Abstracts

Rationale: Impaired consciousness during partial temporal-lobe seizures has puzzled clinicians and researchers for decades. Why would focal seizures in the temporal lobe, which is required for memory and other functions but not for maintaining consciousness, cause impaired consciousness? Clarifying the network mechanisms underlying ictal unconsciousness is crucial for developing effective treatments for this profoundly debilitating side-effect of seizures. According to the network inhibition hypothesis , temporal lobe seizures impair consciousness through inhibition of subcortical arousal pathways. This inhibition in turn causes a transition of the cortex into a sleep-like mode characterized by slow-wave activity.Methods: To test the network inhibition hypothesis, we developed an optogenetic stimulation approach in rats to selectively activate subcortical cholinergic arousal circuits. The light-sensitive non-specific cation channel channelrhodopsin-2 (ChR2) was delivered selectively to cholinergic neurons of the pedunculopontine tegmental nucleus (PPT) using stereotaxic virus injections of a double-floxed ChR2 construct (rAAV5/ EF1 -DIO-ChR2-eYFP, H134R) into the PPT of transgenic rats expressing Cre recombinase specifically in cholinergic neurons (ChAT-Cre rats). 473nm laser light was delivered via a 200 m optical fiber implanted into the PPT.Results: First, we used histology to confirm delivery of ChR2 selectively to cholinergic neurons in the PPT. Second, we conducted extracellular electrophysiological recordings from the PPT while delivering laser light to this brain region. We confirmed and optimized the stimulation paradigm to control the firing of PPT neurons using a broad range of stimulation frequencies (5-60Hz) and light-pulse durations (1-40msec). Using light stimulation under deep anesthesia (unconscious state), we were able to generate PPT neuron firing rates characteristic of the awake state (conscious state).Conclusions: These findings demonstrate, for the first time, the ability to control spiking activity of cholinergic PPT neurons with high spatial and temporal precision. In future experiment we will combine optogenetic stimulation of cholinergic neurons in the PPT with electrophysiological recordings from the neocortex during induced hippocampal seizures. This novel approach will help to clarify the circuit mechanisms of ictal unconsciousness and hopefully pave the way to novel treatments aimed at restoring consciousness during seizures.