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Transcript of 4. Connections to the brain 3. retina 1. lens and optics 2. photoreception event 5. A master switch...
4. Connections to the brain
3. retina
1. lens andoptics 2. photoreception
event5. A master switch
that controls differentiation
6. Time frame for evolution of the major features
BBE/CNS 150 Lecture 14 Wednesday, October 30, 2013Vision 1: Phototransduction and the RetinaandEvolution of the Eye Henry LesterChapter 26, co-written by Markus Meister 1
4. Connections to the brain
3. retina
1. lens andoptics 2. photoreception
event5. A master switch
that controls differentiation
6. Time frame for evolution of the major features
2
BBE/CNS 150 Lecture 14 Wednesday, October 30, 2013Vision 1: Phototransduction and the RetinaandEvolution of the Eye Henry LesterChapter 26, co-written by Markus Meister
“Nothing in biology makes sense except in the light of evolution”Theodosius Dobzhansky
All modern biological processes evolved from related processes.
Every modern gene evolved from other genes.
Every gene has an ortholog in related species, and
most genes have paralogs in the same species.
Because all vertebrate eyes are quite similar,
the hunt for orthologs is straightforward and successful in most cases.
That two organisms share many orthologs is powerful evidence for the view
that those organisms are descended from a common ancestor—a central
aspect of evolution.3
Myr BP
Hemoglobin paralogs in the human genome
Myr BP
Hemoglobin paralogs in the mouse genome
chromosome 7
orthologs resemble each otheracross species (mouse vs human )
human vs mouse vs
paralogs resemble each other, distant or closely, within a species
G vs A
Example: globin genes
orthologs & paralogs
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The lens has an index of refraction greater than water,
because it contains a high concentration of protein.
Many proteins different serve this purpose have been used in various animals.
Some of these proteins, termed crystallins, are also enzymes
that perform metabolic functions in other tissues.
Apparently the only requirement is that the protein have good solubility and no
attached groups (such as vitamins) that might absorb light.
1. Lens and optics
from Lecture 1 How much is 4 mM protein?
A typical protein has 500 amino acid residues.
An average residue has a molecular mass of 110.
Therefore the average protein has a molecular mass of 55,000.
( 4 x 10-3 mol/liter) x (5.5 x 104 g/mol) = 2.2 x 102 g/l = 220 g/l.
The cell is ~22% protein!
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Pax-6 orthologs occur in phyla as diverse as as mammals, insects, and molluscs.
Many genes, including crystallins, have acquired a “Pax-6 responsive element”Pax-6 contains a homeo domain & another-DNA binding domain
Crystallin
Pax-6, a transcription factor with orthologs in many species
Pax-6 (vertebrates)Ey (Drosophila) Presence of multiple
sufficient gene regulatory mechanisms can underlie
“gene sharing”
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Existing proteins have been used for an additional functions. Which way were they adopted? Probably the use in the lens came second. Evidently several distinct transcription factors can “share” activation of a given gene.
The aperture mechanism: controlled by smooth muscles
singlesmooth muscle cell
inextensible fibers
Contracts and thickens: leads to smaller pupil
nerve from brain; muscarinic synapse
Innervated smooth muscles control:
diameter of blood vessels, peristaltic activity of the intestinal tract,
diameter of the bladder neck
In each case, the nervous system has evolved circuits that
(1) extract and integrate information from sensors and
(2) employ smooth muscles in a homeostatic loop.
blocker: atropine from Atropa belladonna
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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 provoke photochemistry such
as cis-trans isomerization.
Shorter-wavelength ultraviolet light is too energetic and would destroy
organic molecules.
2. The photoreception event
8
h
9
h
Free-floating discs
Rhodopsin
Rhodopsin
The photoreceptor cells receive light from “the back”
Like Figs. 26-5, 26-7
Each opsin interacts distinctly with retinal, producing a distinct absorption spectrum.
There are 4 opsin paralogs in the human genome.
Absorption spectra of cone pigments
Blue- green- red-absorbing
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Mutations that change the spectrum
Like Fig. 26-8, 26-9
Detection of light by retinal bound to opsin
From Darnell et al., Mol. Cell Biology11
Enzymes
Like Fig. 26-8
cytosol
The usual GPCR pathway
nucleus
kinase
phosphorylatedprotein
cAMPCa2+
intracellularmessenger
receptor
tsqiG protein
enzymechannel effector
membrane
from Lecture 12outside
inside
outside
inside
12
The GPCR pathway in a photoreceptor
channel
receptor
tsqiG protein
enzymechannel effector
cytosol
intracellularmessenger
Ca2+ cAMPcGMP
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membrane
GTP GDP + Pi
Effector: enzyme or channel
outside
inside
Neurotransmitter or hormonebinds to receptor
activatesG protein
Beginning of the G Protein-Coupled Receptor Pathway
How fast?100 ms to 10 s
How far?Probably less 1 m
like previous lectures
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Photon isomerizesretinal bound to rhodopsin
Special aspects of the G protein-coupled receptor pathway in photoreception
How fast?< 100 ms
How far?< 1 m
GTP
activatesG protein
Effector is an enzyme
GDP + Pi
h
Although the components are not membrane-bound, the membranes effectively
restrict their motion
In rods and cones, these proteins lack lipid tails
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cytosol between disks,or
between folds
Like Fig. 26-7
Intracellular messengers bind to proteins
kinases
phosphorylatedprotein
A few ion channels(olfactory system,
retina)
N
NN
N
NH2
O
OHO
HH
O
P-O
O
cyclic AMP (cAMP)
Ca2+ and
Expanding on a previous lecture, we said . . .intracellularmessenger
Ca2+ cAMPcGMP
Cyclic nucleotide(cAMP or cGMP)
16
Cyclic GMP is the second messenger for phototransduction
High cyclic GMP keeps the plasma membrane
depolarized and keeps glutamate release at the
terminal high.
Increased Hydrolysis of cGMP reduces cGMP concentration, resulting in closing of a cation channel in the outer segment
membrane and transient hyperpolarization of the entire plasma
membrane.
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cAMP ATP
Effector enzyme“cyclase”
Breakdown enzyme“phosphodiesterase”
Inhibited by caffeine uninteresting
cGMP GTP
Enzyme“cyclase”
Breakdown enzyme“phosphodiesterase”
uninterestingThe effector for Gt
like a previous Lecture
A paralog expressed elsewhere in the body is
inhibited by Viagra
channel
receptor
tsqiG protein
enzymechannel effector
intracellularmessenger
Ca2+ cAMPcGMP
“Viagra . . . may cause a perception of bluish haze or increased light sensitivity in
some patients.”
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Rods and Cones have cGMP-activated Na+/Ca2+ Channels
Excised “inside-out” patch
allows access to the inside surface
of the membrane
no cGMP no channel openings
+cGMP*
+cGMP*
closed
open
like a previous Lecture receptor
qiG protein
channel
ts
enzymechannel effector
intracellularmessenger
Ca2+ cAMPcGMP
19
Light Response of the Photoreceptor Cell
The vertebrate photoreceptor functions electrophysiologically opposite to most neurons.
20
1. Rhodopsin absorbs light
2. Cation channels close in the plasma membrane of the outer segment, which hyperpolarizes the entire cell
. 3. The hyperpolarization relays visual information to the synaptic terminal,
where it slows ongoing release of the transmitter glutamate.
The “ribbon synapse” facilitates the tonic high rate of transmitter release
Photoreceptor to horizontal cell synapse
21
The Phototransduction Cascade:1. Amplification 2. Adaptative/homeostatic mechanisms
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 ~1010 fold brighter
Many adaptive and homeostatic mechanisms underlie these phenomena.
Note: it is incorrect to explain that your favorite process (memory, learning, addiction) occurs “because of” homeostasis or adaptation.
Homeostasis and adaptation are not, by themselves, mechanisms.
There are homeostatic and adaptive mechanisms.
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The Phototransduction Cascade:1. Amplification (2. Adaptive/homeostasic mechanisms)
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.
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2a. Transducin hydrolyses GTP to GDP and thus inactivates itself.
The Phototransduction Cascade:(1. Amplification) 2. Adaptive/homeostatic mechanisms
3c. Guanylate cyclase must synthesize new cGMP from GTP(1) Guanylate cyclase is partially inhibited by [Ca2+] > ~75 nM.(2) Ca2+ influx through the tonically open cation channel sets the cytosolic level of Ca2+ to ~ 500 nM.(3) When the cation channel closes upon light stimulation, Ca2+ continues to be pumped out via the usual processes, lowering cytosolic Ca2+ to ~50 nM and activating guanylate cyclase
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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
Rod
Cone
Synapses of outer plexiform layer
3. Neurons of the retina
Glutamate is the major transmitter; Some neurons make
dopamine & acetylcholine.Inhibitory neurons release GABA.
Many paralogs to genes expressed elsewhere:
Channels, receptors, transporters.
Ganglion cell is unique in firing impulses
optic nerve
Synapses of inner plexiform layer
Bipolar cells
Horizontal cells
25
Like Fig. 26-2
Roger Sperry’s Nobel prize-winning experiments (1948) (goldfish): After he cut the optic nerve, individual fibers grew back to their original destination in
the brain.
Sperry also conducted the “Split brain” experiments that form the basis for
modern ideas about the distinct specialties of the two hemispheres.
4. Connections to the brain
Sperry postulated a “chemoaffinity” between
the nerves and their target cells.
A previous Lecture
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Horseshoe crabs(Limulus polyphemus)
Maps may be unique to nervous systems,
but
visual maps arose at least 500 Myr ago.
We will discuss visual maps in the next
lectures.
27
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tyrosine kinase
receptors
peptide ligands for
these receptors
Ephrins:cell-surface proteins that can induce growth cone collapse.
Eph kinases and Ephrins are distributed in gradients in the retina and tectum.
Eph repulsive signaling partially defines Sperry’s “chemoaffinity” that sets up the retinotectal map.
Axons with high Eph kinase expression avoid tectal regions with high
levels of ephrin
Figs 54-13, 54-14
Sperry’s “chemoaffinity”in the retinotectal system:
a 21st Century view
A Normal
A P A PRetina Tectum
Cell bodies Growth cones
A P A P
C Inactivate Ephrin A5
B Confined overexpression of Ephrin A2
A P A P
Discussed in a previous lecture
Pax-6 / Ey functions when expressed at various locations in Drosophila
5. Master switches for eye development?
Little Alberts 8-25 © Garland 29
Eye formation varies enormously among
organisms,
yet even a human Pax-6 ortholog induces an eye in
Ey mutant Drosophila!
30
A Pessimistic Estimate Of The Time Required For An Eye To Evolve, D.-E. Nilsson and S. Pelger,
Proceedings of the Royal Society London B, 1994, 256, pp. 53-58.
Estimate: several hundred thousand yr from primitive eyespot to fisheye with lens
Selective advantages of the intermediate steps are summarized here:http://www.pbs.org/wgbh/evolution/library/01/1/l_011_01.html
1 2 3 4
5 86 7
6. Time frame for evolution of the major structural features
31
32
Henry Lester will not have “office” hours this Friday
BBE/CNS 150 End of Lecture 14
Dark State
Light Channel Closure
Cyclic GMP hydrolysis
Lowered cytosolic
Ca2+
Increased cyclic GMP synthesis
Channel opening
Visual excitation is followed by Recovery and Adaptation
The role of Ca2+ in adaptation also appears to be important, but this process is not understood in molecular detail yet.
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