Synaptic mechanisms of pattern completion in the hippocampal CA3 network

The hippocampal CA3 region plays a key role in learning and memory. Recurrent CA3–CA3 synapses are thought to be the subcellular substrate of pattern completion. However, the synaptic mechanisms of this network computation remain enigmatic. To investigate these mechanisms, we combined functional connectivity analysis with network modeling. Simultaneous recording fromup to eight CA3 pyramidal neurons revealed that connectivity was sparse, spatially uniform, and highly enriched in disynaptic motifs (reciprocal, convergence,divergence, and chain motifs). Unitary connections were composed of one or two synaptic contacts, suggesting efficient use of postsynaptic space. Real-size modeling indicated that CA3 networks with sparse connectivity, disynaptic motifs, and single-contact connections robustly generated pattern completion.Thus, macro- and microconnectivity contribute to efficient memory storage and retrieval in hippocampal networks

Intrinsic membrane properties determine hippocampal differential firing pattern in vivo in anesthetized rats

The hippocampus plays a key role in learning and memory. Previous studies suggested that the main types of principal neurons, dentate gyrus granule cells (GCs), CA3 pyramidal neurons, and CA1 pyramidal neurons, differ in their activity pattern, with sparse firing in GCs and more frequent firing in CA3 and CA1 pyramidal neurons. It has been assumed but never shown that such different activity may be caused by differential synaptic excitation. To test this hypothesis, we performed high-resolution whole-cell patch-clamp recordings in anesthetized rats in vivo. In contrast to previous in vitro data, both CA3 and CA1 pyramidal neurons fired action potentials spontaneously, with a frequency of ∼3–6 Hz, whereas GCs were silent. Furthermore, both CA3 and CA1 cells primarily fired in bursts. To determine the underlying mechanisms, we quantitatively assessed the frequency of spontaneous excitatory synaptic input, the passive membrane properties, and the active membrane characteristics. Surprisingly, GCs showed comparable synaptic excitation to CA3 and CA1 cells and the highest ratio of excitation versus hyperpolarizing inhibition. Thus, differential synaptic excitation is not responsible for differences in firing. Moreover, the three types of hippocampal neurons markedly differed in their passive properties. While GCs showed the most negative membrane potential, CA3 pyramidal neurons had the highest input resistance and the slowest membrane time constant. The three types of neurons also differed in the active membrane characteristics. GCs showed the highest action potential threshold, but displayed the largest gain of the input-output curves. In conclusion, our results reveal that differential firing of the three main types of hippocampal principal neurons in vivo is not primarily caused by differences in the characteristics of the synaptic input, but by the distinct properties of synaptic integration and input-output transformation

Investigation of the hippocampal information processing in freely moving rats

Extracellular electrophysiological recordings of the hippocampus were carried out in freely moving Long-Evans rats. Large-scale and high-density electrodes were used to target all subregions simultaneously. We were able to record both local field potentials (LFPs) and single unit activity, which allowed for the analysis of population activity and individual cells, respectively. We recorded the animals while they performed spatial navigational tasks and during sleep in their home cages. We focused our investigations on the information processing of the hippocampus during its two main general functions: spatial navigation and memory consolidation. In particular, two main questions were addressed in this work:How the different subregions of the hippocampus contribute to spatial coding within the hippocampus. How the distinct subregions of the hippocampus coordinate to generate sharp-wave ripples complexes, known to be essential in memory consolidation To investigate the contribution of the different subregions of the hippocampus to spatial coding, we recorded simultaneously place cells (neurons which selectively respond to crossing locations in the environment) from CA1, CA2 and CA3 during spatial navigation.We analized the fine function of the distinct anatomical portions of these areas separating CA1 into proximal, intermediate and distal and CA3 into CA3c, CA3b and CA3a subregions.We could also record neurons located at several depths of the soma layers so we treated the deep (toward stratum oriens) and the superficially (toward stratum radiatum) located cells inthe CA1, CA2 and CA3 regions separately. We analyzed the distinct properties of the place cells located in the different subregions and found significant differences which characterize the spatial coding in these parts and correlate with the anatomy. The CA1 and CA2 regions in general had a higher number of place cells than the CA3region. Furthermore, more fields per cell were found in CA1 neurons and CA2 compared to their CA3 peers. Firing rates inside the fields and peak firing rates were also higher for CA1 and CA2. The more spatially informative place cells were the ones located in CA3, and the less informative the CA2 ones. Inside the different subregions, CA1 proximal cells appeared 43 to be more spatially informative, showed higher firing rates in-field and a tended to have one single place field whereas toward CA1 distal they fired less, had more place fields and contained less spatial information. For the CA3 region, more spatially informative and selective place cells were located in CA3c, with preferentially single fields but lower firing rates than in their CA3a peers. No significant differences were found within the cells located at different depths in the CA3 area, whereas in CA1 and CA2 a tendency characterized the deep cells with more number of place cells, higher in-field firing rates but less selective and spatially informative than their superficial peers. These findings are in correlation with the axonal distribution of the different afferents to the hippocampus, mainly from the entorhinal cortex. The medial part of the entorhinal cortex (where the highly spatially selective grid cells are located) is preferentially connected with the proximal CA1 (more spatial selectivity than the distal part) and the lateral portion of the entorhinal cortex (where mainly object and environmental cues coding cells are located) mainly target the distal part of CA1 (more place fields per cell and less spatially informative). Unlike the CA1, cells in the CA3 region have strong recurrent collaterals, which can account for the lower number and highly spatially selective place cells in this region. The least informative cells during spatial navigation were the ones located in CA2, which is in line with the discovery of a specific network which code for space during immobility inside this region. The phase precession was weaker in place cells from the CA2 region, which can be also explained by the distinct combination of inputs arriving to this part of the hippocampus. The theta phase locking of the different subregions also correlates with anatomical inputs. While a relatively preserved phase preference was found in CA1 cells from all subregions during RUN (ascending phase of the cycle), a gradual shift was notable from the CA2-CA3a border (ascending phase) toward the CA3c (descending phase). During REM, an important percentage of cells shift their firing with 180º in the CA1 region, but this is not the case for the cells located in CA2 or CA3 regions. Therefore, this is assumed to be related to the specific entorhinal layer III input to the CA1 region..

Dopamine D3 Receptors Inhibit Hippocampal Gamma Oscillations by Disturbing CA3 Pyramidal Cell Firing Synchrony

Cortical gamma oscillations are associated with cognitive processes and are altered in several neuropsychiatric conditions such as schizophrenia and Alzheimer’s disease. Since dopamine D3 receptors are possible targets in treatment of these conditions, it is of great importance to understand their role in modulation of gamma oscillations. The effect of D3 receptors on gamma oscillations and the underlying cellular mechanisms were investigated by extracellular local field potential and simultaneous intracellular sharp micro-electrode recordings in the CA3 region of the hippocampus in vitro. D3 receptors decreased the power and broadened the bandwidth of gamma oscillations induced by acetylcholine or kainate. Blockade of the D3 receptors resulted in faster synchronization of the oscillations, suggesting that endogenous dopamine in the hippocampus slows down the dynamics of gamma oscillations by activation of D3 receptors. Investigating the underlying cellular mechanisms for these effects showed that D3 receptor activation decreased the rate of action potentials (APs) during gamma oscillations and reduced the precision of the AP phase coupling to the gamma cycle in CA3 pyramidal cells. The results may offer an explanation how selective activation of D3 receptors may impair cognition and how, in converse, D3 antagonists may exert pro-cognitive and antipsychotic effects

Histopathological modeling of status epilepticus-induced brain damage based on in vivo diffusion tensor imaging in rats

Non-invasive magnetic resonance imaging (MRI) methods have proved useful in the diagnosis and prognosis of neurodegenerative diseases. However, the interpretation of imaging outcomes in terms of tissue pathology is still challenging. This study goes beyond the current interpretation of in vivo diffusion tensor imaging (DTI) by constructing multivariate models of quantitative tissue microstructure in status epilepticus (SE)-induced brain damage. We performed in vivo DTI and histology in rats at 79 days after SE and control animals. The analyses focused on the corpus callosum, hippocampal subfield CA3b, and layers V and VI of the parietal cortex. Comparison between control and SE rats indicated that a combination of microstructural tissue changes occurring after SE, such as cellularity, organization of myelinated axons, and/or morphology of astrocytes, affect DTI parameters. Subsequently, we constructed a multivariate regression model for explaining and predicting histological parameters based on DTI. The model revealed that DTI predicted well the organization of myelinated axons (cross-validated R = 0.876) and astrocyte processes (cross-validated R = 0.909) and possessed a predictive value for cell density (CD) (cross-validated R = 0.489). However, the morphology of astrocytes (cross-validated R > 0.05) was not well predicted. The inclusion of parameters from CA3b was necessary for modeling histopathology. Moreover, the multivariate DTI model explained better histological parameters than any univariate model. In conclusion, we demonstrate that combining several analytical and statistical tools can help interpret imaging outcomes to microstructural tissue changes, opening new avenues to improve the non-invasive diagnosis and prognosis of brain tissue damage

Development of Epilepsy-on-a-chip System for High-throughput Antiepileptogenic Drug Discovery

Epilepsy is one of the most common neurological disorders and affects millions of people in the United States. Currently available antiepileptic drugs require continuous administration for suppression of seizures and have not been shown to prevent the development of epilepsy (epileptogenesis). The discovery of antiepileptogenic drug is complicated by the long time course of epileptogenesis in animal models of epilepsy and the requirement of continuous monitoring of epileptiform activity in vivo for the assessment of drug efficacy. In recent years, organotypic hippocampal cultures have been increasingly used as an in vitro model of post-traumatic epilepsy in both basic and translational research. Epileptogenesis in this in vitro model has a compressed time scale and can be monitored by detection of electrographic and biochemical markers of seizure-like activity. However, the lack of a scalable chronic electrical recording platform is a significant bottleneck in high-throughput antiepileptogenic drug discovery using organotypic cultures.In an effort to circumvent the throughput limitations of in vitro antiepileptogenic drug discovery, a hybrid microfluidic-multiple electrode array (µflow-MEA) technology was developed for scalable chronic electrical assay of epileptogenesis in vitro. Specifically, the microfluidic perfusion technique was utilized to miniature the culture platform, which enabled the long-term maintenance of an organotypic culture array on a single device. The integration of the microfluidic perfusion system with a customized planar MEA allowed for parallel continuous recordings. As a proof-of-concept demonstration, a pilot screen of receptor tyrosine kinase (RTK) inhibitor library was performed on µflow-MEA based electrical assay platform. The screen results revealed significant antiepileptogenic effect of cFMS RTK inhibitor.This thesis also provides further validation of the organotypic hippocampal culture model of epilepsy by investigating the influence of culture medium composition on epileptogenesis. We found that epileptogenesis occurred in any culture medium that was capable of supporting neural survival, indicating that culture medium composition has limited influence on epileptogenesis in organotypic hippocampal cultures.It is hoped that the techniques presented in this thesis will accelerate the antiepileptogenic drug discovery and contribute to the development of new therapeutics to treat individuals at risk of epileptogenesis

Experience-dependent structural rearrangements of synaptic connectivity in the adult central nervous system

The functioning of the brain critically relies on its capacity to adapt and respond to its environment. The brain’s ability to change in response to experience is called plasticity and underlies principal brain functions, such as learning and memory. My thesis work investigated the ability of the brain to structurally remodel upon altered experiences, and changes that occur during normal aging. Furthermore, I addressed what might be the molecular mechanisms regulating such remodeling. I will therefore start by introducing the term of experience-dependent plasticity and exemplify the brain’s capacity to adapt to changes in experience and usage. I will then attempt to describe mechanisms of experience-dependent plasticity on the functional, molecular and structural level. Furthermore, I will discuss the impact of age and life-style on the brain’s capacity for plasticity. Finally, I will close the introduction by outlining the function and anatomy of the brain region that was the main subject of our investigations, namely the hippocampus, and specifically the mossy fiber pathwa