Detrimental effect of post Status Epilepticus treatment with ROCK inhibitor Y-27632 in a pilocarpine model of temporal lobe epilepsy

Temporal lobe epilepsy (TLE) is the most common type of epilepsy in adults where 20-30% of the patients are refractory to currently available anti-epileptic drugs. The RhoA/Rho-kinase signaling pathway activation has been involved in inflammatory responses, neurite outgrowth and neuronal death under pathological conditions such as epileptic insults. Acute preventive administration of ROCK inhibitor has been reported to have beneficial outcomes in Status Epileptic us (SE) epilepsy. In the present study, we evaluate the effect of chronic post SE treatment with the ROCK inhibitor Y-27632 in a rat pilocarpine model of TLE. We used chronic i.p. injections of Y-27632 for 5 days in 6 week old control rats or rats subjected to pilocarpine treatment as a model of TLE. Surprisingly, our findings demonstrate that a systemic administration of Y-27632 in pilocarpine-treated rats increases neuronal death in the CA3 region and ectopic recurrent mossy fiber sprouting (rMFS) in the dentate gyrus of the hippocampal formation. Interestingly, we found that chronic treatment with Y-27632 exacerbates the down regulation and pathological distribution of the K+-Cl- cotransporter KCC2, thus providing a putative mechanism for post SE induced neuronal death. The involvement of astrogliosis in this mechanism appears to be intricate as ROCK inhibition reduces reactive astrogliosis in pilocarpine rats. Conversely, in control rats, chronic Y-27632 treatment increases astrogliosis. Together, our findings suggest that Y-27632 has a detrimental effect when chronically used post SE in a rat pilocarpine model of TLE.Peer reviewe

Current view on the functional regulation of the neuronal K+-Cl- cotransporter KCC2

Peer reviewe

EZcalcium: Open-Source Toolbox for Analysis of Calcium Imaging Data

Fluorescence calcium imaging using a range of microscopy approaches, such as two-photon excitation or head-mounted “miniscopes,” is one of the preferred methods to record neuronal activity and glial signals in various experimental settings, including acute brain slices, brain organoids, and behaving animals. Because changes in the fluorescence intensity of genetically encoded or chemical calcium indicators correlate with action potential firing in neurons, data analysis is based on inferring such spiking from changes in pixel intensity values across time within different regions of interest. However, the algorithms necessary to extract biologically relevant information from these fluorescent signals are complex and require significant expertise in programming to develop robust analysis pipelines. For decades, the only way to perform these analyses was for individual laboratories to write their custom code. These routines were typically not well annotated and lacked intuitive graphical user interfaces (GUIs), which made it difficult for scientists in other laboratories to adopt them. Although the panorama is changing with recent tools like CaImAn, Suite2P, and others, there is still a barrier for many laboratories to adopt these packages, especially for potential users without sophisticated programming skills. As two-photon microscopes are becoming increasingly affordable, the bottleneck is no longer the hardware, but the software used to analyze the calcium data optimally and consistently across different groups. We addressed this unmet need by incorporating recent software solutions, namely NoRMCorre and CaImAn, for motion correction, segmentation, signal extraction, and deconvolution of calcium imaging data into an open-source, easy to use, GUI-based, intuitive and automated data analysis software package, which we named EZcalcium

Impaired perceptual learning in a mouse model of Fragile X syndrome is mediated by parvalbumin neuron dysfunction and is reversible.

To uncover the circuit-level alterations that underlie atypical sensory processing associated with autism, we adopted a symptom-to-circuit approach in the Fmr1-knockout (Fmr1-/-) mouse model of Fragile X syndrome. Using a go/no-go task and in vivo two-photon calcium imaging, we find that impaired visual discrimination in Fmr1-/- mice correlates with marked deficits in orientation tuning of principal neurons and with a decrease in the activity of parvalbumin interneurons in primary visual cortex. Restoring visually evoked activity in parvalbumin cells in Fmr1-/- mice with a chemogenetic strategy using designer receptors exclusively activated by designer drugs was sufficient to rescue their behavioral performance. Strikingly, human subjects with Fragile X syndrome exhibit impairments in visual discrimination similar to those in Fmr1-/- mice. These results suggest that manipulating inhibition may help sensory processing in Fragile X syndrome

Hippocampal structural reactive plasticity in a rat model of temporal lobe epilepsy : chloride homeostasis as a keystone

Cette thèse a pour objectif spécifique d’explorer les événements précoces pouvant être à l’origine du bourgeonnement aberrant des fibres moussues (FM) du gyrus denté, une réorganisation majeure dans l’Epilepsie du Lobe Tempora (ELT). Nous avons utilisé le modèle pilocarpine d’ELT chez le rat afin de montrer que la transmission GABAergique jouait un rôle prépondérant dans la formation des FM aberrantes au cours de l’épileptogenèse. Ceci étant due à une altération de l’homéostasie chlore, suite à une augmentation de l’expression du co-transporteur NKCC1 et une diminution du co-transporteur KCC2. Nos résultats ont démontré que le récepteur aux neurotrophines p75NTR était un médiateur de l’action trophique de la réponse GABAergique dépolarisante sur le bourgeonnement aberrant des FM. Le blocage de l’action dépolarisante de la transmission GABAergique via l’utilisation de la bumétanide, a permis de réduire le bourgeonnement aberrant des MF en réduisant l’expression de p75NTR. Enfin, l’application transitoire de la bumétanide au cours de l’épileptogenèse a abouti à la réduction du nombre de crises récurrentes et spontanées au cours de la phase chronique d’ELT chez le rat. Ce travail a permis de dévoiler les mécanismes moléculaires sous-jacents de la réorganisation du réseau neuronal glutamatergique consécutif à une crise inaugurale dans un modèle d’ELT. Dans l'ensemble, cette thèse apporte un éclairage nouveau sur l’importance de l’interaction de la signalisation GABAergique avec les neurotrophines afin d’orchestrer la plasticité réactive au sein de l’hippocampe dans TLE.The present dissertation undertakes to investigate the early triggering events of the mossy fiber sprouting (MFS) in the dentate gyrus, a hallmark of hippocampal reactive plasticity in Temporal Lobe Epilepsy (TLE). We used the rat pilocarpine model of TLE to show that altered GABAA receptor-mediated transmission play a key role in the formation of early ectopic MFS during epileptogenesis. This is likely due to a compromised chloride homeostasis, as a result of increased expression of chloride loader NKCC1 and downregulation of the neuronal chloride extruder KCC2. We next addressed the mechanistic action of depolarizing GABAAR responses with regard to neurotrophin signaling. Our findings uncovered that the pan neurotrophin receptor p75 (p75NTR) mediated the sculpting action of depolarizing GABAAR responses on the ectopic MFS. Blockade of depolarizing GABAAR responses using the loop diuretic bumetanide reduced abnormal p75NTR subsequently decreased the ectopic MFS. Finally, transitory application of bumetanide during epileptogenesis resulted in reduction of spontaneous and recurrent seizures during the chronic phase of TLE. The rationale of this work is that unveiling the molecular mechanisms underlying the hippocampal post-seizure glutamatergic network rewiring will help to drive future novel therapeutic avenues involving chloride homeostasis and neurotrophin interplay. Overall, this dissertation shed a new light on how GABAergic transmission and neurotrophin signaling crosstalk can orchestrate reactive hippocampal plasticity in TLE

aGABRacadabra: A surprising new role for GABAA receptors in cortical development

In this issue of Neuron, Babij, Ferrer, and colleagues provide new evidence that β3 subunit of GABAA receptors is critical for the maturation of functional networks in the neonatal somatosensory cortex

Network state transitions during cortical development

Mammalian cortical networks are active before synaptogenesis begins in earnest, before neuronal migration is complete, and well before an animal opens its eyes and begins to actively explore its surroundings. This early activity undergoes several transformations during development. The most important of these is a transition from episodic synchronous network events, which are necessary for patterning the neocortex into functionally related modules, to desynchronized activity that is computationally more powerful and efficient. Network desynchronization is perhaps the most dramatic and abrupt developmental event in an otherwise slow and gradual process of brain maturation. In this Review, we summarize what is known about the phenomenology of developmental synchronous activity in the rodent neocortex and speculate on the mechanisms that drive its eventual desynchronization. We argue that desynchronization of network activity is a fundamental step through which the cortex transitions from passive, bottom-up detection of sensory stimuli to active sensory processing with top-down modulation

The NKCC1 inhibitor bumetanide restores cortical feedforward inhibition and lessens sensory hypersensitivity in early postnatal fragile X mice.

BackgroundExaggerated responses to sensory stimuli, a hallmark of Fragile X syndrome (FXS), contribute to anxiety and learning challenges. Sensory hypersensitivity is recapitulated in the Fmr1 knockout (KO) mouse model of FXS. Recent studies in Fmr1 KO mice have demonstrated differences in activity of cortical interneurons and a delayed switch in the polarity of GABA signaling during development. Previously, we reported that blocking the chloride transporter NKCC1 with the diuretic bumetanide, could rescue synaptic circuit phenotypes in primary somatosensory cortex (S1) of Fmr1 KO mice. However, it remains unknown whether bumetanide can rescue earlier circuit phenotypes or sensory hypersensitivity in Fmr1 KO mice.MethodsWe used acute and chronic systemic administration of bumetanide in Fmr1 KO mice and performed in vivo 2-photon calcium imaging to record neuronal activity, while tracking mouse behavior with high-resolution videos.ResultsWe demonstrate that layer (L) 2/3 pyramidal neurons in S1 of Fmr1 KO mice show a higher frequency of synchronous events at postnatal day (P) 6 compared to wild-type controls. This was reversed by acute administration of bumetanide. Furthermore, chronic bumetanide treatment (P5-P14) restored S1 circuit differences in Fmr1 KO mice, including reduced neuronal adaptation to repetitive whisker stimulation, and ameliorated tactile defensiveness. Bumetanide treatment also rectified the reduced feedforward inhibition of L2/3 neurons in S1 and boosted the circuit participation of parvalbumin interneurons.ConclusionsThis further supports the notion that synaptic, circuit, and sensory behavioral phenotypes in Fmr1 KO can be mitigated by inhibitors of NKCC1, such as the FDA-approved diuretic bumetanide