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Richard Axel and Linda Buck
Win Nobel Prize for Olfactory System Research
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After Leslie Vosshal
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Pear Pear Banana Orange
Pineapple Peach Apple Lemon
Fruit Odors
After Leslie Vosshal
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Menthol Caraway Spearmint
Stereochemistry and Olfaction
After Leslie Vosshal
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A multitude of vertebrate chemosensory receptor genes
After Leslie Vosshal
MOE
VNO
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Retinotopy
Spatial information
Topographic maps
x
y x’
y’
?
Olfactory projections
T y p e o f r e c e p t o r
y’
x’
8Kandel et al.: Principles of Neural Science Purves et al.: Neuroscience
Our brain is consisted of a series of maps
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Sensory Perception
One receptor per cell…………
Blue
allows Brain discrimination
Red Green Qualitative map?
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System Quality
Visual Position in the external world
Somatosensory Position on the body
Auditory Tone
Olfactory Chemical identity
Space is Used in Sense Organs
and the Brain to Encode Quality
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Poorly understood until
molecular recent
advance Nobel Prize to
Richard Axel & Linda
Buck
Best studied because of
precise stimuli and
sophisticated
physiology &
psychophysics
Brain
processing
Odorant to OR to
G protein to cAMP gated
channel opening; slow
Photon to rhodopsin to
G protein to cGMP gated
channel closure; fast
Signaling
mechanism
Poorly defined; poor
spatial organization
Well defined; extremely
well organized spatially Sensory
stimuli
Instinctive:
This smells right.
Complimentary:
He has a vision.
Literature
use
Olfaction Vision
3
Comparisons of Vision and Olfaction
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• Odorants bind to Olfactory Receptors (ORs) in the dendritic tip of Olfactory Receptor Neurons
• In mouse, there are ~1000 OR genes (~ 3% of genes!)
• Each OR neuron expresses only one type of OR: Exclusion
• ORs activate G protein, adenylate cyclase & cAMP-gated channels: Sensory stimuli are sent as action potentials to the brain
• Olfaction is a combinatorial recognition process: each OR is activated by a number of odorants; each odorant activates a number of ORs (ORNs). A vast number of odorants can be distinguished
How is the olfactory information processed in the brain?
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Olfaction
13from Ramon y Cajal (1911) Histology
A: Olfactory receptor neuron
B: Glomerulus C: Mitral cell
D: Granule cell E: Mitral cell axons sending olfactory
information to the olfactory cortex
Anatomy of the Olfaction System
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from Axel (1995) Sci. Am. 273(4), 154
Anatomy of the Olfaction System
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Axel (1995) Sci. Am. 273(4), 154
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Anatomy of the Olfaction System
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Axel (1995) Sci. Am. 273(4), 154
Three models of information processing
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Vassar et al. (1993) Cell 74:309
Random expression of receptors within one of the four zones:
The ORNs expressing common receptors are NOT grouped
Olfactory receptor expression in nasal epithelia
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Axel (1995) Sci. Am. 273(4), 154
ORNs expressing the same receptor appear to target their axons to the same glomeruli
(usually one receptor, two glomeruli per hemisphere)
In situ hybridization for a
particular OR gene: two bilaterally glomeruli in the
olfactory bulb
ORN axon convergence I:
in situ hybridization of receptor expression
1- what is OR’s function in the OB?
2- why is OR expressed in a single glomeruli?
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Is the model B correct?
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- IRES: translation of the P2 receptor and tau-lacZ
- tau-lacZ: Microtubule binding protein tau and lacZ, for axon labeling
- KI: insertion of a transgene into a defined genomic locus (here the P2 locus):
transgene is expressed in the exact same pattern as the endogenous gene
ORN axon convergence II
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P2 in situ tau-lacZ
All P2 receptor expressing neurons
converge their axons onto the same glomeruli
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Mombaerts et al. (1996) Cell 87, 675
nose
olfactory bulb
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• Each of the 1000 ORs in mouse is expressed
“randomly” in ~1000 ORNs
• The ~1000 ORNs expressing a given OR gene
converge their axons onto ~2 of the ~2000
glomeruli
• The olfactory information is represented as a two
-dimensional map in the olfactory bulb; a given
odor should activate a defined set of olfactory
receptors and a defined set of glomeruli
A glomerular map for odor representation
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• Human Brain: ~1012 neurons, ~1015 synapses
• Human genome: ~3x109 bp, ~3-5 x 104 genes
• There are far few genes (proteins) than needed to make
“cytochemical tags” that specify every connection precisely!
• Make many proteins from a single gene
• Use combinatorial code to specify connections
• Use different amount of the same protein to specify different connections: gradients
• Leave some wiring unspecified — allow experience to determine the final connections
Possible Solutions
Olfactory maps are qualitatively different from visual
/somatosensory maps: digital vs. continuous
The Wiring Specificity Problem
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• Knock-in of IRES-tau-lacZ in P2 receptor
• ORN now express only lacZ
• Axon convergence is disrupted
Thus, OR are required for axon convergence to glomeruli
Roles of olfactory receptors in ORN axon targeting
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Wang et al. (1998) Cell 93, 47
P2-I
RE
S-t
auLacZ
P
2-d
ele
tion-I
RE
S-t
auL
acZ
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• P2 receptor is replaced with a different OR-IRES-tau-lacZ
• ORNs normally expressing P2 now express M50 along with the tau-lacZ marker
• Axon convergence is restored
Where do the axons go? P2? M50?
43 Receptor swap restores ORN axon convergence
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29
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Wang et al. (1998) Cell 93, 47
• M50->P2 swap results in targeting of ORN axons to some intermediate glomerulus! (So did several other swaps)
• Some other as-yet unidentified cues must work together with olfactory receptors to instruct targeting specificity
• The ligands that these ORN axons recognize are completely mysterious
Could post-synaptic target cells play a role?
Olfactory receptors play an instructive although
not deterministic role in ORN axon targeting
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Determination
partially by
connections
Independent
specification
How Are Neural Maps Made?
32or Tbr-/-, P2-IRES-lau-lacZ
Hence, ORN axon convergence appears not to be dependent on their major post-synaptic targets
Post-synaptic target cells are not involved in the map
- In Tbr-/- mice, mitral cells (second
order “projection neurons”)
are not formed.
- In Dlx-1,2 -/- mice, the majority of
GABAergic local interneurons (e.g.
granule cells) are not formed.
- Yet ORN axon convergence are
normal
3333
Determination
partially by
connections
Independent
specification
How Are Neural Maps Made?
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Knockout
Wildtype
Electroolfactograms (EOG) show that response to all odorants is absent in CNG
channel KO mice.
(The residual opposite sign response to triethylamine arises from non-neuronal stimulation)
(Brunet et al., Neuron 17:681 (1996)
Cyclic Nucleotide-Gated Channel KO Mice
are Totally Anosmic
35In the absence of odor-induced electrical activity,
ORN axon targeting is normal
Neuronal activity is not involved in the map
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Antennae
Maxillary Palps
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Antennal
lobe
Mushroom
body Lateral
horn
Antenna
Maxillary palpAntennal lobe
Mushroom body
Lateral horn
The Drosophila Brain
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The Antennal Lobe (fly Olfactory bulb) comprises
~ 43 glomeruli, occupied in stereotypic positions.
D
V A P
Neurons expressing a given OR converge
upon a single pair of glomeruli.
Antenna Antenna lobe
Vosshall et al., 2000
Or47a Or47b
Or22a
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• There are ~50 olfactory receptor genes in Drosophila
(DOR); each DOR is expressed in a stereotypical subset of
olfactory receptor neurons (ORNs, ~ 1300 total) in the third
antennal segment, or in the maxillary palp
• Each ORN expresses only one DOR; the exception to this
rule is one distant member of DOR: OR83b, which is
expressed in 80% of all ORNs. OR83b may serve as a co
-receptor
• ORNs expressing the same receptor converge their axons
onto the same glomerulus (~ 40-50)
• Hence, the “1 receptor, 1-2 glomerulus” rule derived from the mouse study also applies in Drosophila, at least to the
first degree of approximation
Clyne et al. (1999) Neuron 22, 327;
Vosshall et al. (1999) Cell 96, 725;
Vosshall et al. (2000) Cell 102, 147; Gao et al. (2000) Nat. Neurosci. 3, 780
Molecular biology of Drosophila olfaction
4141
Determination
partially by
connections
Independent
specification
How Are Neural Maps Made?
Are the identities of projection (OR target) neurons
instructed by ORNs, or are they independently specified?
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MARCM: Mosaic Analysis with a Repressible
Cell Marker
Developed by Luo and Lee
Dorsal and lateral neuroblast clones contain PNs
with stereotypical & complementary projections.
Greg Jefferis, Lisa Marin
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Projection Neurons Connect the Antennal Lobe to
the Mushroom Body and lateral horn.
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Expression of CD2 in a Population of Projection Neurons
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Expression of CD8-GFP in Single Projection Neurons
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Individual Projection Neurons Innervate a Single Glomerulus
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Different Projection Neurons Show Distinct Patterns in the lateral horn.
Similarities of Projection Patterns
Spatial Map in the Higher Brain
cpEGFP G-CaMP
M
13
CaM
N C
Ca++
509 nm
G-CaMP: A Calcium Sensitive GFP
Nakai et al., 2001
925 nm
The Antennae-Brain Preparation
Glomerular Response to Caproic Acid and Pyridine
Caproic Acid
Pyridine
Odor Representations in the Fly Brain
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Sengupta et al. (1996) Cell 84, 899
Troemel et al. (1997) Cell 91, 161
—Cori Bargmann and colleagues
• Of the 959 cells in C. elegans, 302 are neurons; 3 pairs (AWAs, AWBs, AWCs) are responsible for sensing volatile stimuli (olfaction)
• AWAs and AWCs mediate odor-induced attraction; AWBs mediate odor-induced repulsion (laser ablation experiments)
• ~500 G-protein coupled receptors are expressed in sensory neurons, ~100 of which are likely expressed in olfactory neurons; thus individual neurons express multiple olfactory receptors
• Odr-10 is a receptor for diacetyl, is expressed in AWA neurons, and mediates diacetyl induced attraction
• Misexpress Odr-10 in AWB neurons in Odr-10 mutants resulted in diacetyl induced repulsion!
Thus, a simple logic likely applies to C. elegans olfaction: all “attractive” receptors are expressed in AWAs and AWCs; all “repulsive” receptors are expressed in AWBs. Activation of a particular class of sensory neurons determines the behavioral consequence.
—Space code in the simplest form!
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C. elegans olfaction
Mammalian Taste
