Non–Cell Autonomous Epileptogenesis in Focal Cortical Dysplasia

Abstract

Objective: Low-level somatic mosaicism in the brain has been shown to be a major genetic cause of intractable focal epilepsy. However, how a relatively few mutation-carrying neurons are able to induce epileptogenesis at the local network level remains poorly understood. Methods: To probe the origin of epileptogenesis, we measured the excitability of neurons with MTOR mutation and nearby nonmutated neurons recorded by whole-cell patch-clamp and array-based electrodes comparing the topographic distribution of mutation. Computational simulation is used to understand neural network-level changes based on electrophysiological properties. To examine the underlying mechanism, we measured inhibitory and excitatory synaptic inputs in mutated neurons and nearby neurons by electrophysiological and histological methods using the mouse model and postoperative human brain tissue for cortical dysplasia. To explain non-cell-autonomous hyperexcitability, an inhibitor of adenosine kinase was injected into mice to enhance adenosine signaling and to mitigate hyperactivity of nearby nonmutated neurons. Results: We generated mice with a low-level somatic mutation in MTOR presenting spontaneous seizures. The seizuretriggering hyperexcitability originated from nonmutated neurons near mutation-carrying neurons, which proved to be less excitable than nonmutated neurons. Interestingly, the net balance between excitatory and inhibitory synaptic inputs onto mutated neurons remained unchanged. Additionally, we found that inhibition of adenosine kinase, which affects adenosine metabolism and neuronal excitability, reduced the hyperexcitability of nonmutated neurons. Interpretation: This study shows that neurons carrying somatic mutations in MTOR lead to focal epileptogenesis via non-cell-autonomous hyperexcitability of nearby nonmutated neurons.

Non-synaptic Cell-Autonomous Mechanisms Underlie Neuronal Hyperactivity in a Genetic Model of PIK3CA-Driven Intractable Epilepsy

“…Regardless, for the first time, our data indicates that acute seizure mechanisms are distinct and dependent on the location of the mutant protein within the PI3K-AKT-MTOR pathway. This conclusion is further supported by a recent study of a mouse model of MTOR activation, where neuronal hyperexcitation was found be driven by noncell autonomous mechanisms and not associated with altered intrinsic properties (Koh et al, 2021). Significant reduction of burst-generating cells, burst frequency and PPS by blocking voltage-dependent calcium or potassium channels indicate that Pik3ca E545K -driven hyperactivity is predominantly mediated by multiple altered cell-intrinsic properties.…”

“…Intriguingly, rapamycin analog RAD001 had minimal acute effect in our model, yet western analyses demonstrated pathway inhibition with RAD001 was similar to that seen with the other pathway drugs we assessed. While elevated MTOR activity is known to be epileptogenic (Crino, 2016;Kim and Lee, 2019;Koh et al, 2021), our data suggest that alternate PI3K-AKT targets acutely regulate neuronal hyperactivation in PIK3CAdriven epilepsy.…”

Cortical Dysplasia and the mTOR Pathway: How the Study of Human Brain Tissue Has Led to Insights into Epileptogenesis

“…Examination at 3-4 months showed that while cortical lamination was not affected by late production of Rheb CA , the mice exhibited spontaneous seizures and cytomegalic neurons, supporting the notion that neuronal misplacement may not be required for seizure generation [85]. A recent study added further insights into the mechanism of epileptogenesis in FCD by demonstrating that neurons carrying somatic MTOR variants can cause hyperexcitability in nearby non-mutated neurons in a non-cell autonomous manner [86].…”

Not So Innocent Bystanders in Focal Cortical Dysplasia

“…Interestingly, interneuron loss has been well documented in histopathologic studies on human tissues but not in Koh's study. A more detailed analysis of [1][2][3][4][5][6][7][8][9][10] genetically intact interneurons, particularly parvalbu-min+ fast-spiking cells, should be conducted in future studies. Fourthly, the intracellular injection of ADK in individual neurons did not modify extracellular adenosine concentration, suggesting that multiple neurons must have their metabolism altered to allow modification of adenosine levels.…”

“…2 Koh HY et al took advantage of episomal plasmid-based in utero electroporation (IUE) to probe the origin of epileptogenesis in dysplastic brains. 3 They generated a FCD mouse model with ∼2% cells in the IUE site ectopically overexpressing mutant MTOR. Their previous study has shown that these mice recapitulated the pathological features of FCDII and developed frequent generalized tonic-clonic seizures around P21.…”