Fast intrinsic optical signal correlates with activation phase of phototransduction in retinal photoreceptors

Abstract

As the center of phototransduction, retinal photoreceptors are responsible for capturing and converting photon energy to bioelectric signals for following visual information processing in the retina. This article summarizes experimental observation and discusses biophysical mechanism of fast photoreceptor-intrinsic optical signal (IOS) correlated with early phase of phototransduction. Quantitative imaging of fast photoreceptor-IOS may provide objective optoretinography to advance the study and diagnosis of age-related macular degeneration, retinitis pigmentosa, diabetic retinopathy, and other eye diseases that can cause photoreceptor dysfunctions.

Keywords: Optoretinography; age-related macular degeneration; diabetic retinopathy; intrinsic optical signal; optical coherence tomography; optophysiology; photoreceptor; phototransduction; retinitis pigmentosa.

Figure 1.

IOS imaging of a freshly isolated frog retina (Rana pipiens). (a) Representative raw image sequence of the retina. The images were recorded with a CCD camera at a speed of 80 frames/s. The white spot in the third frame shows the visible stimulus pattern. (b) Corresponding IOS images. Each frame is an average over a 100 ms interval (8 frames). 200 ms prestimulus and 500 ms poststimulus images are shown. (c) Enlarged image of the third frame shown in (b). (d) Temporal profiles of IOS change. Tracings 1 to 3 are representative IOS of individual pixels, corresponding to three points of arrows in (c). Tracing 4 represents the averaged optical response of the whole image area. The vertical line indicates the stimulus onset. ERG: electroretinography. Reprinted with permission from Yao and Zhao. (A color version of this figure is available in the online journal.)

Figure 2.

Stimulus-evoked photoreceptor movement in a mouse retina (Mus musculus). (a) NIR microscopic image of mouse photoreceptor mosaic. A 40× objective with 0.75 NA was used. The image size corresponds to a 60 × 60 µm² area on the retina. The green dashed rectangle indicates the oblique stimulation area. (b) Displacements of 10 photoreceptors over time. The stimulus was delivered at time 0 s. These 10 photoreceptors were specified by arrows in (a). Blue arrows in circles indicate the direction of the displacement at 30 ms after light stimulus. (c) Averaged displacement of 10 photoreceptors. The inset panel shows the same data within the period from −0.02 to 0.1 s. Reprinted with permission from Lu et al.⁴⁰ (A color version of this figure is available in the online journal.)

Figure 3.

Stimulus-evoked OS length shrinkage of frog rod photoreceptor (Rana pipiens). (a) Representative microscopic images of a single isolated rod OS acquired at intervals of 0.5 s. To better show the light-evoked OS shrinkage, the base of the rod OS in each image is aligned horizontally following the black solid line at the bottom. The black-dash line at the top represents the position of the rod OS tip at time −1 s. Scale bars represent 5 µm. (b) Enlarged picture of the white rectangle in (a). Scale bars represent 2 µm. (c) Normalized OS length and diameter changes from 8 different rod OSs. A shaded area indicates the 1 s stimulation period. Reprinted with permission from Lu et al.⁴⁴ (A color version of this figure is available in the online journal.)

Figure 4.

Comparative measurement of stimulus-evoked ROS movement and ERG of frog retina (Rana pipiens). (a) Representative image of ROS acquired at a frame speed of 500 fps. Temporal profiles of (b1) averaged ROS movement and (c1) averaged ERG response from 10 different retinal locations. (b2) Enlarged picture of the dashed square in (b1). (c2) Enlarged picture of the dashed square in (c1). The red triangles in (b2) and (c2) indicate the onset times determined by the 3-δ threshold of ROS movement and ERG a-wave, respectively. Shaded areas in (b1), (b2), (c1), and (c2) represent stimulation periods. ERG: electroretinography; ROS: rod outer segment. Reprinted with permission from Lu et al.⁴⁵ (A color version of this figure is available in the online journal.)

Figure 5.

Comparative measurement of stimulus-evoked ROS movements and ERGs acquired from frog retinas (Rana pipiens) with (a) control Ringer’s medium, (b) low sodium medium, and (c) recovery groups. Shaded areas represent stimulation periods. Modified with permission from Lu et al.⁴⁵ ERG: electroretinography; ROS: rod outer segment. (A color version of this figure is available in the online journal.)

Figure 6.

(a) In vivo IOS imaging and ERG measurement of frog retina (Rana pipiens). Each illustrated frame is an average over 100 ms interval (20 frames). The black arrowhead indicates the onset of the 10 ms green flash stimulus. 200 ms pre-stimulus baseline and 900 ms post-stimulus IOS recordings are shown. (b) Representative IOS responses of individual pixels randomly selected from the image area. The gray bar indicates the stimulus onset and duration. (c) The top black trace shows the IOS magnitude (i.e. absolute value of the IOS) averaged over the whole image area, corresponding to the image sequence shown in (a). The gray trace shows one control experiment without stimulation. The black trace below shows concurrent frog ERG. The gray bar indicates the stimulus onset and duration. ERG: electroretinography; IOS: intrinsic optical signal. Reprinted with permission from Zhang et al.⁵⁴ (A color version of this figure is available in the online journal.)

Figure 7.

Functional OCT of stimulus-evoked neurovascular responses in a mouse retina (C57BL/6J). (a1) Representative flattened OCT B-scan and (a2) spatiotemporal neural-IOS map. (b1) Representative flattened OCTA B-scan and (b2) spatiotemporal hemodynamic-IOS map. Scale bars in (a1) and (b1) indicate 500 µm. (c) Neural-IOS changes of PL, OPL, IPL, and GCL. (d) Hemodynamic-IOS changes of SVP, ICP, and DCP. (e) Averaged onset times of neural-IOS changes at PL, OPL, IPL, and GCL. (f) Averaged onset times of hemodynamic-IOS changes of SVP, ICP, and DCP. SVP: superficial vascular plexus; ICP: intermediate capillary plexus; DCP: deep capillary plexus; GCL: ganglion cell layer; IPL: inner plexiform layer; OPL: outer plexiform layer; PL: photoreceptor layer. Reprinted with permission from Yao et al. (A color version of this figure is available in the online journal.)

Figure 8.

Phototransduction activation. Step 1: Incident photon (hν) is absorbed and activates a rhodopsin to R*. Step 2: R* makes repeated contacts with transducin molecules, catalyzing its activation to G*. Step 3: G* binds inhibitory γ subunits of the PDE activating its α and β subunits. Step 4: Activated PDE hydrolyzes cGMP. Step 5: GC synthesizes cGMP. Reduced levels of cytosolic cGMP cause cyclic nucleotide gated channels to close, preventing further influx of Na⁺ and Ca²⁺. GC: guanylyl cyclase; PDE: phosphodiesterase; GDP: guanosine diphosphate; GTP: guanosine triphosphate; GMP: guanosine monophosphate; cGMP: cyclic guanosine monophosphate. Reprinted from https://en.wikipedia.org/wiki/Visual_phototransduction. (A color version of this figure is available in the online journal.)

Figure 9.

Comparative photoreceptor-IOS measurements in WT (C57BL/6J) and rd10 (B6.CXB1-Pde6b^rd10/J) mice at P14 and P16. (a1) and (a2) show averaged photoreceptor-IOS responses recorded from 12 WT and 12 rd10 retinas at P14. (b1) and (b2) are statistics of peak amplitudes and time-to-peaks corresponding to the data shown in (a1) and (a2), respectively (n = 12, NS = not significant). Averaged magnitude-temporal curve of photoreceptor-IOS responses from (c1) 12 WT mouse retinas and (c2) 12 rd10 mouse retinas at P16. (d1) and (d2) are statistics of peak amplitudes and time-to-peaks corresponding to the data shown in (c1) and (c2), respectively (n = 12, NS = not significant, **P < 0.05). Significance was determined by a two-sample t-test with equal variance assumed. The normality of data was determined using the Kolmogorov-Smirnov test. Each solid curve in (a) and (c) represents the mean values, and the accompanied colored area represents the corresponding standard deviations of the photoreceptor-IOS responses. The gray shaded areas represent stimulus duration. Reprinted with permission from Lu et al.⁴⁶ (A color version of this figure is available in the online journal.)