It is shown that a pathogenic variant of the G protein subunit Gβ1 impairs activation of neuronal G-protein-coupled inwardly rectifying potassium (GIRK) channels by inhibitory synaptic GABAB receptors, which leads to increased dendritic excitability and longer duration dendritic calcium spikes in mouse hippocampal neurons in response to stimulation of synaptic inputs.
Abstract
GNB1 encephalopathy (GNB1-E) is a rare neurodevelopmental disorder associated with motor dysfunction, epilepsy and learning disability caused by mutations in the gene encoding the G protein subunit Gβ1. Previous work has shown that altered Gβ1 can disrupt activation of G-protein-coupled inwardly rectifying potassium (GIRK) channels, dysregulate neuronal excitability and cause seizures. However, the relevant upstream regulators of Gβ1 and the consequences of GIRK dysfunction for neuronal synaptic, cellular and circuit function have not been characterized. Here we report that mice of both sexes carrying the deleterious p.I80T mutation in Gnb1 present features consistent with GNB1-E, including developmental delay, decreased locomotion and increased anxiety. Using histology, whole-cell patch-clamp electrophysiology and pharmacology in ex vivo brain slices, we find that hippocampal neurons in heterozygous Gnb1I80T/+ mice exhibit simplified dendritic morphologies, decreased synaptic inhibition mediated by metabotropic GABAB receptors and increased dendritic excitability. These phenotypes result in longer duration dendritic calcium spikes in response to synaptic afferent stimulation, an effect that is reversed by a specific activator of GIRK channels, ML297. Given the known roles of dendritic calcium spikes in driving burst firing and inducing synaptic plasticity, these findings suggest that targeting dendritic excitability has therapeutic potential to address both the seizure susceptibility and learning deficits associated with GNB1-E. Significance Statement GNB1 encephalopathy (GNB1-E) is a rare neurodevelopmental disorder associated with motor dysfunction, epilepsy and learning disability for which there are currently no mechanism-based treatments. Here we show that a pathogenic variant of the G protein subunit Gβ1 impairs activation of neuronal G-protein-coupled inwardly rectifying potassium (GIRK) channels by inhibitory synaptic GABAB receptors. This leads to increased dendritic excitability and longer duration dendritic calcium spikes in mouse hippocampal neurons in response to stimulation of synaptic inputs. We find that this phenotype is reversed by a drug that activates GIRK channels, opening pathways to develop therapies for GNB1-E that specifically target dendritic excitability.
The first mouse model of SCN1A GoF epilepsy with heterozygous Cre-dependent expression of the recurrent patient variant Scn1a-p.R1636Q is developed, the first study of SCN1A GoF epilepsy in a preclinical model in vivo and further investigation in the Scn1aflox(R1636Q)mouse will yield new mechanistic insights into disease mechanisms to drive advances in the treatment of SCN1A GoF epilepsy.
Sophie F. Hill, Z. P. Rosenthal, Ethan M. Goldberg· bioRxiv· 0 citations
The heterotrimeric G proteins are ubiquitous membrane-bound complexes specialized in the transduction of receptor-mediated extracellular signals into intracellular responses. Mutations in the G protein subunit alpha O1 (Gαo), encoded by GNAO1, have been associated with neurodevelopmental disorders characterized by prominent movement disorder with or without epilepsy. The Gnao1[C215Y]/+ mouse model recapitulates key features of the human disease, with a relatively mild phenotype, normal viability and late onset movement abnormalities, detectable in specific motor tasks. Here we report functional alterations in cortical layer V pyramidal neurons from Gnao1[C215Y]/+ mice, with reduction of the inward currents (Ih) and impairment of the GABAB-mediated outward current. We propose an adeno-associated virus (AAV)-based gene therapy aimed at overexpressing the wild-type form of Gαo by intracerebroventricular injection in newborn heterozygous Gnao1[C215Y]/ + mice. Our results demonstrate that overexpression of the wild-type protein mitigates behavioral abnormalities and restores functional neuronal deficits in young adult mice, thereby supporting the development of AAV-mediated gene augmentation strategies to counteract the effects of GNAO1 variants in patients. The potential of gene supplementation therapy is further supported by data from genetically modified C. elegans strains carrying not only the relatively mild C215Y variant but also a panel of goa-1/GNAO1 mutations associated with more severe phenotypes, suggesting a broader potential of this therapeutic strategy.
Germana Cocozza, T. D’Andrea, Martina Di Rocco et al.· Acta Neuropathologica Commun...· 0 citations
The integrated analysis establishes the first cross-species model for GNB1E, suggests common neurological mechanisms and molecular pathways linked to GNB1E, and provides a framework for mechanistic and therapeutic studies.
Haritha P. Reddy, Vigneshwar Ranjan, Merav Klo et al.· bioRxiv· 1 citation
Epilepsy is one of the most common neurological disorders, affecting more than 50 million people worldwide. Among the genetic etiologies of epilepsy, variants in genes coding for ion channels are vastly represented and characterized. Notably, loss-of-function (LoF) mutations in voltage-gated sodium channels (NaV) genes can result in a wide range of phenotypes including West syndrome, autism spectrum disorder, or Dravet Syndrome. Although the implication of NaV subtypes in epileptic syndromes and the relationship between seizures and inflammation have been extensively described, subtype-specific neuronal responses to inflammation in the context of NaV loss-of-function remain poorly understood. In this study, we investigated the consequences of subtype-specific downregulation of NaV expression in primary mouse cortical neurons. Using shRNA-mediated silencing of Scn1a, Scn2a, or Scn8a, we generated neuronal cultures with reduced expression of NaV1.1, NaV1.2, or NaV1.6 and evaluated neuronal survival, inflammatory gene expression, and global transcriptomic responses under basal conditions and following an inflammatory challenge. Subtype-specific NaV downregulations did not produce a uniform phenotype. Rather, minor differences under basal conditions led to important discrepancies following exposure to an inflammatory stimulus. Notably, NaV1.1 reduction was associated with synaptic transcriptional changes, whereas NaV1.6 downregulation led to a substantial inflammatory signaling remodeling. Our observations suggest that the consequences of NaV dysfunction are not only determined by their role in neuronal excitability but also depend on subtype-specific responses to inflammatory cues. They notably shed light on the relevance of inflammatory events in the onset and progression of epileptic syndromes related to NaV loss-of-function mutations.
Daniela Jacobsohn, David Guenoun, Nathalie Hertrich et al.· bioRxiv· 0 citations
The first comprehensive human mechanistic model of PPP1R9A haploinsufficiency using an isogenic CRISPR/Cas9-engineered iPSC system differentiated into cortical neurons is established, providing a human-specific mechanistic framework linking reduced Neurabin I dosage to neurodevelopmental and psychiatric disease risk.
Fragile X syndrome (FXS) is the leading monogenetic cause of intellectual disability and autism, yet how basic hippocampal circuit properties evolve across development for this condition remains unclear. Here, we studied CA1 pyramidal neurons in male Fmr1 knockout rats at postnatal day (P)12–15 and 6–10 weeks using ex vivo electrophysiology, pharmacology, imaging and biochemistry. P12–15 knockout neurons showed impaired sustained firing, progressive action potential broadening and enhanced activity-dependent synaptic vesicle replenishment. These defects recovered by 6–10 weeks. These phenotypes were linked to Kv7 channel dysfunction, as Kv7 activation altered action potential dynamics and neurotransmission in WT but not Fmr1 KO neurons. Rather than directly altering Kv7 channel function, Fmr1 knockout altered axon initial segment (AIS) development such that Kv7 became functionally inert. These findings suggest that this drives early, transient Kv7-dependent CA1 dysfunction in FXS and highlight how alterations in AIS development can have profound functional impacts.
Jack F. Webster, M. Pronot, Michael A. Cousin· bioRxiv· 0 citations
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