1. BBE/CNS 150 Lecture 13 Wednesday, October 29, 2014 Vision 1: Phototransduction and the Retina Bruce Cohen Kandel Chapter 26 1

2. Rod Cone Synapses of outer plexiform layer Organization of the retina Glutamate is the major transmitter Some neurons make dopamine & acetylcholine (amacrine cells) Inhibitory neurons release GABA.. Ganglion cell is unique in firing impulses optic nerve Synapses of inner plexiform layer Bipolar cells Horizontal cells 2 Like Fig. 26-2

3. Photoreceptor organs have evolved independently at least 40 times, each time responding to the visible spectrum and near-UV. How do we explain the use of a limited part of the spectrum? Infrared light is not sufficiently energetic to drive photochemical reactions such as the cis-trans isomerization of retinal. Shorter-wavelength ultraviolet light is too energetic and would destroy organic molecules. Photoreception 3

4. h 4 h Free-floating discs Rhodopsin Outer segment of photoreceptors contains the visual pigment Like Figs. 26-5, 26-7

5. Each opsin interacts distinctly with retinal, producing a distinct absorption spectrum. There are 3 types of opsin in human color-sensitive cones. Absorption spectra of cone pigments Blue- green- red- absorbing 5 Mutations that change the spectrum Like Fig. 26-8, 26-9

6. Rods dominate the visual response at low light levels 6

7. Detection of light by retinal bound to opsin From Darnell et al., Mol. Cell Biology 7 Enzymes Like Fig. 26-8

8. The GPCR pathway in photoreceptors channel receptor tsqi G protein enzymechannel effector cytosol intracellular messenger Ca 2+ cAMP cGMP 8 membrane

9. 9 Phototransduction Starts with photon absorption by rhodopsin Transducin binds to activated rhodopsin, exchanges GTP for GDP Activated transducin dissociates into  and  subunits The  subunit binds to, and activates, phosphodiesterase Intracellular cGMP concentration decreases Reduction in cGMP closes cGMP-gated cation channels in the plasma membrane Membrane potential hyperpolarizes Closing of cGMP-gated channel reduces intracellular calcium Reduced calcium counteracts the effects of light absorption

10. Rods and Cones have cGMP-activated Na + /Ca 2+ Channels Excised “inside-out” patch allows access to the inside surface of the membrane no cGMP no channel openings +cGMP* closed open like a previous Lecture receptor qi G protein channel ts enzymechannel effector intracellular messenger Ca 2+ cAMP cGMP 10

11. The “ribbon synapse” facilitates the tonic high rate of transmitter release Photoreceptor to horizontal cell synapse 11

12. The Phototransduction Cascade: 1. Amplification2. Adaptation 1. When fully dark-adapted, many species can detect ~1 photon per photoreceptor cell 2. When fully light-adapted, many species can accurately analyze light at intensities ~10 10 fold brighter 12

13. The Phototransduction Cascade: 1. Amplification 1a. When the rod is dark adapted, the activated Receptor (O*) can activate 500 transducin proteins. 1b. The phosphodiesterase has a turnover number of 4200/sec, near the diffusion limit for catalysis. 1c. Each millisecond that the cGMP-dependent cation channel in the rod outer segment plasma membrane is open,10,000 ions flow through it. 13

14. 2a. Transducin hydrolyses GTP to GDP and thus inactivates itself. The Phototransduction Cascade: (1. Amplification)2. Adaptive mechanisms 3c. Guanylate cyclase must synthesize new cGMP from GTP (1) Guanylate cyclase is partially inhibited by [Ca 2+ ] > ~75 nM. (2) Ca 2+ influx through the tonically open cation channel sets the cytosolic level of Ca 2+ to ~ 500 nM. (3) When the cation channel closes upon light stimulation, Ca 2+ continues to be pumped out via the usual processes, lowering cytosolic Ca 2+ to ~50 nM and activating guanylate cyclase 14 2b. The activated receptor (O* or R*) must also be deactivated. (1) Rhodopsin kinase phosphorylates the carboxyl tail of the receptor (2) The phosphorylation permits binding of the inhibitory protein, arrestin

15. Dark State Light Channel Closure Cyclic GMP hydrolysis Lowered cytosolic Ca 2+ Increased cyclic GMP synthesis Channel opening Visual excitation is followed by Recovery and Adaptation The role of Ca 2+ in adaptation also appears to be important, but this process is not understood in molecular detail yet. 15

16. 16 Information flow in the retina Visual information flows from photoreceptors to ganglion cells through the bipolar neurons There are two types of bipolar cells, “on” and “off” On bipolar cells are depolarized by light at the center of their receptive field Off bipolar cells are hyperpolarized by light at the center of their field Off bipolars have glutamate-gated ion channels that shut off when the photoreceptor stops releasing glutamate On bipolars have glutamate GPCRs that activate phosphodiesterase and inhibit cGMP- gated channels from opening in the dark When the photoreceptor stops releasing glutamate, the cGMP channels open and depolarize the on bipolar cell

17. 17 Retinal ganglion cells have center- surround receptive fields

18. 18 BBE/CNS 150 End of Lecture 13