme: i) aukkoliitosten roolia hyperaktiivisuuden säätelyssä, ii) kaksi-ja kolmiulotteisen verkkotopologian vaikutusta hermosolujen aktiivisuuteen, iii) lasken- nallisten mallien ja laboratoriokokeiden vertailua, hyödyntäen erityisiä hermosolu- astrosyyttiviljelmiä mikroelektrodimatriisialustalla (MEA). Väitöskirjatyön toisessa osassa, jossa keskityttiin laboratoriotuloksiin, tutkimme astrosyyttien roolia lääkeaineiden neutraloimisessa, jotka aiheuttavat hermosoluissa konvulsiivisia vaikutuksia (lisääntynyttä sähköistä aktiivisuutta tai kouristuksia). Näitä lääkkeitä annettiin suoraan erilaisiin in vitro -viljelmiin. Tutkimme, miten astrosyytit ja niiden solunulkoinen ionien puhdistus vaikuttavat hermosolujen aktiopotentiaalien muotoon ja aktiivisuuden kaavoittumiseen kehityksen aikana. Laskennalliset tulokset osoittavat, että sekä astrosyyttien että hermosolujen verkoston rakenteella on merkittävä rooli hermosolujen toimintapotentiaalin leviämisisessä hermoverkostossa. Itse asiassa tämä rakenne vaikuttaa siihen, kuinka astrosyytit jakelevat välittäjäaineita, mikä puolestaan vaikuttaa niiden lähialueen hermosolujen aktiivisuuden lisääntymiseen tai vähenemiseen. Lisäksi laboratoriokokeet osoittavat, että hermosolujen sähköinen vaste estettäessä jänniteherkkien kaliumkanavien toimintaa 4-aminopyridiinillä, tai gamma-aminovoihappo tyypin A reseptoreja gabatsiinilla, riippuu astrosyyttien määrästä viljelmissä. Tämä viittaa siihen, että astrosyyttien lukumäärä hermoverkostossa on olennaisen tärkeä ioni- ja välittäjäainetasapanon ylläpitämiselle. Väitöskirjan yhteenvetona voidaan todeta, että astrosyytit todennäköisesti säätelevät hermosolujen toimintaa poistamalla ioneja ja välittäjäaineita solunulkoisesta tilasta, mikä puolestaan säätelee hermosolujen aktiopotentiaaleja ja niiden etenemistä hermoverkostossa. Astrosyytit aiheuttavat signaalien eriaikaistumista, mikä voi viitata in vivo ihmis- ja eläinkokeissa vähentyneeseen alttiuteen epileptisille kohtauksille. Tämä väitöskirjatyö esittelee ensimmäiset tutkimukset, jossa on käytetty tarkasti määriteltyjä viljelmiä eri hermosolujen ja astrosyyttien määrällisillä suhteilla. Kaiken kaikkiaan tämä työ korostaa hermosolujen ja astrosyyttien suhteellisten osuuksien huomioimisen tärkeyttä uusia lääkeaineita arvioitaessa ja yleisesti yhteisviljelmiä hyödyntävissä laboratoriokokeissa.The human brain is composed of more than 86 billion neurons, interconnected in synapses and in constant communication with other cell types, like for example the astrocytes. Astrocytes shape the cytoarchitecture of the brain and provide metabolic support to the neurons. Furthermore, they promote synapse formation, maturation, and elimination by exchanging transmitters with them. Thus, astrocytes can modulate neuronal activity. Impairments in astrocytes may lead to hyper- or hypoactivity and thus to diseases like Alzheimer’s and epilepsy. Computational models became more commonly used to better understand unanswered questions on the astrocytic modulation of neuronal activity. This thesis combines computational studies and in vitro experiments. We first utilized computational methods to study how astrocyte network topology affects neuronal spiking and synchronization in neuron-astrocyte networks. Then, we examined in vitro how astrocytic ionic and gliotransmitter regulation controls neuronal activity. In the computational studies, we extended a computational model previously developed in our group to: i) test the role of gap junction uncoupling in hyperactivity control, ii) compare 2D and 3D co-culture network topology effects on neuronal activity and iii) compare the results obtained from the model with experimental results obtained using microelectrode arrays recording of neuron-astrocyte co-cultures with fixed relative amounts of neurons and astrocytes. In the second part of the thesis, which is focused on the in vitro experiments conducted, we explored the role of astrocytes in counteracting the effect of convulsant drugs applied to the different in vitro cultures. Furthermore, we studied how astrocytes and extracellular ionic clearance affect the spike shape and the activity patterning during development. The results of the computational studies suggest that the topology of both the astrocyte and neuron networks is crucial for the activity propagation in the neurons. The topology, in fact, affects how the transmitters are distributed in the astrocytes, resulting in an up- or down-regulation of the activity in the neighboring neurons. The experimental studies, moreover, show that blocking the activity of either voltage-gated K+ channels with 4AP or of GABAA receptors with gabazine resulted in effects that depended on the particular relative proportions of neuron and astrocytes in the co-culture. This suggested that the amount of astrocytes in the net- works is fundamental for proper control of alterations of the ionic and transmitter homeostasis. The collective findings of this thesis indicate that indeed astrocytes have a regulatory effect on neuronal activity, probably by eliminating ions and gliotransmitters from the extracellular space, thereby controlling the neuronal action potentials, signal propagation within the network, and synchronicity and patterning of the signals. Astrocytes induced desynchronization in the signals and led to less patterned signals; in vivo this effect might be reflected in a reduced seizure susceptibility. This thesis presents the first studies where the cultures had defined relative neuron-astrocytes proportions. Overall, our studies stress the importance of considering the ratio of astrocytes in the cultures when testing new drugs or in general in co-culture experiments

Investigation of the input-output relationship of engineered neural networks using high-density microelectrode arrays

Bottom-up neuroscience utilizes small, engineered biological neural networks to study neuronal activity in systems of reduced complexity. We present a platform that establishes up to six independent networks formed by primary rat neurons on planar complementary metal–oxide–semiconductor (CMOS) microelectrode arrays (MEAs). We introduce an approach that allows repetitive stimulation and recording of network activity at any of the over 700 electrodes underlying a network. We demonstrate that the continuous application of a repetitive super-threshold stimulus yields a reproducible network answer within a 15 ms post-stimulus window. This response can be tracked with high spatiotemporal resolution across the whole extent of the network. Moreover, we show that the location of the stimulation plays a significant role in the networks' early response to the stimulus. By applying a stimulation pattern to all network-underlying electrodes in sequence, the sensitivity of the whole network to the stimulus can be visualized. We demonstrate that microchannels reduce the voltage stimulation threshold and induce the strongest network response. By varying the stimulation amplitude and frequency we reveal discrete network transition points. Finally, we introduce vector fields to follow stimulation-induced spike propagation pathways within the network. Overall we show that our defined neural networks on CMOS MEAs enable us to elicit highly reproducible activity patterns that can be precisely modulated by stimulation amplitude, stimulation frequency and the site of stimulation.ISSN:0956-5663ISSN:1873-423