Abstract
Andrews JP, Geng J, Voitiuk K, Elliott MAT, Shin D, Robbins A, Spaeth A, Wang A, Li L, Solis D, Keefe MG, Sevetson JL, Rivera de Jesús JA, Donohue KC, Larson HH, Ehrlich D, Auguste KI, Salama S, Sohal V, Sharf T, Haussler D, Cadwell CR, Schaffer DV, Chang EF, Teodorescu M, Nowakowski TJ. Nat Neurosci 27(12):2487–2499. Seizures are made up of the coordinated activity of networks of neurons, suggesting that control of neurons in the pathologic circuits of epilepsy could allow for control of the disease. Optogenetics has been effective at stopping seizure-like activity in nonhuman disease models by increasing inhibitory tone or decreasing excitation, although this effect has not been shown in human brain tissue. Many of the genetic means for achieving channel rhodopsin expression in nonhuman models are not possible in humans, and vector-mediated methods are susceptible to species-specific tropism that may affect translational potential. Here we demonstrate adeno-associated virus-mediated, optogenetic reductions in network firing rates of human hippocampal slices recorded on high-density microelectrode arrays under several hyperactivity-provoking conditions. This platform can serve to bridge the gap between human and animal studies by exploring genetic interventions on network activity in human brain tissue.
Commentary
Optogenetics, the precise control of neural activity using genetically encoded, light-sensitive ion channels, has revolutionized systems neuroscience in animal models. Yet translation to human therapeutic applications has remained challenging. To date, the clinical use of optogenetics in humans has been extremely limited, with only one notable example: the partial restoration of vision in retinitis pigmentosa via optogenetic modulation of retinal ganglion cells. 1 Despite impressive results in animal models demonstrating its ability to rapidly and precisely modulate neuronal networks, 2 optogenetic technology has not yet been successfully translated into treatment for human epilepsy. Until now, network-level optogenetic modulation had not been demonstrated in slices of human brain tissue, though previous studies had shown optogenetic modulation at the single-cell level. 3
Andrews et al 4 address this crucial gap by establishing an innovative platform for evaluating optogenetic interventions in human hippocampal slices. Human hippocampal tissue was obtained from surgical resections in patients with drug-resistant temporal lobe epilepsy, including those with and without mesial temporal sclerosis. These hippocampal slices were transduced using adeno-associated virus serotype 9 to deliver the HcKCR1 channelrhodopsin, whose expression was driven specifically by a CAMK2A promoter. This promoter choice is significant because CAMK2A expression is predominant in excitatory, glutamatergic neurons—a cell type implicated in generating pathological hyperexcitability in epilepsy. 5 Thus, the study directly targets the therapeutic hypothesis that epilepsy results from an imbalance between neuronal excitation and inhibition.
HcKCR1, the inhibitory channelrhodopsin employed by Andrews et al, 4 opens a potassium channel upon illumination with 530 nm light, hyperpolarizing neurons and thus reducing their firing rate. Notably, this represents the first demonstration of inhibitory optogenetics successfully applied to human neural tissue at the network level. Before performing experimental validations, the authors introduced a computational model simulating the effect of optogenetic inhibition within excitatory neuronal populations. This computational approach is particularly commendable, providing a rigorous theoretical framework for predicting the expected impact of targeted neuronal inhibition and clearly outlining the assumptions underlying their experimental predictions.
As predicted by the computational model, optogenetic activation of HcKCR1 resulted in robust reductions in neuronal firing. The authors examined multiple pharmacologically induced hyperexcitable states, including a zero-magnesium (0-Mg) model of epilepsy, kainic acid (KA) administration (which further exacerbates excitability), and bicuculline-induced gamma amino butyric acid receptor blockade. Under each condition, optogenetic activation of HcKCR1 reduced neuronal firing, but importantly, the responses were heterogeneous: some neurons exhibited profound reductions (≥90% reduction in firing), while others showed partial or minimal responses. In particular, rhythmic bursting induced by KA was incompletely suppressed by optogenetic inhibition, highlighting the complex dynamics inherent in epileptic networks.
Despite effective neuronal inhibition, complete suppression of pathological network activity was not achieved. This indicates that presynaptic inhibition of a subset of cells might not be sufficient to prevent coherent epileptic activity, which is mediated by a complex interplay of postsynaptic potentials and intrinsic currents. This residual activity was observed even when accounting for differences in viral transduction efficiency. Additionally, using clustering methods based on extracellular spike waveforms, the authors identified distinct neuronal subpopulations—particularly granule cells of the dentate gyrus—that responded differently to optogenetic stimulation. These findings emphasize that understanding cell-type-specific impacts of optogenetic modulation is crucial for developing effective therapeutic strategies.
As another significant advance, Andrews et al 4 developed a closed-loop approach to optogenetic control, using real-time monitoring of neuronal firing rates. This approach allowed dynamic, automated illumination when epileptiform activity surpassed predefined thresholds, mimicking clinical paradigms such as responsive neurostimulation but with the added advantage of cellular specificity. 6 This offers insights unattainable through conventional electrical stimulation, which indiscriminately modulates all nearby cells, including excitatory neurons, interneurons, and glial cells.
There remain several limitations of this study. While the leap from animal models to human slices from patients with epilepsy gets closer to the type of brain we wish to treat clinically, a slice, by definition, lacks connectivity with the wider neural substrate in which it was originally embedded. This connectivity, both near and far, has repeatedly been implicated in the expression of seizures in people with epilepsy, and tied to surgical outcomes. 7 It will be fruitful to tie the insights uncovered by these simpler slice models with models of intact brains.
Another limitation that may prompt exciting additional work involves the acute versus chronic effects of this modulation. 6 In the study, modulation was, by the necessity of the slice model, brief. But other models such as organotypic cultures or organoids could begin to probe how consistent, prolonged stimulation, even if delivered in a closed-loop fashion, may begin to remodel these circuits. It may be that stimulation that works best acutely may fade in impact over time as the network adapts, and that different optogenetic stimulation paradigms work better at producing more long-lasting effects. It is an exciting area of exploration that is only now possible through the pioneering work done by this article and others.
In conclusion, Andrews et al 4 significantly advance our understanding of the translational potential of optogenetics in epilepsy research. By demonstrating network-level modulation of neuronal activity in human hippocampal slices, this study provides a crucial step toward clinical translation. Their approach brings us closer to realizing the clinical use of optogenetics, offering precise, cell-type-specific modulation, and highlighting both the promise and complexity of translating optogenetics into human epilepsy therapies.
