In order to achieve the proof-of-concept of the feasibility of the biohybrid approach to brain repair, Re.B.Us has combined microelectrode array (MEA) electrophysiology from brain slices with computational and engineering tools to restore the physiological function in an in vitro model of temporal lobe epilepsy (TLE). TLE is the most common partial complex epileptic syndrome and the most frequently unresponsive to pharmacological therapy. The hippocampus, a key brain area for learning and memory processes, is primarily involved in TLE and its functional impairment leads to progressive psychiatric and cognitive co-morbidities. In in vitro models of TLE, the functional interactions between the hippocampus and the parahippocampal cortex have been extensively characterized. Specifically, the CA3 hippocampal subfield plays a crucial role in controlling the propensity of the parahippocampal cortex to generate seizure activity. In epileptic tissue, the CA3 function is impaired and seizure activity generated by the cortex is unrestrained. The experimental work pursued in Re.B.Us has followed sequential logical steps based on these known network interactions in experimental TLE models. First, analysis of the CA3-driven pattern has revealed the presence of lognormal temporal dynamics and their preservation during acutely induced epileptiform behavior and in chronic epilepsy. Mimicking this pattern via open-loop lognormal electrical stimulation delivered to the subiculum (the hippocampal output gate) could successfully control seizure activity generation in the parahippocampal cortex. Further, this paradigm proved more efficient than periodic pacing, in that it delivered less electrical pulses, thereby imposing less stress to the brain tissue, while promising to improve the battery life of neuromodulators. These results anticipated the possibility of bridging the gap of communication between the hippocampus and the parahippocampal cortex to treat TLE. Thus, in the next set of experiments, the project pursued a closed-loop control strategy to functionally reconnect these two structures via electronic bridges. Specifically, three closed-paradigms were implemented: (i) unidirectional bridging within the same brain slice, (iii) unidirectional bridging between two distinct brain slices, and (ii) bidirectional bridging between two distinct brain slices. These experiments demonstrated the feasibility of bridging disconnected brain areas via a feedback loop to re-establish their physiological dialogue; specifically, they demonstrated that it is possible to exploit the intrinsic ability of CA3 to generate a sustained anti-ictogenic pattern to control cortical ictogenicity. Further, they heralded the possibility of using graft brain tissue to repair epileptic brain circuits, or, in a wider perspective, as a biological neuromodulator.
In pursuing the experimental work, Re.B.Us has developed software and hardware tools for advanced signal processing and seizure detection, stimulus pattern generation, and unidirectional and bidirectional closed-loop control, including a custom PCB purposely designed for Re.B.Us (Fig. 1) in collaboration with the company PCB Project (Lucca, Italy).
