Effect of Sensory Deprivation of Nasal Respiratory on Behavior of C57BL/6J Mice

Abstract

Nasal breathing is a dynamic cortical organizer involved in various behaviors and states, such as locomotion, exploration, memory, emotion, introspection. However, the effect of sensory deprivation of nasal respiratory breath (NRD) on behavior remain poorly understood. Herein, general locomotor activity, emotion, learning and memory, social interaction, and mechanical pain were evaluated using a zinc sulfate nasal irrigation induced nasal respiratory sensory deprivation animal model (ZnSO₄-induced mouse model). In the open field test, the elevated O-maze test, and forced swim test, NRD mice exhibited depressive and anxiety-like behaviors. In memory-associated tests, NRD mice showed cognitive impairments in the hippocampal-dependent memory (Y maze, object recognition task, and contextual fear conditioning (CFC)) and amygdala-dependent memory (the tone-cued fear conditioning test (TFC)). Surprisingly, NRD mice did not display deficits in the acquisition of conditional fear in both CFC and TFC tests. Still, they showed significant memory retrieval impairment in TFC and enhanced memory retrieval in CFC. At the same time, in the social novelty test using a three-chamber setting, NRD mice showed impaired social and social novelty behavior. Lastly, in the von Frey filaments test, we found that the pain sensitivity of NRD mice was reduced. In conclusion, this NRD mouse model showed a variety of behavioral phenotypic changes, which could offer an important insight into the behavioral impacts of patients with anosmia or those with an impaired olfactory bulb (OB) (e.g., in COVID-19, Alzheimer's disease, Parkinson's disease, etc.).

Keywords: C57BL/6J mice; behavioral test battery; nasal breathing; sensory deprivation of nasal respiratory.

Figure 1

An illustration of the experimental timeline. Open field test, OFT; elevated O maze, EOM; forced swim test, FST; Y-maze test, YMT; novel object recognition test, NORT; contextual fear conditioning test, CFC; tone-cued fear conditioning test, TFC; social novelty test, SNT; von Frey filaments test, VFFT.

Figure 2

Normal locomotor activity and anxiety-like behavior in NRD mice. (A) Diagram of the open field apparatus (the peripheral (between solid and dotted lines) and central (inside the dotted line) regions). (B) Representation of the locomotion tracing for control and NRD mouse in the open field test. (C–F) Performances of the control and NRD mice in terms of distance in open field (C), moving speed in open field (D), in center zone (E), and number of center squares entered (F). * p < 0.05 as determined by unpaired Student’s t-test. Data are expressed as the means ± SEMs. n (control) = 14, n (NRD) = 14.

Figure 3

Anxiety-like behavior in NRD mice. (A) Diagram of the O-maze apparatus. (B–D) Performances of the control and NRD mice regarding the distance covered in open arms (B), time in open arms (C), and number of open arms entered (D). * p < 0.05 as determined by unpaired Student’s t-test. Data are expressed as the means ± SEMs. n (control) = 13, n (NRD) = 13.

Figure 4

Decreased struggle time in NRD mice. (A) Diagram of the forced swim test. (B) Struggle times of the control and NRD mice in the forced swim test. ** p < 0.01 as determined by unpaired Student’s t-test. Data are expressed as the means ± SEMs. n (control) = 17, n (NRD) = 17.

Figure 5

Impairment of Y-maze spontaneous alternation rate in NRD mice. (A) Diagram of the Y-maze working memory task. (B) The number of entries in all arm of the Y maze. (C) Spatial memory measured as alternation percentages in the Y maze. * p < 0.05 as determined by unpaired Student’s t-test. Data are expressed as the means ± SEMs. n (control) = 15, n (NRD) = 14.

Figure 6

Defected cognition in object recognition memory in NRD mice. (A) Diagram of the novel object recognition test. (B) Performances of the control and NRD mice in the novel object recognition test. ** p < 0.01 as determined by unpaired Student’s t-test. Data are expressed as the means ± SEMs. n (control) = 13, n (NRD) = 13.

Figure 7

Enhanced contextual fear memory in NRD mice. (A) Diagram of the contextual fear conditioning test. (B) Performance of control mice and NRD mice before and during training. **** p < 0.0001 as determined by the two-way ANOVA. n (control) = 20, n (NRD) = 20. (C) Freezing behavior tests after 1 h of training. n (control) = 20, n (NRD) = 20. (D) Freezing behavior tests after 24 h of training. n (control) = 20, n (NRD) = 18. * p < 0.05 as determined by the unpaired Student’s t-test. Data are expressed as the means ± SEMs.

Figure 8

Reduced tone-cued fear memory in NRD mice. (A) Diagram of the cued fear conditioning test. (B) Performance of NRD mice and control mice before and during training. n (control) = 16, n (NRD) = 16. (C) Freezing behavior tests after 1 h of training. n (control) = 15, n (NRD) = 15. (D) Freezing behavior tests after 24 h of training. n (control) = 15, n (NRD) = 16. * p < 0.05, as determined by unpaired Student’s t-test; **** p < 0.0001 as determined by the two-way ANOVA. Data are expressed as the means ± SEMs.

Figure 9

Both social behavior and social novelty behavior were impaired in NRD mice. (A) Diagram of the social novelty test. (B) Sniffing time of empty and stranger 1in the control and NRD mice during sociability session. * p < 0.05, *** p < 0.001, as determined by the two-way ANOVA. (C) Quantitative sociability assessment. ** p < 0.01 as determined by unpaired Student’s t-test. (D) Sniffing time of stranger 1 and stranger 2 in the control and NRD mice during social novelty session. *** p < 0.001, **** p < 0.0001, as determined by the two-way ANOVA. (E) Quantitative social novelty assessment. * p < 0.05 as determined by unpaired Student’s t-test. Data are expressed as the means ± SEMs. n (control) = 13, n (NRD) = 13.

Figure 10

Declined pain sensitivity in NRD mice. (A) Schematic of von Frey filaments test. (B) Quantitative allodynia assessment by fifty percent paw withdrawal threshold testing. **** p < 0.0001 as determined by unpaired Student’s t-test. Data are expressed as the means ± SEMs. n (control) = 14, n (NRD) = 14.