Abstract
Zhao J, Sun J, Zheng Y, Zheng Y, Shao Y, Li Y, Fei F, Xu C, Liu X, Wang S, Ruan Y, Liu J, Duan S, Chen Z, Wang Y. Nat Commun. 2022;13(1):7136. doi:10.1038/s41467-022-34662-2
Epileptic seizures are widely regarded to occur as a result of the excitation-inhibition imbalance from a neuro-centric view. Although astrocyte-neuron interactions are increasingly recognized in seizure, elementary questions about the causal role of astrocytes in seizure remain unanswered. Here we show that optogenetic activation of channelrhodopsin-2-expressing astrocytes effectively attenuates neocortical seizures in rodent models. This anti-seizure effect is independent from classical calcium signaling, and instead related to astrocytic Na+-K+-ATPase-mediated buffering K+, which activity-dependently inhibits firing in highly active pyramidal neurons during seizure. Compared with inhibition of pyramidal neurons, astrocyte stimulation exhibits anti-seizure effects with several advantages, including a wider therapeutic window, large-space efficacy, and minimal side effects. Finally, optogenetic-driven astrocytic Na+-K+-ATPase shows promising therapeutic effects in a chronic focal cortical dysplasia epilepsy model. Together, we uncover a promising anti-seizure strategy with optogenetic control of astrocytic Na+-K+-ATPase activity, providing alternative ideas and a potential target for the treatment of intractable epilepsy.
Commentary
Astrocytes are resident glial cells which regulate neuronal functions to an extent that could at times make them sovereigns of neurons. One astrocyte may oversee more than 100,000 synapses and can control them by versatile pathways. 1 One such pathway, “gliotransmission” is the calcium-dependent release of neurotransmitters. In addition, astrocytes contribute to neuronal development, store and distribute energy substrates, buffer the extracellular levels of ions, help maintain the integrity of the blood–brain barrier (BBB), and participate in immune functions. 1,2
Given the multitude of astrocytic duties, it is not surprising that their dysfunction has been implicated in both epileptogenesis and ictogenesis. Virtually any of the aforementioned astrocytic functions can go wrong in epilepsy. Yet we are still far from understanding how astrocytes contribute to the generation of seizures, due to the many challenges in their investigation. First, astrocytes are a heterogenous group, with small astrocytic subsets being capable of promoting or limiting neurological disease 3 ; second, reactive and compensatory phenotypes are not readily distinguishable 1 ; and third, separating the precise impact of astrocytes from those of nearby neurons has been impeded by the inability to selectively modulate astrocyte activity. The third hurdle has been addressed in a recent study by Zhao and colleagues, 4 who investigated in vivo the role of astrocytes in the generation of neocortical seizures through the application of optogenetics.
The selectivity of optogenetics resides in the cellular coexpression of an opsin gene and a driver protein. Consequently, cells that are naturally light-insensitive can be manipulated by illumination. 5 To selectively manipulate astrocytes, the authors used mice which express the driver protein Cre in astrocytes but not in neurons or oligodendrocytes. 4 Astrocytes were transfected in vivo with the opsin channelrhodopsin-2 (ChR2) using an adeno-associated virus injected into the primary motor cortex. Next, neocortical seizures were induced by local injection of kainic acid. Photostimulation of the astrocytes attenuated seizure progression and completely eliminated generalized seizures. The astrocyte-mediated anti-seizure activity did not depend on the stimulation frequency, was detected in both the early and late phases of neocortical seizures, sustained over a long duration (minutes), and the occurrence of generalized seizures was delayed even in remote brain regions. These experiments demonstrate that astrocytes can effectively control both seizure initiation and seizure spread.
In contrast to the impressive effects of indirect neuronal inhibition (via astrocytic photostimulation), direct optogenetic inhibition of pyramidal neurons had only a narrow temporal window for its anti-seizure effects and it did not inhibit seizure spread. In addition, the blue light that stimulated astrocytes did not alter locomotor activity, whereas neuronal activation by a yellow light was associated with impaired motor function, suggesting that at least it that experimental setting, astrocytic stimulation had fewer adverse effects.
Zhao et al next investigated the mechanism of this anti-seizure effects of astrocyte stimulation. ChR2 is a cation channel which translocates Ca2+ and Na+ into cells. Indeed, Ca2+ infusion into astrocytes during ChR2 channel activation resulted in higher intracellular calcium level. Calcium was involved in seizure initiation in the early stage but was not obligatory for the seizure-suppressing effect of astrocyte stimulation. Instead, the anti-seizure activity was mediated by stimulation of the astrocytic Na+K+ATPase as a result of Na+ influx. A similar effect was observed in a rat model of focal cortical dysplasia. This time, activation of the cation pump required pretreatment with pentylenetetrazole, because spontaneous epileptiform activity in this model isinfrequent, requiring stimulation with light over several hours to capture a seizure. 4
Over the Rainbow
Translational aspects of optogenetics are not straightforward. One key limitation is tissue heating as a result of light application over a prolonged period, which may occur if seizures are rare and stimulation is continuous (in contrast to closed-loop systems). Other potential barriers include the absence of safe and effective methods for gene and light delivery into deep brain tissue and the need for activating larger brain structures in humans compared with rodents. 5 In addition, asimilarity between the operation manuals of rodent and human astrocyte-neuron communication has yet to be validated. Yet the featured study demonstrates the key strength of optogenetic techniques: controlling the activity of selected cells within a heterogeneous tissue.
In neuroscience, the use of optogenetics has been generally limited to the investigation of neuronal activity. 5 Previous articles in this journal provided insights into potential applications of this powerful tool in studying the pathogenesis of epilepsy, including a general overview of optogenetics and neuronal activity 6 and a review of studies in interneurons. 7 The current optogenetic toolbox includes new opsins (e.g., enzyme, potassium, and channel kinase signaling rhodopsins) with new sensors of diverse excitation spectra, a wider assortment of delivery methods for improved penetration of genes across BBB and for cell targeting, and novel light delivery techniques. Optogenetics may be applied to refine deep brain stimulation protocols 5,6 and new implications in cardiology can shed light on the mechanisms that lead to sudden unexpected death of epilepsy. Moreover, instead of a gene product, the photoswitchable entity may be a drug, making gene delivery unnecessary. 5,8
Old Targets, New Tricks
Astrocytes are usually bystanders rather than the main targets of anti-seizure treatments, including the ketogenic diet and the anti-seizure medications valproic acid, phenytoin, and gabapentin. 1 The paradigm shift over the past 2 decades from neurons to additional cell types of the brain raises the question: can we develop drugs that selectively affect astrocytes? Such treatments may have fewer adverse effects than those directing neurons, further increasing their translational appeal. Novel astrocyte-targeting agents have shown promise in rodent models of epilepsy and epileptogenesis. These include the purinergic receptor antagonists MRS2179 and MRS2500 and the Panx1 channel inhibitors probenecid and mefloquine. 1 Unfortunately, the physicochemical characteristics of many of these compounds meet the criteria to become CNS drugs but not anti-seizure medications, for example, due to their relatively large molecular weight. 9
The featured study not only extends the milieu of potential astrocytic drug targets but also paves the way for novel stimulation-based treatments, for example, during deep-brain stimulation. The 2 approaches (drug development and optogenetics) may be combined. One example is the use of photoswitchable astrocytic activators. Optogenetic interventions are being evaluated for vision restoration in patients with retinal diseases (clinicaltrials.gov). With further improvements, they are hoped to contribute to the control of drug-resistant epilepsy.
