1. Bi / CNS 150 Lecture 18 Wednesday, November 5, 2015
Olfaction: vertebrates / Worms / insects
Henry Lester
Reading: Kandel Chapter 32, pp (not taste)

2. 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.

3. Part of the Olfactory System
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.
Figure 32-1

4. 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
A single odorant receptor can respond to a number of different compounds
Nonetheless, there can be a high degree of selectivity in odorant binding
Left, chemical structure of carvone
Carvone is an interesting odorant molecule that has a stereo center and, thus, isomers
There are two enantiomers (mirror images) of this compound
One smells like spearmint and the other, caraway
Thus, there appears to be two different odorant receptors for this compound
The different smells evoked by these isomers suggests that odorant receptors have highly selective binding sites for certain odorant molecules

5. 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.

6. Cells of the mammalian main olfactory epithelium
To olfactory bulb
Axon
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.
Basal
cells
Olfactory
sensory
neuron
Dendrite
Supporting
cells
Cilia
Mucus
Like Figure 32-2

7. 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.

8. a a b g The start of the G protein pathway in vertebrate olfaction
Part of
Fig. 32-3
The start of the G protein pathway in
vertebrate olfaction
How fast?
100 ms to 10 s
How far?
Probably less 1 mm
Effector:
membrane-bound
enzyme
activates
G protein
Odorant binds to receptor
outside a b g a
inside
GTP
GDP + Pi

9. The usual GPCR pathway membrane receptor t s q i G protein enzyme
from Lecture 12
receptor t s q i
G protein
enzyme
channel
effector
kinase
phosphorylated
protein
cAMP
Ca2+
intracellular
messenger
cytosol
The usual GPCR pathway

10. (olfactory system, retina)
intracellular
messenger
Ca2+
cAMP
cGMP
but in a previous lecture, we said . . .
Intracellular messengers bind to proteins
kinases
A few ion channels
(olfactory system, retina)
phosphorylated
protein
Previous lecture
Ca and

11. The GPCR pathway in an olfactory cell receptor G protein i q olf t
effector
channel
enzyme
Very similar to Gs
intracellular
messenger
Ca2+
cAMP
cGMP
The GPCR pathway in an
olfactory cell
channel

12. Olfactory neurons have cAMP-activated Na+/Ca2+ Channels
receptor q i
G protein s t
effector
channel
enzyme
intracellular
messenger
Ca2+
cAMP
cGMP
Excised
“inside-out” patch
allows access
to the inside surface of the membrane
+cAMP
closed
open
channel
olfactory neurons have paralogs: cAMP-activated channels
no channel openings
no cAMP

13. 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.

14. Olfactory
bulb
Expression zones of 4 individual olfactory receptors (rat nose, coronal section)
Olfactory
epithelium
Olfactory
receptor
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.
K20
K20
L45
A16
Another gene class, expressed in all olfactory neurons
Figure 32-5

15. 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.
Projections to the olfactory bulb
glomus, ball of yarn (Latin)
like a bishop’s miter (hat)
To lateral olfactory tract
Inhibitory
Mitral cell
Tufted
cell
Periglomerular cell
perforated (Latin)
Figures 32-1, 32-6
Olfactory sensory neuron

16. 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.

17. 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.
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.

18. 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.

19. 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.

20. 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.
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).
Figure 32-9

21. The GPCR pathway in a VNO cell
receptor
G protein i q s t
effector
channel
enzyme
intracellular
messenger
Ca2+
cAMP
cGMP
IP3
DAG
channel

22. VNO signal transduction
(like the GPCR lecture)
VNO signal transduction
phosphatidyl inositol
4,5 bisphosphate = PI(4,5)P2
TRPC2 channel
Like Alberts 15-36
© Garland

23. 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

24. 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.

25. 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

26. Loss of VNO signaling eliminates aggressive responses to intruders
TRP2 = TRPC2
TRPC2 knockout mice
lack this response.
Normal male mice attack intruders introduced into their territory, especially intruders swabbed with male pheromone.
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

27. 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).

28. 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

29. 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

30. 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.

31. 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. “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

33. End of Lecture 18