Background and significance

Characterizing olfactory generalization in Fischer 344 rats using behavioral reaction timesWendy M Yoder1 � Olivia Munizza1 � Leslie Gaynor2,4 � Ethan Windham3 � Michelle Lyman4 � Barry Setlow4,5,7 � Jennifer L Bizon4,5,6,7 � David W Smith4,6

1Program in Interdisciplinary Studies, Neurobiological Sciences � 2Department of English � 3Health Science Program � 4Program in Behavioral and Cognitive Neuroscience, Department of Psychology � 5Department of Psychiatry � 6Center for Smell and Taste � 7Department of Neuroscience, University of Florida College of Medicine, Gainesville, Florida

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Behavioral methodology: Operant Chamber

Acknowledgments

References

Modified Operant Chamber

Odor Port

Odor Stream

Stimulus Behavior Response Measure Observation

Figure 2: Psychophysical principle of reaction time. Reaction times can be used to measure stimulus intensity. In the illustration above, the hotter the flame, the faster the hand will withdrawal. Similar behavioral methodology may be applied to odor-guided tasks to create a generalization gradient and increase difficulty.

Stimulus Behavior Response Measure Observation

Figure 4: Procedure for using non-reinforced probe trials. (A) If the rat identifies the mixture (S+ : S-) as the target (S+), the rat will lick. If the animal decides the mixture (S+ : S-) is NOT the target (S+), the animal will not lick. (B) How fast the rat licks in response to the mixture is a measure of how certain the rat is that the mixture (S+ : S-) is the same as the target (S+).

=

MixtureS+ : S-

Rat SniffsS+ or S- ?

Decision: S+Rat Licks

Delay to Decide / Lick

Probe smells like S+

Figure 3: Operant chamber. Licking in the presence of the target odorant (S+) results in 5 µl of liquid reinforcement (Ensure). Conversely, incorrectly licking in the presence of the control odorant (S-) results in a 5 second time-out; the rat cannot initiate new trials during this interval. Reaction times are defined as the interval from the time of stimulus onset until the rat makes contact with the lick tube.4

Behavioral

BiomarkerSeveral

LimitationsNeed Better

TranslationalMeasures

OdorIdentification

Target

Choices

A) Can rats be trained to “say” POPCORN?B) Can the odor-guided task be more

difficult to minimize ceiling effects?

• Olfactory changes are an early sign of many forms of neural and cognitive dysfunction.1

• To thoroughly investigate the underlying mechanisms involved in these processes, cross-species assessments are needed that are both sensitive and specific.

Figure 1: Odor identification schematic. Rodents are frequently used in olfactory research, but current measures are often not comparable to olfactory tests used with human subjects.2

Cross-species

Comparisons

Step 1: Obtain thresholds for individual ratsStep 2: Reinforcement (100%) on S+ trials

Step 3: Partial reinforcement (50%)

Step 4: Non-reinforced probe trials S+ : S- S+ : S- S+ : S-S+ S-

+ + +

Criterion:100% Accuracyvs.

S+ S-

Criterion:100% Accuracyvs.

S+ S-

Figure 3: Stimuli Presentation. Each regular block contains 20 trials (10 S+ and 10 S-). Probe trials (2 per block) are pseudorandomized and presented during 5 separate sessions (n=10 per mixture ratio).

Individual thresholds are used to generate binary mixturesFigure 6. Comparison of individual, threshold estimates for octanol and citral. Columns refer to descending concentrations (% v/v) of the dilutions. Each panel/symbol represents discrimination accuracy for the target odorant (octanol or citral) vs. the odorless, control odorant (diethyl phthalate). Lines represent accuracy across three consecutive blocks for each concentration. The lowest concentration at which the rat receives 85% or greater on at least one of the three blocks is recorded as threshold. These are used in subsequent steps to match intensity levels between odorants.

Threshold:Lowest concentration the rat receives ≥ 85% vs.S+ S-

vs.S-S+

vs.

individual Mixture Probe Trials

Figure 8: Group mean reaction times (ms) for binary mixture ratio probe trials. Circles represent mean reaction times for non-reinforced probe trials containing different concentrations of citral and octanol. For visual simplification, only the two highest ratios recorded as S- are shown here.

Figure 9: Individual reaction times for non-reinforced, probe trials. Each mixture ratio contains 10 probe trials (2 per block) randomized across sessions. Ratios not shown indicate the rats did not lick – responded to the mixture as S-.

Figure 7: Comparison of mean reaction times (ms) for reinforced target (S+) trials and non-reinforced, probe trials of equal concentration (threshold level). Mean values are not statistically different (i.e., reinforcement contingency does not affect reaction times).

Funding for this research was provided by NIH R01 AG024671 and the McKnight Brain Research Foundation.

Asymmetrical Response

Aging is a complex process and a key risk factor for cognitive and neural dysfunction, but declines are rapid without early intervention. Researchers are scrambling to identify biomarkers.

Is olfactory dysfunction a promising behavioral

biomarker?

This measure could be included in a battery with

other olfactory tests.

Importantly, our laboratory has expertise assessing olfactory acuity in both

humans subjects and animal models.

Fischer 344 rats may be an important piece of the puzzle. As these rats age, some show cognitive declines that correlate with olfactory impairments. Olfactory changes may be predictive.4

For both citral and octanol, reaction times were rapid (~150-200 ms) when the probe trials consisted of a single, monomolecular component.

Binary mixtures that differed from the target odorant by as little as 1% v/v were different enough to increase reaction times -- rats hesitated before responding.

Reaction times changed systematically as a function of task difficulty for both odorants, but the steepness of the generalization gradients depended on the target odorant presented.

95:5 84:16

Figure 10. Asymmetrical responses.5 Less citral was required to “suppress” the presence of octanol than vice versa. On average, animals ceased responding to the probe trials as the target at the ratio 95:5 for octanol and 85:15 for citral.

• Probe trials “ask” the rat to respond without the contingency of reinforcement.

• This measure captures subtle, behavioral changes and minimizes ceiling effects.

• Mixtures can be more or less complex depending on the experimental objectives.

Disadvantages

Advantages

• Threshold matching is labor intensive.• Stimuli interaction effects should be

considered.

1. Atanasova, B., Graux, J., El Hage, W., Hommet, C., Camus, V., Belzung, C. 2008. Olfaction: a potential cognitive marker of psychiatric disorders. Neuroscience and biobehavioral reviews 32(7), 1315- 25.

2. Doty R, Frye R, Agrawal U. 1989. Internal consistency reliability of the fractionated and whole University of Pennsylvania Smell Identification Test. Perception & Psychophysics 45: 381-384.

3. LaSarge, C., Montgomery, K.S., Tucker, C., Slaton, S., Griffith, W., Setlow, B., Bizon, J. 2007. Deficits across multiple cognitive domains in a subset of aged Fischer 344 rats. Neurobiology of aging 28(6), 928-36.

4. Slotnick, B. 2007. Odor-Sampling Time of Mice under Different Conditions. Chemical Senses 32(5), 445-54. 5. Sokolic L, Laing D, McGregor I. 2007. Asymmetric suppression of components in binary aldehyde mixtures: behavioral studies in the laboratory rat.

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