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Bi / CNS 150 Lecture 18 Wednesday, November 5, 2015

Olfaction: vertebrates / Worms / insects

Henry Lester

Reading: Kandel Chapter 32, pp 712-726 (not taste)

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Proust, Remembrance of Things Past

“as soon as I had recognized the taste of the piece of madeleine soaked in her decoction of lime-blossom which my aunt used to give me (although I did not yet know and must long postpone the discovery of why this memory made me so happy) immediately the old grey house upon the street, where her room was, rose up like a stage set to attach itself to the little pavilion opening on to the garden which had been built out behind it for my parents (the isolated segment which until that moment had been all that I could see); and with the house the town, from morning to night and in all weathers, the Square where I used to be sent before lunch, the streets along which I used to run errands, the country roads we took when the weather was fine . . . “

Olfactory memory

The nose can detect and (in principle) classify thousands of different compounds.

The ‘mapping’ of these compounds probably occurs by matching to memory templates stored in the brain; thus, a smell is categorized based on one’s previous experiences of it and on the other sensory stimuli that correlate with its appearance.

Part of the Olfactory System

Figure 32-1

Odorants are volatile chemicals that can be detected by olfactory sensory neurons in the nose

Odorants can evoke an emotional response that is more immediate and compelling than the response to visual or auditory stimuli

Outputs from the olfactory bulb go directly to the cortex without passing through the thalamus

Part of the reason that olfactory stimuli may be able to evoke a strong emotional response is that the olfactory bulb projects to areas of the limbic system that mediate emotional and motivational responses

These areas are the amygdala and anterior hypothalamus.

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Olfactory system can distinguish stereoisomers of a compound

•A single odorant receptor can respond to a number of different compounds

•Nonetheless, there can be a high degree of selectivity in odorant binding

•The nose can distinguish similar compounds, such as stereoisomers, as different smells.

•An example: the two stereoisomers of carvone smell like spearmint (R) and caraway (S).

•This implies that there are stereoisomer-specific carvone receptors.

•Also implies that odorant receptors are proteins

Carvone

Stereo center

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Anatomy of the mammalian olfactory system

In many mammals (rodent shown here), the

olfactory organs within the nose are split

into the main olfactory epithelium (MOE)

and the vomeronasal organ (VNO).

MOE neurons project to the main olfactory

bulb (MOB).

VNO neurons project to the accessory

olfactory bulb (AOB).

MOB output neurons project to regions of

cortex, while AOB output neurons project

only to the (ventral) amygdala.

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Cells of the mammalian main olfactory epithelium

Olfactory neurons have

apical dendrites with long

ciliary extensions, where

the transduction

components are located.

Cilia are embedded in the

mucus layer.

Olfactory neurons turn over

and are replaced every 60

days.

Axon

Olfactorysensory neuron

Dendrite

Cilia

Basalcells

Supportingcells

Mucus

To olfactory bulb

Like Figure 32-2

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Olfactory receptor proteins in vertebrates and most other phyla

Odorants bind to 7-helix (G-protein coupled) receptors.

In mice, >1000 genes (2-3% of genes!) encode these receptors.

Humans have ~ 350 odorant receptors.

Receptor sequences also are quite variable, especially in putative odorant-binding helices.

Thus, the repertoire is extremely diverse.

In mammals, each neuron probably expresses only a single receptor.

8GTP GDP + Pi

Effector: membrane-bound

enzymeoutside

Odorant binds to receptor

a

activatesG protein

The start of the G protein pathway in vertebrate olfaction

How fast?100 ms to 10 s

How far?Probably less 1 mm

b g

a

inside

Part of Fig. 32-3

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cytosol

The usual GPCR pathwaykinase

phosphorylatedprotein

cAMPCa2+

intracellularmessenger

receptor

tsqiG protein

enzymechannel effector

membrane

from Lecture 12

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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

but in a previous lecture, we said . . .intracellularmessenger

Ca2+ cAMPcGMP

Previous lecture

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The GPCR pathway in anolfactory cell

channel

receptor

tolfqiG protein

enzymechannel effector

intracellularmessenger

Ca2+ cAMPcGMP

Very similar to Gs

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Olfactory neurons have cAMP-activated Na+/Ca2+ Channels

Excised “inside-out” patch allows access to the inside surface of the membrane

no cAMP no channel openings

+cAMP

+cAMP

closed

open

receptor

qiG protein

channel

ts

enzymechannel effector

intracellularmessenger

Ca2+ cAMPcGMP

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More about olfactory channels and their role in olfactory transduction

Olfactory cAMP-gated channels are permeable to Na+ and Ca2+

Thus, odorant binding causes depolarization of the olfactory neuron through

Na+ entry.

Ca2+ also enters and activates a Cl - channel, increasing depolarization (ECl

is near zero in these cells).

This process stimulates the olfactory neuron to fire action potentials.

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Expression zones of 4 individual olfactory receptors(rat nose, coronal section)

The olfactory turbinates display four ‘expression zones’.

Each receptor is expressed in a small, randomly

distributed subset of neurons within one of the 4 zones

.

As there are ~1000 receptors, about 1/250 of neurons

within a zone express each receptor.

Neurons within each expression zone send axons to a

different quadrant of the olfactory bulb.

Another gene class, expressed in all olfactory neuronsFigure 32-5

K20

K20

L45

A16

Olfactoryreceptor

Olfactoryepithelium

Olfactorybulb

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Projections to the olfactory

bulb

Olfactory neurons send axons to the glomeruli (synaptic balls shielded by glia) of the olfactory bulb.Olfactory neurons excite mitral cells, which are the bulb output cells.

Olfactory sensory neuron

Mitral cell

Periglomerular cell

Tuftedcell

Inhibitory

Figures 32-1, 32-6

To lateral olfactory tractlike a bishop’s miter (hat)

perforated (Latin)

glomus, ball of yarn (Latin)

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Projections to specific glomeruli

Neurons expressing a specific

olfactory receptor project their

axons to a single glomerulus in

each half-bulb.

Axons converge from many

directions onto the target.

This projection specificity is at

least partly determined by the

receptor itself, but the

mechanisms are unknown.

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Mice: glomeruli connected to neurons expressing I7, a receptor for octanol, respond

to octanol.

This was also the case if the I7 glomerulus was moved to the wrong place in the bulb

by transplacing the I7 gene into the genomic locus for another receptor.

Glomerular odorant responses: Ca2+ imaging in a fish

Individual glomeruli are selectively activated by specific odorants.

In fish, “odorants” are soluble amino acids.

Imaging studies now show that specific glomeruli in mammals are also activated in response to odorants.

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Maps of mitral cell projections to higher olfactory areas

Piriform cortex neurons receive projections from mitral cells corresponding to

many glomeruli that receive input from ORNs expressing different receptors.

Mitral cells also project to olfactory tubercle and other areas.

Integration of odorant responses and odorant identification may take place in

cortex, although some integration is also likely to occur in the bulb.

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The vomeronasal organ

The VNO is thought to respond to pheromones.

It is a cup-shaped organ near the front of the rodent nose; its neurons are divided into basal and apical (near the lumen) layers.

The microvilli of the VNO neurons face the lumen.

The transduction channel and the receptors are located on the microvilli at the edge of the lumen.

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VNO receptor molecules

The 2 distinct families of VNO G protein-coupled receptors are all unrelated to MOE receptors.

Each VNO neuron probably expresses only one receptor, as in the MOE.

Figure 32-9

V2Rs (~100 genes in the rodent) are

expressed in a random pattern by basal layer

neurons (Go-expressing neurons). V2Rs have

large N-terminal extracellular domains.

V1Rs (~180 genes) are expressed by

different subsets of neurons within the

apical layer (Gi-expressing neurons).

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The GPCR pathway in a VNO cell

channel

receptor

tsqiG protein

enzymechannel effector

intracellularmessenger

Ca2+ cAMPcGMPIP3

DAG

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VNO signal transduction

Like Alberts 15-36© Garland

phosphatidyl inositol4,5 bisphosphate = PI(4,5)P2

TRPC2 channel

(like the GPCR lecture)

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Response characteristics of VNO neurons

VNO neurons respond to urine.

Some neurons selectively respond to urine from mice of the same sex,

others to urine of the opposite sex.

Unlike ORNs, their responses are narrowly tuned; no neurons were ever

observed to respond to more than one compound.

A behavioral assay: mice produce ultrasonic calls (‘whistling’) in response

to contact with urine from the opposite sex; production of these calls

requires both the VNO and the MOE.

In TRPC2 knockout mice, VNO neurons do not respond to urine; and mice

do not vocalize in response to urine

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AOB projections to the brain

Mitral cells in the AOB have apical dendrites that arborize in multiple glomeruli.

The AOB projects to the amygdala (directly), and the hypothalamus (via the

amygdala).

The projections from the rostral and caudal AOB halves are superimposed in the

amygdala.

This implies that integration of pheromone signals may take place primarily in the

AOB.

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Generalities about main olfactory system and vomeronasal system function

The main olfactory system mediates cortical responses to volatile odorants,

and these cortical responses are used to drive conscious behavior (food-

seeking, predator avoidance, etc).

The VN system is thought to mediate unconscious responses to water-soluble

pheromone compounds found in urine and secretions of other individuals.

Despite the apparent absence of the vomeronasal organ in humans, we still

apparently detect and respond to some pheromones, including ones that

control the menstrual cycle

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Loss of VNO signaling eliminates aggressive responses to intruders

TRPC2 knockout males mate normally with females.

Remarkably, though, they also mount males, which control mice never do.

The TRPC2 knockout phenotype suggests that the ‘default’ pathway in the absence of VNO input is to mate with everything.

VNO input causes male mice to fight rather than attempt to mate

Normal male mice attack intruders introduced into their territory, especially intruders swabbed with male pheromone.

TRPC2 knockout mice lack this response.

TRP2 = TRPC2

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Chemical nature of pheromones

The various pheromones include

prostaglandins in fish,

androstenone in pigs, and

protein ligands such as hamster aphrodisin.

In most cases, however, individual pure compounds don’t elicit strong responses.

Natural pheromones are mixtures of many substances,

perhaps combinations of (protein carriers) plus (bound small organic compounds).

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A genetic model system: nematode olfaction

C. elegans can chemotax toward and away from volatile attractants and repellents.

It uses only two pairs of neurons, AWA and AWC, to respond to volatile attractants.

It has many olfactory receptors, however, so each chemosensory neuron must express many of these.

The ODR-10 receptor is expressed in AWA and localized to its dendrite.

ODR-10 is a receptor for the odorant diacetyl (2,3 butanedione).

Worms lacking ODR-10 are not attracted to diacetyl.

Figure 32-11

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ODR-10 phenotype

ODR-10 is specific for diacetyl and does not respond to 2,3-pentanedione,

which differs by only one methylene group.

ODR-10 mutants still chemotax to 2,3-pentanedione

The AWC cell has receptors for 2,3-pentanedione

Deletion of AWC destroys chemotaxis to 2,3-pentanedione

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ODR-10 recognizes components of a metabolic pathway

What is the selective advantage of a worm’s response to diacetyl?

Diacetyl results from respiration by certain bacteria.

These bacteria often use citrate as a carbon source.

ODR-10 also recognizes the metabolic intermediates citrate and pyruvate.

Diacetyl is a volatile signature compound for certain bacterial species.

Many other bacteria do not make diacetyl but do make acetoin or lactate as

respiratory endproducts.

Diacetyl attraction thus allows the worm to recognize specific food sources at

a distance in the soil.

Citrate and pyruvate (nonvolatile) interactions with ODR-10 may provide

taste-like recognition of these bacteria after the worm arrives at their colony.

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A Recent Surprise: Insect Olfactory Receptors are Probably Ligand-gated Channels

Encoded by one of ~ 60 genes

An auxiliary subunit, common to most insect olfactory receptors.

Also called Or83b, Or1, Or2, and Or7.

Terminology is converging on “Orco”

For structure of the Drosophila

olfactory system, see Fig. 32-10

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“Metabotropic signalling in vertebrates provides a rich panoply of positive and negative regulation, whereas ionotropic signalling in insects enhances processing speed.” Kaupp, Nature Revs. Neuro, 2010

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End of Lecture 18