Olfaction 1 Flashcards Preview

Sensational Neuroscience > Olfaction 1 > Flashcards

Flashcards in Olfaction 1 Deck (55):

Billig (2011)

Ano2 highly expressed in MOB / MOE, Ano1 in the vomeronasal organ, but not MOE (western blot). Cre-LoxP (tissue-specific) Ano2 KO: still expressed CNG channel (immunostaining). Patch clamp: KO only had 10% of the Cl- currents that WT had.

R: KO = no change in air-phase olfactograms, - chloride current is not required for physiological levels of olfaction. Functional relevance: go/no-go discrimination: KOs = similar levels of sensitivity to different odourants (CnGa2 KOs showed considerably less sensitivity in task, demonstrating vital importance of CnGa2 in olfactory transduction).

L: NFA (Cl- channel inhibitor) suppressed some of the transduction activity in KO. Another Cl channel in the KO contributing to depolarisation?? OFF-TARGET effects of NFA cannot be ruled out - may be suppressing electrical activity of OSNs by a mechanism independent of Cl- channels. In vitro: IONIC GRADIENTS may not reflect physiological ion gradients generated by in vivo MUCUS.

May be subtle types of discrimination, or other contexts (e.g. lower concentration odorant mixtures), where amplification by the chloride current becomes important.


Kelliher (2003)

CNGA4 KO: go/no-go task: impaired ability to discriminate 10-3M of octanal against a background of 10-4M. Background was reduced to 10-5M, KO mice significantly improved ability to discriminate (90% accuracy).

KO mice had significantly reduced ability to detect octanal on a background of heptanal (they developed cross-adaptation), but were accurately discriminated octanal from a background of cineole.

R: Effect of CNGA4 subunit on adaptation, and CNGA4-mediated adaptation necessary for discriminating odours from background odours, BUT this role is dependent upon the similarity of the odours.

L: KO causes lack of localisation of multiple CNG subunits, to the cilia. BUT KO in this paper still demonstrated olfactory function - if not localised would expect complete lack of ability to discriminate (Billig et al 2011 showed that CNGA2 KOs are unable to discriminate odours). Unclear whether we can trust the findings of this subunit KO, - further experiments with similar methodology (same staining methods, electrophysiology) would be needed to clarify the effects of CNGA2 KOs.


Mombaerts et al (1996)

Cre-LoxP recombination in mice: replace the P2 coding region (OR) with an M12 coding region. Inserted tau-lacZ so that the axonal projections of OSNs could be visualised.

Position of the P2 glomerulus to shifted posteriorly. Suggests that OR type may determine spatial patterning in the MOB. (But when remove transduction with CnGa2 KO - still get same glomerular position, so not dependent on transduction ability?).

However, this cannot be the only determining factor of glomerular positioning, as the glomerulus remained within the same zonal projection, and was nearer to the P2 position than the M12 position.


Imai et al (2006)

T: IRES technology in mice to label a specific (I7) OR with GFP (visualise the associated glomerulus). Changed the DRY sequence of the G protein to ‘RDY’ (shown to eliminate transduction using calcium imaging). Also inserted constitutively active G-protein (caG3).

R: I7(RDY) mutant failed to converge with glomerulus but when caG3 inserted, glomerular convergence restored, and glomerulus shifted posteriorly.

caG3 in WT mouse also caused posterior shift, whereas blocking PKA signalling in WT resulted in an anterior glomerular shift.

Illustrates how altering Gs signalling can determine the anterior-posterior axis of a glomerulus.


Arevian et al (2008)

1) Calcium imaging: more granule cells active when two glomeruli simulatenously activated (rather than individual excitation of each glomerulus) - granule cells are activated by SUMMATION of mitral cell input.

2) Whole cell recordings of pre and post synaptic mitral cells, & evoked lateral IPSPs in the post-synaptic cells, by stimulating nearby mitral cells. Showed activity-dependent lateral inhibition across dendrodendritic reciprocal synapses where lateral inhibition between two neighbouring mitral cells DEPENDS ON ACTIVITY OF CELL BEING INHIBITED

L: in vitro mouse olfactory bulb slices, therefore, it is possible that the process may behave differently in vivo.

3): Computational model of activity-dependent lateral inhibition, based on these electrophysiological findings, where connectivity was ‘all-to-all’, with no spatial structure. Performed contrast enhancement of input patterns, even when the spatial patterning of the input patterns were completely randomised.

Supports the idea that MOB does not act as a spatial topographic map, but acts as a PATTERN SEPARATOR, which uses contrast enhancement to DECORRELATE similar input patterns.


Whitesell et al (2013)

T: patch clamp recordings, sectioned external plexiform layer (where the dendrodendritic synapses between mitral and granule cells are located)

R: Interglomerular lateral inhibition occurred after sectioning EPL - contradicts the idea that granule cells are responsible for lateral inhibition.

Authors argued for functional separation of GABAergic cells in the MOB:

1) GABAergic short axons cells in glomerular layer: mediate inhibition of mitral cells between glomeruli,

2) GABAergic granule cells that project to EPL, which are responsible for synchronising glomeruli.

L: in vitro rat slices


Niessing + Freidrich (2010)

T: Calcium sensitive dyes - image MC activity, in response to different AA odorants (diff concs. Deconvolution matched calcium activity with the underlying AP dynamics, and principle component analysis showed that temporal patterns of activity for different odorants are highly correlated between similar concentrations of an odorant.

R: MIXTURES of SIMILAR odorants represented by patterns that correlated with an INDIVIDUAL.

Mixtures of DISSIMILAR odorants produced patterns of activity that were entirely SEPARATE to those that represented the individual components.

illustrates the role of the MOB as a pattern SEPARATOR, where synthetic patterns are generated to represent odours, rather than representing the proportions of individual components.

L: zebra fish slices, may not translate to activity patterns in mammalian MOBs.


Schusterman et al (2011)

T: studied AWAKE mice (most sniff work on anesthetsied rats) electrophysiological recordings from electrodes implanted in DORSAL & VENTRAL MC layers.

R: INCREASE in MC activity that was more closely aligned with the ONSET OF INHALATION, as opposed to the onset of stimulus presentation. PEAK responses of INDIVIDUAL MCs occurred throughout the course of the sniff cycle, and that ANALYSIS of PEAK firing within the sniff cycle may be used by a COMPUTER to potentially DISCRIMINATE different odours. (highlights how temporal representation of information MOB, in relation to sniff cycle, important in odour discrimination in mammals)


Smear et al (2011)

T: OPTOGENETICs, AWAKE MICE, more precisely characterise the timing of MC input in mice. Inserted channel rhodopsin gene into mature OSNs, and used light stimuli to depolarise the OSNs whilst measuring the sniff cycle.

Go/no-go behavioural task trained to DISCRIMINATE between LIGHT STIMULI delivered at 2 diff points in the SNIFF CYCLE

R: ACTIVELY USE temporal coding of information in the MOB to DISCRIMINATE stimuli.

ELECTROPHYS recordings showed that when light stimuli were delivered at different time points in the sniff cycle, INDIVIDUAL MCs responded to these stimuli with varying AMPLITUDE and varying SPEEDS of activation. illustrates how the intensity and timing of activity in the MOB in relation to the sniff cycle can be used to encode odourant stimuli.


Verhagen et al (2007)

Calcium imaging MOB: AWAKE, head-fixed mice. During HF sniffing: glom. activity LOWER than predicted (suggesting that adaptation occurring). Repeat stimulus presentation = LF sniffing with expected glom. activity.

When ethyl butyrate presented on butanone bckgrnd - if EB was novel, mice showed HF-sniffing + mitral cell activity that reflected the DIFFERENCE between the activity patterns for the individual odourants. If EB was familiar, LF-sniffing occurred, and glomerular activity reflected a SUMMATION of the activity patterns for each component.

R: HF-sniffing is important for adapting out bckgrnd odours, so novel odours can be clearly discriminated from bkgrnd odourants.

However, mechanism by which SNIFF FREQUENCY-DEPENDENT ATTENUATION of glomerular input occurs still somewhat unclear. Primary mechanism: receptor adaptation? higher sniff frequencies allow less time for OSNs to recover from adaptation between each inhalation.

Glomerular activity became attenuated within ~2s during HF-sniffing - supported by Reisert (2001): OR currents decayed to ~20% of their original value within ~2s of first presenting odourant. Supports idea that HF sniffing enhances adaptation at level of OR by preventing recovery from adaptation.


Stettler + Axel (2009)

Over 100 mice: calcium imaging: pyramidal cells. Responses to 16 individual odourants.

R: No spatial patterning across the anterior and posterior PC, not even at coarse level, (like coarse clustering seen in MOB)

Large number of cells responded to octanal or a-pinine - fewer number of cells became active in response to a mixture of both the odourants.

R: Piriform cortex does not respond to mixtures in additive manner - generates new patterns of activity in response to mixtures.


Davidson & Ehlers (2011)

Mice: Glutamate uncaging to optically activate dorsal MOB, recorded mitral cells + pyramidal cells (anterior piriform cortex).

Optically activating single glomerulus not sufficient to activate associated pyramidal cell, but CO-ACTIVATION of 3-4 GLUTAMATE UNCAGING SITES (= ~6-10 GLOMERULI) = sufficient to generate pyramidal cell firing.

Each pyramidal cell responded to different patterns of glomerular activity (where patterns were generated by 16 active glomeruli), which suggests that the anterior PC acts as CO-INCIDENCE DETECTOR, and detects slightly different patterns of glomerular input + therefore different patterns of OR activation.


Chapuis and Wilson (2012)

(relevant to role of piriform cortex)

Awake rats, forced choice task, then anaesthetised and electrophysiology

Learn to discriminate between an 10c and 10cR1, less able to discriminate between 10c and 10c-1. Patterns of activity in MOB were very different for 10c and 10c-1, but anterior piriform cortex produced similar activity patterns for them.

Highlights role of aPCX as odour object recogniser, as can generate complete pattern of activity from an incomplete pattern of input, in order to generalise similar odourants.


Chapuis + Wilson (2010)

(relevant to role of learning)

Forced choice task, rats, then anaesthetised and electrophysiology

Trained to discriminate between 10c and either 10c-1 / 10R1. Initially, 10c-1 more challenging than 10R1 (because anterior PC usually generates similar patterns for 10 and 10c-1).

After 8 days of training: anterior piriform cortex showed DECORRELATION of activity patterns for 10c and 10c-1.

When rats retrained to treat odours as the same: increase in correlation of activity patterns for the two different odourants.

Highlights how the process of generalising similar odours can be CHANGED by LEARNING, by directly altering the patterns of activity in the piriform cortex.

Note: recordings were performed under anaesthesia. As anaesthesia shown to change activity patterns in MOB (Kato et al), to further confirm these findings, a similar experiment with a method of recording pyramidal cell activity in awake animals would be desirable.


Haddad et al (2013)

Mice - Optogenetics, OSNs inserted with channel rhodopsin- optically stimulate 2 separate areas of MOB.

Firing rate of some pyr. cells unaffected by timing of the 2 stimuli. Some pyr. cells exhibit SYMMETRICAL TEMPORAL RESPONSE PROFILES (pry. cell firing rate dependent on synchronicity of the 2 MCs stimulated). Some have ASYMMETRICAL temporal response profiles (order in which the 2 MCs are activated determines firing rate of the pyramidal cell).

R: Pyramidal cells in piriform cortex are SENSITIVE to the relative TIMINGS of glomerular activation, within the order of 10s of milliseconds. This is similar to the timing differences that altered the behaviour of mice in Smear et al’s (2011) study.

L: contrasts with Davison + Ehler's as in this study single stimulations of one glomerulus could activate the pyramidal cells BUT authors admit using larger spots of light so prob activating multiple glomeruli


Wilson (2001)

Prolonged stimulation with odourant causes reduction in response in both the MOB and piriform cortex

After 10-20s, piriform firing rate reduced to ~25% of its initial rate, whereas MOB reduced to ~75% of its initial firing rate. Therefore, HABITUATION in the piriform cortex is much more PROFOUND than adaptation in MOB (partly explained by mGlu2 on MC axon terminals - Glu release causes auto-inhibition - reduces post-synaptic activation of pyramidal cells, even when MC firing maintained)

MOB + piriform cortex also differ in the SELECTIVITY of their adaptation responses. CROSS-adaptation occurs readily in the MOB, (adaptation to one odourant = reduced response to odorants that activate same OR).

Very little occurs in piriform- supports idea that MOB recognising individual odourant features (similar structures) because MC will adapt to all of the odourants it responds to, whereas the PC is recognising odour objects.


Reisert (2005)

RT-PCR: mouse OSNs express NKCC1, but not NKCC2

Electrophysiology: KO mice for NKCC1 did not show the outward chloride current that is seen in WT mice.

Suggests that the NKCC1 channel is primarily responsible for generating the electrochemical gradient that is required for the outward chloride current.

note - reisert 2001 showed that there is 80% decay of current after 2 seconds of presenting odourant - different study, supports HF sniffing/adaptation role


Kim et al (2012)

- in terms of transduction

M: Recordings from in vitro mouse slices of VSNs, in response to urine. Also used KO GIRK/SK3 and GIRK/SK3 inhibitors.

In vitro slices: Potassium efflux attenuated TRPC2-mediated / CACC-mediated depolarisation

Extracellular recordings from intact preparations: both channels enhance urine-induced depolarisation in VSNs.

Ion-sensitive electrode in VNO mucus: ↑[potassium]: ∴ in vivo, SK3 / GIRK activation = depolarising, inward current


Kim et al (2012)

- in terms of behaviour..

SK3/GIRK1 KO/inhibitors: channels enhance depolarisation of VSNs in response to urine. Innate behaviours = resident-intruder aggression assays + mating assays.

SK3, GIRK1 and TRPC2 KO = impaired aggressive + sexual behaviour.

TRPC2 KO: MORE REDUCTION in AGGRESSIVE behaviour whereas the SEXUAL behaviour was somewhat LESS IMPAIRED (than SK3/GIRK1 KO)

L: Gene KOs used in this experiment were not restricted the VNO. Therefore, it is possible that gene silencing in other areas of the CNS may have contributed to the behavioural deficits that were measured.

“Alternatively, it is possible that distinct populations of neurons reside in the VNO and that the relative contributions of TPRC2, SK3 and GIRK1 differ in different cells.” then these neurons activate different behaviours.


Nara et al (2011)

Calcium imaging of dissociated OSNs (female mice) - 3000 OSNs, 13 mixtures.

81 OSNs responsed to 36 different sets of aldehydes

Some odourant mixtures activated 3-5x more OSNs than others, and some individual odourants activated 5-10x more OSNs (particularly citrus/fruity).

Most OSNs narrowly tuned (responded to similar structures) but some broadly tuned.

Most odour codes were shown to be unique and combinatorial i.e. individual OR responses are ambiguous but if examine whole population of receptors – unique activation pattern.

However: dissociated OSNs removes sustentacular and microvillar cells (supporting cells), which may influence the firing rates of OSNs - although less relevant to ligand responses.

Recordings from intact preparations result in a 100% response rate to ligands, lower response rate to dissociated neurons - also kept in different solutions.


Dibattista (2012)

Whole cell patch clamp isolated VSNs

In response to calcium uncaging, a large inward current is seen when the cell is held at -50mV, which was partially inhibited by NFA.

Furthermore, if chloride was removed from the extracellular solution and replaced with other membrane impermeable anions, the reversal potential of this current was altered.

Strong evidence that the calcium activated current is a chloride current


Isogai et al (2011)

Egr1 expression

48% of V1R-expressing VSNs = ‘promiscuous’ >1 of: male, female, mammalian non-predator, mammalian predator, reptile, avian predator.

9% of V2Rs = promiscuous

Clades of V2Rs evolved to detect urinary proteins of specific mammalian predators, other clades for non-mammalian predators.

Sulphated steroids: (steroid hormones sulphated by liver then excreted): mixture activated V1Ref and V1Rjk clades - a specific oestrogen, androgen and glucocorticoid were found to activate specific receptors within these clades.


Kobayakawa et al (2007)

CreLoxP: diphtheria toxin in neurons with O-MACS promotor (dorsal zone specific). ΔD = ablate dorsal zone OSNs ΔII = ablate class II (both dorsal + ventral).

ΔD: glomeruli restricted to ventral MOB zone. ΔII: glomeruli restricted to D1 domain (hypoplasia of whole MOB).

ΔD: no innate aversion to odourants (e.g. predator, spoiled food), but still distinguish them & learn conditioned attraction response (detect with ventral domain receptors & generate learned responses via general odour analyser system, but lack innate aversion). ΔII: no innate aversion to predator odours, but aversion to spoiled FOOD.

R: avoidance to spoiled food mediated by Class I olfactory receptors, whereas avoidance to predator odourants is mediated by dorsal Class II receptors.

L: ΔD mice actually demonstrate an innate attraction to some aversive odourants. If dorsal receptors are simply responsible for innate aversion, loss of dorsal receptors in these mice cannot adequately explain why innate attraction was observed.

Mice tested as adults, therefore cannot completely disregard role of learning / previous exposure to some of the odourants.


Main olfactory epithelium - innate responses?

Guanyl cyclase (GC) dependent OSNs (separate transduction pathway) -project to necklace glomeruli.

Bleymehl et al (2016): calcium imaging - Gucy1b2 neurons are activated by low O2. Conditioned place preference test: activation results in avoidant behavioural response (conditioned place aversion).

May have role in mediating innate behavioural responses to O2 levels - mice select higher O2 areas to burrow nests (survival).

TRPM5 OSNs: project to the ventral MOB (social stimuli)

Lopez et al (2014): calcium imaging in mice to show that these receptors respond to pheromones, urine, and MHC peptides, but not general odourants

ALSO: TAARs, OR37 neurons


What are TAARs

Trace amine associated receptors

Subtype of OSN, distinct from the canonical OSNs: highly conserved between species.

Follow classical transduction pathway of canonical receptors, but express a distinct family of TAAR genes (15 different receptors exist in mice).


Zhang et al (2013)

Mice: in vitro patch clamp: dendritic knobs of TAAR expressing OSNs

In vivo imaging of the associated glomeruli

Individual TAAR-expressing OSNs can respond to variety of structurally diverse amine compounds (broadly tuned), TAAR4 expressing OSNs are also highly sensitive to phenylalanine (PEA), a compound found in cat urine - can respond at concs as low as 10⁻¹² M!

Human TAAR5 (hT5), when expressed in mouse OSNs, also responded to some amine compounds, but with much lower sensitivity that the mouse TAAR receptors.


Dewan et al (2013)

Tau-associated fluorescent markers in TAAR-expressing OSNs: converge onto glomeruli in the dorsal region of MOB, (therefore - mediate innate aversive responses- Kobayakawa?)

In vivo Ca imaging (TAAR-glomeruli): KO TAAR 2-9 - no amine response, but preserved non-amine responses. Loss of innate aversion to predator odours (PEA) and other amines in a place preference test, although some aversion was demonstrated when amines were presented at very high concentrations.

TAAR4 KO: loss of innate aversion to PEA and predator urine, but aversion to other amine compounds is preserved.

Very strong evidence that TAAR4 evolved to be highly sensitive to predator urine, and mediates an adaptive behavioural response - mice avoid locations where predator been.


What are OR37 OSNs?

Highly evolutionary conserved subtype of OR in MOE. OR37 genes show 80% homology between humans and mice.

Project to the ventral MOB, but are distinct from the rest of the main olfactory system, as have a direct projection to the paraventricular nucleus (PVN).

Respond to long-chain aldehydes, (the optimal stimuli for OR37A, OR37B and OR37C).

Shown that when these neurons respond to mixtures of aldehydes, cFOs expression in CRH neurons within the PVN is inhibited.

Suggests that OR37 system may have an anti-stress effect, where the release of glucocorticoids from the adrenal glands is inhibited.


What are pheromones?

1959: “substances secreted to the outside of an individual, and received by a second individual of the same species, in which they release a specific reaction, for example, a definite behaviour or developmental process”.

Today, pheromones are often described as substances that are produced and received by a single species, and provide an evolutionary benefit to both animals.


What are chemosensory organs that have roles in innate behavioural responses?

In addition to innate responses mediated by OSNs within the MOE: distinct chemosensory organs.

In mice: septal organ, vomeronasal organ, and Grueneburg ganglion.

Some of thought to have a role in mediating responses to pheromones.

The Grueneburg ganglion (GG) projects axons to the NECKLACE glomeruli of the MOB, and Brechbuhl et al (2008) showed that sectioning these axons in mice, results in degeneration of the chemosensory cells within the GG, and a loss of freezing behaviour in response to alarm pheromones.


What are alarm pheromones?

Alarm pheromones are released by mice under stress, and usually result in a fear response in other mice. This is thought to be an adaptive response in order to avoid the attention of predators.


Soucey et al (2009)

Imaged glomeruli in rats & mice & identified them based on responses to many odorants.

Many individual glomeruli in similar positions between left & right bulb within individuals, and between animals of the same species.

Supports idea that odours may be spatially represented within MOB, but further investigation showed glomeruli responding to similar stimuli are only clustered in a coarse manner, with glomeruli responding to dissimilar molecules scattered among them.

Suggests that stimulus-response mapping is not present in the MOB at a precise level - MOB does not represent odours in a functional topographic map.

L: patterns of glomerular activity may be altered under anaesthesia (Kato et al, 2012).


He et al (2008)

Many mouse V1R VSNs respond to both female and male urine, and urine from different strains of mice.

Further examination with principle component analysis: unique patterns of activity across the V1R population could separate different strains of mice. (similar to the population coding that occurs in the main olfactory epithelium)

Also identified very small number of cells that could individually recognise gender: female urine specific cells (FUSCs) & male urine specific cells (MUSCs). FUSCs = 2.6% of VSNs that were imaged & MUSCs <1%.


Wagner et al (2006)

Evidence for axonal guidance within VN system (regenerating VNE axons converge on appropriate glomerulus).

Flourescent labelling of range of different V1Rs within different clades

Neurons expressing closely related V1Rs (within the same clade) converge within the same domains of the AOB (similar to the zonal projections seen in the MOB).

In the AOB, the domains are arranged along the dorso-ventral axis of the bulb, like a checkerboard. Receptors from individual V1R clade all project to the same domain, however, >1 receptor clade may project to a single domain (e.g. V1Rj clade & V1Rk clade project to same domain).


Wagner et al (2006)

Evidence for selective heterotypic connectivity

Selective heterotypic connectivity = mitral cells innervate glomeruli that receive input from receptors all belonging to same clade.

Injected fluorescent dye into AOB mitral cells: visualised their apical dendritic tufts, and genetically labelled different glomeruli for their inputs from different receptor clades.

Single mitral cell could form dendritic connections with glomeruli that received input from receptors that were of different subtypes, but within the SAME CLADE.


What is homotypic connectivity?

In the main olfactory system, OSNs expressing single OR will converge on individual glomerulus. A single mitral cell will only have dendritic projections to a single glomerulus, therefore mitral cells receive information in a homotypic way, as each mitral cell only receives input from one OR type.


What is heterotypic connectivity?

VSNs that express a single vomeronasal receptor type do not converge onto a single glomerulus - can converge onto 5-20 glomeruli.

In the VN system, mitral cells have branched primary dendritic trees, which can synapse with ~5 several glomeruli. Therefore, mitral cells may integrate information from many different receptor types at the level of the first synapse.


Sakomoto et al (2011)

Ablated new neurons by using tamoxifen-treated (NSE-DTA) mice (tamoxifen-inducible Cre recombinase causes DTA expression in differentiating neurons - efficient ablation of newly born neurons in the forebrain).

Mutant: discriminate odors as well as controls.
Both showed freezing to fox odor (predator), but MUTANT mice APPROACHED this odor when they were conditioned to associate the odor with a reward, whereas control mice did not approach the odor.

Mutant males + females showed normal social recognition behaviors to opposite sex mice

However: mutant MALES = DEFICITS in MALE–MALE AGGRESSION + SEXUAL behaviours to females, whereas mutant FEMALES = deficits in FERTILITY and nurturing behaviours, indicating that sex-specific activities, which are known to depend on olfaction, are impaired.

Suggest continuous neurogenesis is required for predator AVOIDANCE and SEX-SPECIFIC responses that are olfaction dependent and innately programmed.


Luo et al (2003)

AOB mitral cell recordings: mice investigating other mice.

Peak firing after 9-10s, remained 10-30s (slower response time & more sustained response then MOB). May link to role e.g. FELD4 (cat urine) = long term avoidance to areas frequented by predators (stays in environment > volatiles that affect MOB). AOB relatively low firing rates (peak 20-30 Hz), whereas MOB mitral cells can fire at 60Hz.

INDIVIDUAL mitral cells in AOB show excitatory + inhibitory responses to diff mice strains and to diff sexes within these strains. Even at mitral cell (first level of encoding), neurons can respond selectivity to strain + sex, and may have system of CONTRAST ENHANCEMENT where inhibitory mechanism sharpens mitral cell responses to diff strains.

High selectivity to conspecifics: mice may create a “pheromonal image” of conspecifics by using relatively small but highly specific pop of neurons, rather than extracting picture from large pop of broadly tuned neurons. Consistent with 'labeled lines' model for conspecifics (info about their gender and genetic makeup)

L: Number of mouse strains and substrains is large, and the number actually tested SMALL (BALB6 / C57BLB6 / CBA)

integration likely for encoding complete pheromonal images of conspecifics - substantial number of cells responded in some manner to one of the strains used, animals with different genetic backgrounds must have overlapping pheromonal composition and thus activate overlapping populations of AOB cells, implying that some mechanisms providing additional discriminatory power will be required)



Ben-Shaul et al (2010)

Electrophysiological recordings of AOB in anaesthetised mice: many mitral cells able to respond to pheromones from either male or female mice, and many able to respond to predator urine in addition to mouse urine.

Contrasts idea of ‘labelled lines’ in the vomeronasal system, where specific signals are linked to very specific responses in the olfactory bulb, and suggests that individual patterns of activity across a wide range of mitral cells will convey information about an individual vomeronasal stimulus.

Strength = FUNCTIONAL - based on stimulus delivery (stimulus delivery to vomeronasal organ very difficult to get so this was good) but anaesthesied (think Kato).



Kato et al (2012)

In vivo calcium imaging of head-fixed mice (Cre-recombinase, express GCaMP3 in mitral)

Selectively imaged same 100 mitral (& inhibitory) cells to see how activity changed

R: Wakefulness enhances activity of inhibitory granule cells and makes principle mitral cell odour responses more sparse (efficient) & temporally dynamic.

Longitudinal exposure to odourant over several days = progressive sparsening of mitral cell activity. If animals removed from odourant for 2 months, the pattern of activity in response to this odourant returned to the original, naïve pattern. These data show that experiences of an odour can cause a long-term, but reversible, sharpening / fine tuning of mitral cell activity in response to a given odourant.

Limitations: Only two types of anaesthetic used, could be mode of actions of anaesthetic causing different effects.


Kay and Laurent (1999)

Awake mice, electro recordings mitral cells in go/no-go task training (2 odourants).

When reward contingencies changed, (one paired with sucrose reward, one with bitter taste), mitral cell firing rates differed for each odour. When contingencies returned to equal (both sucrose), mitral cell firing rates were equal for each odourant.

Mitral cell activity particularly associated with VALUE of odourant rather than odourant itself.
Only 11% mitral cells changed firing rate during odour sampling, whereas 94% changed firing during behaviours surrounding the sampling period.

Further suggests that MOB activity continually changing due to centrifugal neuromodulation, and changes to intrinsic connectivity that are associated with the experience and behaviours of an animal.


Schoenbaum (2002)

Recorded PC in awake rats during go/no-go

Activity in PC dependent on meaning of the odour, previous odours that rat had been previously exposed to, & which odour would be predicted next within a sequence. Ultimately highlights the complexity of the information conveyed in the patterns of activity within the piriform cortex, and that learning and previous experience can alter these patterns in a vast number of ways.


Doucette ?

Compared multi-unit MC recordings with single-unit recordings in mice- go/no-go task.

Start: synchronisation of MC action potentials similar for the rewarded and unrewarded odours: as task progressed, ↑ synchronised APs in response to rewarded odour. When reward contingencies swapped, newly rewarded odour produced ↑ in firing rate.

R: Divergence in rate of synchronised mitral cell spikes occurs during the process of learning (supports Kay and Laurent- MC activity highly dependent on meaning rather than odour itself). ADRENERGIC receptor ANTAG 10 mins before go/no-go task ↓magnitude of the divergence in synchronised firing.

L: Little justification for the 250µs time period used to determine whether 2 MC APs were synchronised. If diff window selected, results may have differed significantly, although currently no established time window that has clear functional relevance to the process of summation /synchronisation in the piriform cortex.

Although largely attributed to centrifugal influences, could also be neuroplasticity within MOB? Mechanisms underlying development of differential synchronised responses not well understood, but NA projections from the LC to the MOB may be modulating these responses (supports centrifugal idea).


Leinders Zufall (2000)

V1R-expressing VSNs are highly selective for individual small, volatile pheromones.

Calcium imaging in mouse slices: activity of VSNs in the apical VNE in response to 6 putative pheromones.

Distinct subset of VSNs responded to each pheromone: very little overlap between groups in response to each pheromone. Very few VSNs responded to >1 of the pheromones.


Leinders-Zufall (2004)

MHC-dependent peptides can act as ligands at Class II VSNs (which are found on the basal region of the vomeronasal epithelium, and are coupled to Gao proteins).

Extracellular field potentials from intact VNO preparations of C57BL/6 mice

Individual V2Rs could selectively respond to MHC-dependent peptides from either a BALB/c mouse or a C57BL/6 mouse. (highly sensitive, dose-dependent response - near max responses at concs of 10⁻¹³M!!)

Changing anchor residues of peptides obliterated responses, whereas altering sequences outside the anchor residues preserved the dose-response curve.

Very strong evidence that Class II VSNs are selective for anchor residues of MHC-dependent peptides.


Munger (2001)

CNGA4 -/- and WT mice showed equal magnitudes of depolarisation in response to an odourant, however, KO showed less adaptation to the application of odourant, and application of a PDE inhibitor (chemical stimulation by increasing cAMP).

Reduced adaptation was shown in response to both prolonged and repeated stimulation.

L: excised membrane patches and therefore may have little relevance to olfactory function in vivo.


What are sig mixtures?

Chemical signals olfactory system has evolved to detect (or evolved to be detected) which don’t really fit with pheromone definition as don't mediate an innate response (way information is acted upon depends on learning)

Convey information about individual identity or state e.g. hormonal state,
* The way this information is acted upon, depends on learning

Behavioural contexts requiring discrimination of individual identity (mice vary greatly in the chemical signals they produce, these signals can be used by other mice to distinguish/recognise individuals).


Signature mixtures can influence?

Territorial behaviour (males marking territory, signal identifies producer)

Mate choice (males maintain fully marked territory much more successful/competitive mates - female relies on olfaction to choose best mates)

Mother-offspring recognition (several species in which young recognises mother or vice versa on basis of odours)

Social organisation and kin recognition

Mate recognition (Bruce effect) - when female mouse recognises male she has just mated with - prevents pregnancy being lost - protective effect on pregnancy


What are MUPs?

Mouse major urinary proteins

MUPs themselves highly POLYMORPHIC and directly activate vomeronasal sensory cells: each animal can express 5-10 different MUPs, each with distinctive profile


Also bind small testosterone dependent urinary volatiles which can activate VN and OSNs (can bind ligands with pheromonal activity) e.g. urinary volatiles that are effective in eliciting aggression


What is Darcin?

A specialised MUP = Darcin

Involved in territorial behaviour and eliciting male aggression
(but whole family of MUPs with different role - signalling individual identity)


What are involatile MHC-dependent peptide chemo-signals?

Cellular proteins degraded intracellularly by proteosome: shorter peptides (9 AAs). Peptides retrieved from the DEGRADATION pool & presented on surface by MHC CLASS 1 proteins. During development of immune system, recognise presented peptides as endogenous intracellular protein - learning ‘self’, but if viral proteins presented, marked for destruction.

MHC Class I = highly polymorphic (differ between individuals), therefore subset of peptides fished from the degradation pool will be different

Differences in the peptides retrieved from the degradation pool is determined by the BINDING CLEFT of the MHC Class I protein - binds ANCHOR residues, which are in particular positions on the peptide (large, hydrophobic amino acids)

If VNO receptors have the same anchor residue binding specificity as the MHC proteins, diff VNO receptors able to selectively respond to anchor residues of peptides typically presented on cells of specific strains


What responds to MHC peptide ligands?

v2Rs respond to anchor residues (Leinders-Zufall 2004)

BUT MOE can also respond to MHC peptide ligands (Spher et al 2006) - will respond to these peptides down to about 10^-9M concentrations - not as sensitive as vomeronasal system, but still quite sensitive

Would not expect peptides to be volatile - volatility usually below ~400 molecular weight. However, if main olfactory system can respond to MHC peptide ligands - likely that they are reaching MOE? Could this mean that main olfactory epithelium can respond to other relatively involatile components as well?

little known about this *direct contact - snorting up mucus/skin secretions - transported up to MOE? perhaps more likely that fine particles of dust get into MOE

Painted some rhodamine (fluorescent red dye) onto anogenital region of female mouse
* Then male investigated mouse, and found rhodamine right up in the main olfactory epithelium of the mouse
* This shows that non-volatile stimuli can also stimulate the main olfactory system


What is ESP1?

Exocrine secretory peptide

Pheromone in male mice tears - sensed by VN system - painted on males, dramatically increases lordosis in females (secreted from glands then into tears)

Knocking out V2Rp5 receptor abolishes effect of ESP1

Produced by WT animals & by certain inbred strains of mice, but not produced at all by C57 Black 6 (C57BL/6) mice, common inbred mouse strain used in labs

*but because they don’t produce ESP1, don’t get the pheromonal effect in the mouse

*lots of inbred strains of mice don’t have responses to certain pheromones - lost during process of selective breeding - how useful is it to use inbred strains - espec. if looking at pheromonal responses / behaviour of animals?


Evidence for role of urinary volatiles and MUPs in aggression?

Castrated male not seen as normal male (ignored, like juvenile males - don’t produce testosterone)

Then normal male urine painted on back of castrated male: normal mouse now aggressive towards mouse

Aggression induced by both high molecular weight component & low molecular weight fractions of the urine when compared to no urine controls

*Supports evidence that volatile ligands are able to elicit aggressive behaviour - normally they bind to major urinary proteins (MUPs): cups that bind the volatile molecules- but if strip the ligands away from the MUP, the MUP alone is sufficient to cause aggressive behavioural response