1. 1 Bi / CNS 150 Lecture 16 Wednesday, November 5, 2014 Olfaction Bruce Cohen Reading: Kandel Chapter 32, pp 712-726 (not taste) Lulu

2. 2 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 A smell is categorized based on one’s previous experiences of it and on the other sensory stimuli correlated with its appearance.

3. 3 Olfactory system can distinguish stereoisomers of a compound The nose can distinguish similar compounds, such as chemical isomers, 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

4. 4 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.

5. 5 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 Olfactory sensory neuron Dendrite Cilia Basal cells Supporting cells Mucus To olfactory bulb Figure 32-2

6. 6 Olfactory receptor proteins in vertebrates and most other phyla (except insects which use ligand-gated channels) Odorants bind to 7-helix (G-protein coupled) receptors. In mice, >1000 genes (2-3% of genes!) encode these receptors. Humans have about 350 odorant receptors Receptor sequences also are quite variable, especially in putative odorant-binding helices. Thus, the repertoire of receptors is extremely diverse. In mammals, each neuron probably expresses only a single receptor. Surprisingly, odorant receptors were recently found in human skin cells, and are also expressed in liver, heart, kidney, and even sperm cells

7. 7 Odorant binding and signal transduction occurs in the cilia (top left) Amino-acid sequences of odorant receptors are highly variable (black dots indicate most variable residues) (bottom left) Odorant binding to GPCR triggers a cascade that opens a cAMP-gated cation channel (right)

8. 8 Olfactory neurons have cAMP-activated Na + /Ca 2+ Channels Excised “inside-out” patch allows access to the inside surface of the membrane no cAMP no channel openings +cAMP closed open receptor qi G protein channel ts enzymechannel effector intracellular messenger Ca 2+ cAMP cGMP

9. 9 More about olfactory channels and their role in olfactory transduction Olfactory cAMP-gated channels are permeable to Na + and Ca 2+ Thus, odorant binding causes depolarization of the olfactory neuron through Na + entry. Ca 2+ also enters and activates a Cl - channel, increasing depolarization (E Cl is near zero in these cells). This process stimulates the olfactory neuron to fire action potentials.

10. 10 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. There are ~1000 receptors within a zone. Neurons within each expression zone send axons to a different quadrant of the olfactory bulb. Another gene class, expressed in all olfactory neurons Figure 32-5 K20 K21 L45 A16 Olfactory receptor Olfactory epithelium Olfactory bulb

11. 11 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 Tufted cell Inhibitory Figures 32-1, 32-6 To lateral olfactory tract like a bishop’s miter (hat) perforated (Latin) glomus, ball of yarn (Latin)

12. 12 Each glomeruli receives inputs from sensory neurons expressing the same odorant receptor 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.

13. 13 Mapping glomerular odorant responses: Ca 2+ 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.

14. 14 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.

15. 15 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. Neurons in the apical layer express the G protein α subunit G αi2, while those in the basal layer express G αo. The transduction channel and the receptors are located on the microvilli at the edge of the lumen.

16. 16 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 odorant 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).

17. 17 The GPCR pathway in a VNO cell resembles the G q pathway channel receptor t s qi G protein enzymechannel effector intracellular messenger Ca 2+ cAMP cGMP IP 3 DAG

18. 18 VNO signal transduction Like Alberts 15-36 © Garland phosphatidyl inositol 4,5 bisphosphate = PI(4,5)P 2 TRPC2 channel (like the GPCR lecture)

19. 19 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

20. 20 Acessory olfactory bulb (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.

21. 21 Odorant perception by the vomeronasal system is largely unconscious 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

22. 22 Chemical composition 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).

23. 23 End of Lecture 16