Abstracts

Rationale: The emergence of optogenetics allows select modulation of discrete subpopulations of neurons at a millisecond temporal resolution. The genetic sequence required for opsin expression can be virally transduced, and post-expression, specific wavelengths of light may be delivered to the transduced neurons to activate opsins. Depending on the opsin, channelrhodopsin-2 (ChR2) or halorhodopsin (NpHR), the neurons may be depolarized or hyperpolarized with light. This technology offers a new scientific methodology to study epilepsy and even offers possible clinically relevant therapies. Currently, most optical technologies for in vivo central nervous system studies use laser light sources coupled to a tethered optical fiber. Additionally, optical windows with high intensity, large current (100's of mA) consuming LEDs have been implemented. However, to activate ChR2, a minimum of 1 mW per square meter is needed, and a benchmark of 10 mW per square meter is often reported. These irradiance levels can be theoretically reached with low-current LEDs, and development of such a device would offer the potential to be powered wirelessly. Because this enables experiments in untethered freely moving mice, it would be an important contribution to the epilepsy field.Methods: A 200 um core diameter optical fiber is stripped to the cladding and butt coupled to a low powered LED. An off-the-shelf constant current stimulator is matched to the LED specifications to operate between 2.7 and 3.2 V and supply up to 20 mA. The device is referred to as the optogenetic stimulator (OGS). Optical irradiance was measured with an optical power meter, and optical stimulation was tested in transverse hippocampal slices from PV-Cre mice virally (AAV5) transduced with ChR2. Expression of ChR2 was Cre-dependent.Results: Stimulation at 2.85 mA produced an irradiance of 2.7 (SD 0.0002) mW per square millimeter at the fiber tip. 10.55 (SD 0.008) mW per square millimeter was reached at 13.01 mA. Robust stimulation capable of eliciting action potentials from PV cells expressing ChR2 was observed at a current as low as 3 mA. At 3 mA, an action potential firing frequency of over 200 Hz was achievable.Conclusions: The low current consuming OGS was able to deliver sufficient irradiance at 473 nm to activate ChR2 expressing neurons. Such a device can be wirelessly powered and wirelessly programmed for use in virally transduced optogenetic studies. Funding sources: Cyberonics, Inc. (to S.L., O.L., P.P.I.) George E. Hewitt Foundation for Medical Research (to E.K.-M.), NIH NS074702 (to I.S., E.K.-M., C.A.)