Suryavanshi P, Langton R, Fairhead K,  Glykys J. Brief and diverse excitotoxic insults increase the neuronal nuclear membrane permeability in the neonatal brain, resulting in neuronal dysfunction and cell death. J Neurosci. 2024 Aug 30:e0350242024. doi: 10.1523/JNEUROSCI.0350-24.2024.          [Cover of J. Neuroscience journal Oct 30]

• Our study demonstrates that different insults that model diverse neurological conditions enlarge the neuronal nuclear pores in the developing brain. 
• The enlargement of nuclear pores caused a rapid nuclear translocation of cytosolically localized proteins like GCaMP6s. 
• The increased nuclear translocation in neurons was associated with attenuated neocortical circuit activity and neuronal death. 
• Calpain inhibition prevented excitotoxicity-induced nuclear translocation of GCaMP6s and neuronal swelling. 
• Since GCaMP6 variants are widely used in neuroscience, their abnormal nuclear translocation can be used to study early neuronal injury.
• Our findings have clinical implications for treating brain injury and neuronal swelling during early brain development, which currently lack direct pharmacological treatments. 

Suryavanshi P, Baule S, Glykys J. Trauma in neonatal acute brain slices alters calcium and network dynamics and causes calpain-mediated cell death. eNeuro 17 June 2024, ENEURO.0007-24.2024; https://doi.org/10.1523/ENEURO.0007-24.2024

• This study characterizes the spatiotemporal dynamics of neocortical injury in acute neonatal slices, mimicking severe penetrating traumatic brain injury, using PI labeling and elevated neuronal Ca²⁺ load as markers for cytotoxicity. 
• There was depth- and time-dependent neuronal damage after brain slicing, leading to altered neuronal responses. 
• Elevate neuronal Ca²⁺ and cytotoxicity were mitigated by pharmacologically inhibiting calpains, a family of Ca²⁺-dependent proteases involved in multiple cell death mechanisms.
• Our study provides evidence for injury-dependent neuronal and circuit function alterations in neonatal acute brain slices. Calpain inhibition decreased trauma-induced cell death in the neonatal brain, identifying them as potential therapeutic targets at this age.

Liddiard GT, Suryahvanshi PS, Glykys J. Enhancing GABAergic tonic inhibition reduces seizure-like activity in the neonatal mouse hippocampus and neocortex. Journal of Neuroscience 4 January 2024, e1342232023; DOI: https://doi.org/10.1523/JNEUROSCI.1342-23.2023

• Around 30-50% of neonatal seizures resist current treatments.
• Whether enhancing tonic inhibition decreases seizure-like activity in the neonate when GABA is mainly depolarizing at this age is unknown. 
• We showed that THIP, a δ-subunit-selective GABA_A receptor agonist, enhanced GABAergic tonic inhibitory conductances in layer V neocortical and CA1 pyramidal neurons and increased their rheobase without altering sEPSCs characteristics.
• Two-photon calcium imaging demonstrated that enhancing the activity of extrasynaptic GABA_ARs decreased neuronal firing in both brain regions.
• In in vitro seizure models, THIP reduced epileptiform activity in the neocortex and CA1 hippocampal region of neonatal and adult brain slices in a dose-dependent manner.

• Our results suggest that augmenting tonic inhibition could be an alternative treatment for neonatal seizures, especially in pharmacy-resistant ones.

  
   

Islam MS, Suryavanshi P, Baule SM, Glykys J*, Baek S*. A Deep Learning Approach for Neuronal Cell Body Segmentation in Neurons Expressing GCaMP using a Swin Transformer. eNeuro. 2023 Sep 13; ENEURO.0148-23.2023. PMID: 37704367. [*, Corresponding authors]

• Neuronal segmentation is challenging due to the complex morphological structures of neuronal cell bodies and their surroundings.
•  We developed a deep-learning-based approach for automated, fast, robust 2D neuronal cell body segmentation of neurons expressing fluorescent markers using a state-of-the-art vision transformer (Swin transformer).

• This approach for neuronal cell body segmentation can assist researchers in evaluating the changes in neuronal cell body sizes under different pathological conditions.

  
   

Suryavanshi P, Langton R, Fairhead K, Glykys J. Brief excitotoxic insults cause a calpain-mediated increase in nuclear membrane permeability in neonatal neurons. bioRxiv. 2023, Aug 23;2023.08.22.554167. doi: 10.1101/2023.08.22.554167. 

• We showed rapid nuclear translocation of a genetically encoded calcium sensor (GCaMP6s) a few minutes after different and common brain insults in neonatal neurons using in and ex-vivo multiphoton imaging.
• We demonstrate that various brief brain insults that model hypoxic events, seizures, and brief NMDA receptor activation enlarge nuclear pores through a calpain-mediated mechanism.
• The enlargement of the nuclear pores caused large proteins like GCaMP6s to lose their subcellular localization and disrupt nuclear function.
• This mechanism may underlie neuronal injury in the neonatal period. 

Takezawa Y, Langton R, Baule SM, Zimmerman MB, Baek J. Glykys, J. Role of NKCC1 and KCC2 during hypoxia-induced neuronal swelling in the neonatal neocortex. Neurobiol. Dis. 2023. Mar;178:106013. PMID: 36706928

• We used two-photon laser scanning microscopy to image neurons expressing a genetically encoded fluorescent protein sensitive to chloride concentrations.
• We analyzed the neurons using a convolutional neural network.  
• We demonstrated that cation-chloride cotransporters (CCCs) contribute to water movement in neocortical neurons during oxygen-glucose deprivation (OGD) in the neonatal period.
• Blocking NKCC1, a type of cation-chloride cotransporter, decreased neuronal swelling during early OGD.
• Furosemide, a diuretic, decreased neuronal swelling during early and prolonged OGD through additional pathways besides CCCs.
• We conclude that CCCs and other non-CCCs contribute to water movement in neocortical neurons during OGD in the neonatal period.

Langton R, Sharma S, Tiarks GC, Bassuk AG, Glykys, J. Lacosamide decreases neonatal seizures without increasing apoptosis. Epilepsia. 2022. Dec;63(12):3051-3065. PMID: 36168798

• This paper showed that the antiseizure medication lacosamide decreased epileptiform activity in neonates in vitro (A) and behavior seizures in vivo. 
• Significantly, lacosamide did not increase neuronal apoptosis acutely in vitro or in vivo (B, C). 

Tong L, Langton R, Glykys J, Baek S. ANMAF: An automated neuronal morphology analysis framework using convolutional neural networks. Scientific Reports. 2021 April 14;11(1):8179. PMID: 33854113. 

• We developed a novel high-throughput neuronal morphology analysis framework (ANMAF), using convolutional neural networks (CNN) to automatically contour the somatic area of fluorescent neurons in acute brain slices.
• ANMAF produced consistent neuronal contours compared to humans, who exhibited significant variability in repeated measurements.
• ANMAF was generalizable across different imaging protocols and trainable even with a small number of humanly labeled neurons. 
• Our framework can facilitate more rigorous and quantitative studies of neuronal morphology by enabling the segmentation of many fluorescent neurons in thick brain slices in a standardized manner.