Biophotons Contribute to Retinal Dark Noise

Abstract

The discovery of dark noise in retinal photoreceptors resulted in a long-lasting controversy over its origin and the underlying mechanisms. Here, we used a novel ultra-weak biophoton imaging system (UBIS) to detect biophotonic activity (emission) under dark conditions in rat and bullfrog (Rana catesbeiana) retinas in vitro. We found a significant temperature-dependent increase in biophotonic activity that was completely blocked either by removing intracellular and extracellular Ca(2+) together or inhibiting phosphodiesterase 6. These findings suggest that the photon-like component of discrete dark noise may not be caused by a direct contribution of the thermal activation of rhodopsin, but rather by an indirect thermal induction of biophotonic activity, which then activates the retinal chromophore of rhodopsin. Therefore, this study suggests a possible solution regarding the thermal activation energy barrier for discrete dark noise, which has been debated for almost half a century.

Keywords: Biophoton; Biophoton imaging; Ca2+; Phosphodiesterase 6; Rat and bullfrog retinas; Retinal dark noise.

Fig. 1

Imaging biophotonic activity in rat and bullfrog retinas. A Representative regular image of a whole rat retina. B–G Representative biophoton gray images (B-D) and corresponding biophoton number images (E–G) at 34, 36, and 38 °C. H Representative regular image of a whole bullfrog retina. I–N Representative biophoton gray images (I–K) and corresponding biophoton number images (L–N) at 34, 36, and 38 °C. Each image in B–G and I–N was obtained from the merger of 90 continuously-processed original gray images or biophoton number images (1-min imaging time for each original image). Temperatures (34, 36 and 38 °C) indicated in B, C, and D are the same for E, I, and L; F, J, and M; and G, K, and N, respectively. Scale bars 1 mm for A and H.

Fig. 2

Temperature-dependent biophotonic activity in rat and bullfrog retinas. A–G Dynamic changes in biophotonic activity in a representative rat retina at 34, 36, and 38 °C shown as relative gray value (RGV) (A), biophoton number image (BNI) (C), and real number of biophotons (RNB) (E), as well as the sum of the time course of the average change of RGV (B), BNI (D), and RNB (F) (n = 5). H–N Similar patterns in bullfrog retina (n = 6). G, N Significant temperature-dependent increases in biophotonic activity in rat and bullfrog retinas from comparison of average change of RNB from 90 continuously-processed biophoton images at 34, 36, and 38 °C. O Schematic showing the method of estimation of RNBs in the whole retina based on BNIs, considering the sample as a point light source. H Distance between sample and lens (7.0 cm); R, radius of lens (2.5 cm); EMCCD, electron-multiplying CCD camera; the detection quantum efficiency of the EMCCD was 75%.

Fig. 3

Effects of Ca²⁺ and PDE on biophotonic activity in rat retina. A–D Removing intra- and extracellular Ca²⁺ together (A and B, 10 µmol/L BAPTA-AM + 0.5 mmol/L EGTA, n = 5) or introducing a PDE inhibitor (C and D, 100 nmol/L Zaprinast, n = 6) almost completely blocked biophotonic activity in the retina at 34, 36, and 38 °C. The duration at each temperature was 0–90 min at 34 °C, 91–180 min at 36 °C, and 181–270 min at 38 °C. The control data (n = 5) from Fig. 2 B and F were used for statistical comparison. Data show the mean ± SEM. n = number of retinas. *P < 0.05; **P < 0.01.