Sensory/ Motor Systems March 10, 2008 · 12 • Odorants bind to Olfactory Receptors (ORs) in the...

Sensory/ Motor Systems

March 10, 2008

Greg Suh

greg.suh@med.nyu.edu

Title: Chemical Senses- periphery

2

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

12

• 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?

2

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