Embryonic nutritional hyperglycemia decreases cell proliferation in the zebrafish retina.

Diabetic retinopathy (DR) is one of the leading causes of blindness in the world. While there is a major focus on the study of juvenile/adult DR, the effects of hyperglycemia during early retinal development are less well studied. Recent studies in embryonic zebrafish models of nutritional hyperglycemia (high-glucose exposure) have revealed that hyperglycemia leads to decreased cell numbers of mature retinal cell types, which has been related to a modest increase in apoptotic cell death and altered cell differentiation.

Optical Switching Between Long-lived States of Opsin Transmembrane Voltage Sensors.

Opsin-based transmembrane voltage sensors (OTVSs) are membrane proteins increasingly used in optogenetic applications to measure voltage changes across cellular membranes. In order to better understand the photophysical properties of OTVSs, we used a combination of UV-Vis absorption, fluorescence and FT-Raman spectroscopy to characterize QuasAr2 and NovArch, two closely related mutants derived from the proton pump archaerhodopsin-3 (AR3). We find both QuasAr2 and NovArch can be optically cycled repeatedly between O-like and M-like states using 5-min exposure to red (660 nm) and near-UV (405 nm) light.

Analog Retinal Redshifts Visible Absorption of QuasAr Transmembrane Voltage Sensors into Near-infrared.

Opsin-based transmembrane voltage sensors (OTVSs) are increasingly important tools for neuroscience enabling neural function in complex brain circuits to be explored in live, behaving animals. However, the visible wavelengths required for fluorescence excitation of the current generation of OTVSs limit optogenetic imaging in the brain to depths of only a few mm due to the strong absorption and scattering of visible light by biological tissues. We report that substitution of the native A1 retinal chromophore of the widely used QuasAr1/2 OTVSs with the retinal analog MMAR containing a methylamino-modified dimethylphenyl ring results in over a 100-nm redshift of the maxima of the absorption and fluorescence emission bands to near 700 and 840 nm, respectively.

Electronic Preresonance Stimulated Raman Scattering Imaging of Red-Shifted Proteorhodopsins: Toward Quantitation of the Membrane Potential.

Voltage imaging allows mapping of the membrane potential in living cells. Yet, current intensity-based imaging approaches are limited to relative membrane potential changes, missing important information conveyed by the absolute value of the membrane voltage. This challenge arises from various factors affecting the signal intensity, such as concentration, illumination intensity, and photobleaching.

Raman spectroscopy of a near infrared absorbing proteorhodopsin: Similarities to the bacteriorhodopsin O photointermediate.

Microbial rhodopsins have become an important tool in the field of optogenetics. However, effective in vivo optogenetics is in many cases severely limited due to the strong absorption and scattering of visible light by biological tissues. Recently, a combination of opsin site-directed mutagenesis and analog retinal substitution has produced variants of proteorhodopsin which absorb maximally in the near-infrared (NIR).