Abstracts

Rationale: Hemodynamic surrogates of epileptic activity are being used to map epileptic foci with optical imaging methods. Recent data indicate that cerebral blood flow, although focally increased at the onset of a seizure, may be temporarily inadequate to meet the metabolic demands of both interictal and ictal epileptic events. However, much of this work has been performed on anesthetized animal models.  Since anesthesia is known to impact the excitability of the neuronal network as well as neurovascular coupling, we need to develop a new method to study the relationship between optical maps of blood flow, hemoglobin oxygenation and light scattering during epileptic events in awake animals in order to eliminate the effects of anesthesia.  Methods: We used a soft, penetrable, elastic, and transparent, silicone-based polydimethylsiloxane (PDMS) cranial window as a substitute for the skull and dura in mice (Heo C et al Scientific Reports 2016). A lightweight four-winged headplate was implanted to provide a long-term stable head fixation. The air-lifted table (Mobile HomeCage, Neurotar) was used for precise optical and electrophysiological recording in head-fixed awake mice. We induced acute focal ictal discharges in the mouse neocortex by injection of 4-aminopyridine (4-AP, 10mM, 0.5 µl).  The local field potential was recorded to identify the ictal discharges. Hemodynamic signals and light scattering were performed by intrinsic optical imaging at 565 nm, 617 nm and 780 nm. Results: Clear and healthy cortical vasculatures were observed up to 12?weeks post-implantation. Furthermore, the PDMS window allowed for easy insertion of microelectrodes and micropipettes into the cortical tissue for electrophysiological recording and chemical injection at any location without causing any fluid leakage. The mice can be habituated to the air-lifted table after 4 days of training. The mechanical stability of mouse’s head fixation allowed stable recordings in the brain of head-fixed and naturally behaving mice. No burst suppression were found during recording. Real-time intrinsic signals including hemodynamic responses and light scattering were successfully monitored during ictal discharge. Conclusions: We developed a novel method combining air-lifted platform and a chronic silicone-based injectable cranial window for optical imaging in awake epileptic mice. This method can be used for chronic in vivo intrinsic optical imaging of the neocortex over months. It can also be extended for imaging of neocortical structure (e.g., dendritic and axonal dynamics) or neuronal activity (e.g. calcium and glutamate signals) for long-term studies in awake animals.   Funding: Cornell University Ithaca-WCMC seed grant and the Daedalus Fund for Innovation