Melanopsin, a tri-stable photopigment found in intrinsically-photosensitive retinal ganglion cells (ipRGCs), drives circadian rhythms and other non-image forming functions in the nervous system. Despite increased understanding of the biomolecular and spectroscopic properties of melanopsin, its multiphoton and ultrafast optical absorption properties remain underexplored. We demonstrate the effects of two-photon absorption of melanopsin using 900-1160 nm optical stimulation. Excitation in this bandwidth causes consistent increases in calcium levels in transfected HEK293T cells. Our results demonstrate the first reported nonlinear optical properties and corresponding functional responses of two-photon excitation of melanopsin in vitro, along with the effects of spectral-phase modulation on activation.
One of the challenges in indirect measurements of electrical activity is its representation as biologically-relevant features. Non-invasive techniques for controlling neural activity, such as optogenetics with simultaneous optical imaging, have emerged as powerful and versatile tools. We demonstrated Superfast Polarization-sensitive Off-axis Full-field (SPoOF) OCM to image changes to both the optical phase and birefringence from the electrical activity of neurons at cellular resolution for an entire network at a millisecond scale. Here, we demonstrate all-optical neurophysiology with SPoOF OCM and optical excitation as a non-invasive versatile technique for studying neural circuits at high throughput, and a method to convert optical metrics to biologically relevant electrical features.
Active neurons experience rapid changes in their metabolic states since they have dynamic energy requirements. In this presentation, we demonstrate fast dual-channel label-free fluorescence lifetime imaging microscopy (FLIM) of NAD(P)H and FAD as a method for neurophysiology by performing computational photon counting in the onboard FPGA of the digitizer. The data throughput is reduced by 4x for each channel by compressing the photocurrents (16 bits) to photon counts (4 bits); the parallel processing on the FPGA ensures no lag. The setup was demonstrated for mammalian stem-cell-derived neurons under chemical stimulation, ion-channel blockers, and optical excitation. Fast FLIM on the FPGA enables dual-channel label-free metabolic optophysiology of neural activity in real time.
Access to the requested content is limited to institutions that have purchased or subscribe to SPIE eBooks.
You are receiving this notice because your organization may not have SPIE eBooks access.*
*Shibboleth/Open Athens users─please
sign in
to access your institution's subscriptions.
To obtain this item, you may purchase the complete book in print or electronic format on
SPIE.org.
INSTITUTIONAL Select your institution to access the SPIE Digital Library.
PERSONAL Sign in with your SPIE account to access your personal subscriptions or to use specific features such as save to my library, sign up for alerts, save searches, etc.