The main question: How is the topographical information in the olfactory bulb transmitted to and interpreted in the brain to decode the odor map?

Zone-to-Zone Projection (receptor identity)

Zone-to-Zone Projection (zone specific markers + olfactory receptor subtypes) Class I receptor Class II receptor (O-MAC) (OCAM) coronal slice Markers: O-MAC- olfactory specific medium-chain acyl-CoA synthetase (expressed only in the D-zone). OCAM (NCAM2) - olfactory cell adhesion molecule (expressed only in the V-zone).

ΔII ΔDΔD Synaptotagmin GFP OCAM wt Receptor expression Two strains of mice (specific expression of diphtera toxin)

ΔD mice ΔDΔD Synaptotagmin GFP OCAM

Morphological consequences of ΔD (1) (Arrangement of glomeruli in the olfactory bulb) Appropriate glomeruli are missing in ΔD mice (“empty” spaces left – competition?)

Morphological consequences of ΔD (2) (otherwise normal cytoarchitecture with distinct layers) - Appropriate layers were formed (except for missing glomeruli) TH+ = PGNs GABA+ = GCs Synaptotagmin = synapses Reelin+ = MCs - Mitral cells in the D domain do not form dendritic terminal tufts (probably no input from OSNs).

Functional consequences of ΔD (Zif268 + ISI) Dorsal surface of the OB (ISI): “unrolled” whole OB (Zif268): ISI= intrinsic signal imaging Zif268= immediate early gene - No ISI odor response in the dorsal surface of the OB. - Odorants which evoke a response in the V and D domains only activate the V domain in ΔD mice.

Behavioral consequences of ΔD (1) (odor detection) - The detection thresholds for pentanal and TMT were not affected, whereas it was ten- times higher for 2MB acid. - In rats, the most responsive glomerulus for pentanal is located in the V-domain. The same seems to be true in mice, for pentanal and for TMT. Habituation-dishabituation test: Pentanal - spoiled food (avoidance response) 2MB acid - spoiled food (avoidance response) TMT - fox urine (fear avoidance response) Thus, ΔD mice can detect odors fairly normally

Behavioral consequences of ΔD (2) (odor discrimination) Thus, ΔD can discriminate between odors normally - In the odor discrimination test, hungry mice associate 1 odorant with sugar thus and dig longer when smelling it. - Both wt and ΔD have similar discrimination capabilities. -Wt mice cannot be trained to associate TMT with sugar (fear avoidance). - ΔD do not have this problem (but they do detect it).

Behavioral consequences of ΔD (3) (Innate avoidance) Aversive odors - Innate preference test: in contrast to wt mice, ΔD mice do not display innate avoidance. They are even attracted in some cases. - Innate avoidance test: in contrast to wt mice, ΔD mice do not display innate avoidance. They are even attracted in some cases. The same is seen in increasing concentrations of 2MB acid. Thus, the D domain is necessary for innate avoidance

So… ΔD mice can detect and discriminate between odors, but they do not exhibit innate avoidance. Can they be trained?

Behavioral consequences of ΔD (3) (Learned avoidance) - Conditioned avoidance (preference test): ΔD mice can be conditioned to avoid aversive odors (LiCl-induced nausea). Thus, the ΔD mice do not exhibit innate avoidance BUT they do exhibit learned avoidance

The D domain is necessary for innate avoidance, but is it sufficient ? ΔII mice

(Do not have any glomeruli except the DI glomeruli)

(DII but not DI responds to TMT so it is not avoided) Behavioral consequences of ΔII (1) (Innate avoidance) - Innate avoidance test: in contrast to ΔD mice, ΔII mice do display innate avoidance just like wt mice (except in response to TMT). Thus, the D domain is necessary and sufficient for innate avoidance

Two different “fear pathways”: nature and nurture BST - In wt mice, the BST was strongly activated in the BST-MA, and moderately in the BST-LD. - This is consistent with the previous observation that TMT activates the BST- MA, leading to the stimulation of the HPA axis (the “stress pathway”) in rats. - In contrast, the BST-MA was not activated by TMT in the ΔD mice, although the BST-LD was activated as in wild-type mice. - 2MB acid response was similar between wt and ΔD mice.

Two different “fear pathways”: nature and nurture - TMT strongly activated the HPA axis (measured by ACTH) in wt mice but not in ΔD mice. - 2MB acid also moderately activated the HPA axis. “The Stress Pathway”:

Two different “fear pathways”: nature and nurture The proposed model: - TMT activates two different neuronal pathways: one for the innate fear response (in the D-domain) and the other for the learned fear response (in the V-domain). - For TMT, the D-domain glomeruli activate the olfactory cortex, and subsequently the BST-MA, which activates the HPA axis and causes an increase the plasma ACTH concentration.

Eliminate a specific population of neurons Detect morphological consequences Detect functional consequences Detect behavioral consequences Show opposite behavioral consequences Eliminate all but the specific population of neurons “Guidelines” for discovering what a neural circuit “does”: and to be even more persuasive, establish sufficiency (in addition to necessity):

Question?