Targeting Kir4.1 To Control Brain Excitability And Seizures

Neuronal activity can rapidly elevate extracellular potassium in the brain. Potassium elevations are mechanistically related to increases in nerve cell excitability, potentially leading to abnormal network behaviours such as in epilepsy. Astroglia are essential for maintaining potassium balance, through a buffering mechanism engaging their sodium-potassium pumps. However, new evidence attributes the dynamic regulation of extracellular potassium mainly to astroglial channels of the Kir4.1 type whereas downregulation of these channels has been associated with enhanced seizure susceptibility and, in the long term, sclerotic abnormalities of brain tissue.

Despite the importance of Kir4.1 for brain excitability control, the causal relationships between Kir4.1 expression, potassium dynamics, local neuronal excitability and synaptic function remain poorly understood. The lack of progress stems from the poorly understood and often counteracting consequences of extracellular potassium rises, from the lack of tools to monitor potassium dynamics in brain tissue, and from the poor access to the sponge-like, nanoscopic morphology of astroglia.

The main goal of the present proposal is therefore to understand how the Kir4.1-dependent potassium buffering by astrocytes regulates neural excitability and synaptic circuit function, and whether targeting these mechanisms, pharmacologically or genetically, can alter susceptibility to runaway excitation and seizures. Thus, the central hypothesis is that the varied expression of astroglial Kir4.1 regulates, in a mechanistically predictable manner, cell excitability and synaptic signal transfer. The key translational aspect of the proposal is that the controlled manipulation of Kir4.1 expression should ameliorate pathological changes in neural excitability, such as those during epilepsy or cortical spreading depression.

To achieve our goal, we will take advantage of our novel and cutting-edge experimental and theoretical approaches. We have established gene-targeting protocols to enable single-cell studies in Kir4.1-overexpressing astrocytes while simultaneously monitoring neurotransmitter release at local synaptic connections. We have embarked on a novel nanoengineering technology to monitor potassium, which involves encapsulation of the ratiometric optical sensor into ion-permeable, biologically compatible microcapsules. We have established a novel biophysical modelling platform that enables theoretical probing of the intra- and extracellular potassium dynamics in realistic astrocyte models. We have developed a novel multi-electrode electrocorticography technique based on flexible graphene transistor arrays enabling full-band current recordings in awake animals.

These and related methodological breakthroughs, backed by a large body of pilot and proof-of-principle data, helped us to formulate a feasible research strategy for achieving our main goal. The plan includes five specific objectives addressed in five work packages. The results will provide new knowledge about the mechanisms by which astroglia regulate neuronal excitability through the Kir4.1-dependent control of extracellular potassium dynamics. Based on such knowledge, a therapeutic strategy could be developed that helps reduce brain susceptibility to runaway excitation, such as seen in epilepsy and related disorders.

Neural activity elevates extracellular K+ and thus boosts cell excitability, potentially leading to runaway excitation, such as epileptiform activity. Astrocytes dynamically control K+, mainly by engaging Kir4.1 channels, and Kir4.1 downregulation has been related to seizure susceptibility. However, the causal relationships between the Kir4.1 expression patterns, the external K+ dynamics, local neuronal excitability and synaptic function remain poorly understood. This knowledge gap hampers our search for feasible routes of therapeutic intervention.

We aim therefore to understand how the Kir4.1-dependent K+ buffering regulates network excitability and synaptic circuit function in the brain, and whether targeting Kir4.1 function, pharmacologically or genetically, can alter susceptibility to overexcitation and seizures. Our research strategy involves five specific objectives, taking advantage of the newly established techniques in acute slices and in vivo, including: gene-targeting protocols enabling visualisation of Kir4.1-overexpressing astrocytes while monitoring glutamate release at local synapses; a nano-technology to monitor extracellular K+ landscapes in situ using a microencapsulated ratiometric FRET dye; a computational platform ASTRO for theoretical probing of K+ dynamics in realistic astrocyte models; and a novel multi-electrode electrocorticography technique based on flexible graphene transistor arrays enabling full-band current recordings in awake animals.

These and related breakthroughs, backed by a large body of pilot results and proof-of-principle tests, helped formulate a feasible strategy to achieve our goal. The outcome will provide new knowledge about the mechanisms by which astroglia regulate neuronal excitability through the Kir4.1-dependent control of K+ dynamics. Based on such knowledge, a therapeutic strategy could be developed to help reduce brain susceptibility to runaway excitation, such as in epilepsy and related disorders