Mouse Exploratory Behaviour in the Open Field with and without NAT-1 EEG Device: Effects of MK801 and Scopolamine
Abstract
:1. Introduction
2. Methods
2.1. Experimental Design
2.2. Animals
2.3. Stereotactic Surgery and Home-Cage Recordings (Experiment 2 Only)
2.4. Drugs and Drug Formulation
2.5. Apparatus and Testing Protocols
- total distance travelled in the arena;
- line-crossings in a 16-grid overlay;
- meandering (a measurement of an animal’s absolute turn angle over the total distance travelled, which refers to the winding movement of animal);
- frequency of rotations (total of 360 degree turns to the left and 360 degree turns to the right);
- degree of thigmotaxis (a ratio of distance travelled in the periphery zone against the total distance travelled);
- average distance from the centre point (to provide meaningful indications of the general location of the mice without utilizing zonal outlines).
2.6. Exploratory Data Analyses
2.7. Statistical Analysis
3. Results
4. Discussion
Supplementary Materials
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Acknowledgments
Conflicts of Interest
References
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Experiment 1 | Experiment 2 | |
---|---|---|
Study Outcome | Pharmacological studies without EEG attachments | Pharmacological studies in animals with EEG attachments |
Persons involved (Blinder and experimenter) | 2 | 3 |
Animals | ||
Strain | C57BL/6J (Wildtype; total n = 29) | C57BL/6J (Wildtype; total n = 37; lost 10, see Section 2.3) |
Supplier/ Breeder | Charles River Laboratories, Kent, UK | Charles River Laboratories, Kent, UK |
Transport | By road | By road |
Specific pathogen free conditions | Yes | Yes |
Age at arrival and acclimatisation to facility | 10 weeks; 2 weeks of acclimatisation | 9 weeks; 2 weeks of acclimatisation |
Housing/ husbandry; test room conditions | ||
Cage type | Conventional shoe box cages with wire top feeders; single housing | Conventional shoe box cages with wire top feeders (only 1 was moved to IVC cage); single housing |
Enrichment; bedding | Cardboard rolls in all cages; corncob and aspen shavings (ratio 3:1) | No enrichment; corncob and aspen shavings (ratio 3:1) |
Temperature, humidity | 21 ± 2 °C; 50 ± 5% | 21 ± 2 °C; 50 ± 5% |
Exposure of animal to handling | Two/three times a day by facility staff | Two/three times a day by facility staff |
Noise/background noise | Background noise (music) during light phase | Background noise (music) during light phase |
Dark/light cycle | 12:12 h; start of light phase at 07:00, no red light during dark phase | 12:12 h; start of light phase at 07:00, no red light during dark phase |
Diet | Normal chow pellets (SDS CRM) and room-temperature water ad libitum | Normal chow pellets (SDS CRM) and room-temperature water ad libitum |
Handling of cages | Cleaning once every 2 weeks; water and food changed/topped up twice a week. | Cleaning once every 2 weeks; water and food changed/topped up twice a week. |
Experimental: animals | ||
Age of animal at start to experiment | 12–13 weeks (at the start of testing) | 10–11 weeks (at surgery) and 12–13 weeks (at the start of testing) |
Transport of animals in facility | Silent trolley type; cages were not blacked out or covered during transport | Silent trolley type; cages were not blacked out or covered during transport |
Time of testing, day of week | Commencement at 10:00; testing was conducted from Monday to Thursday on the first week, and Monday to Wednesday on the second week. | Commencement at 10:00; testing was conducted from Monday to Thursday on the first week, and Monday to Wednesday on the second week. |
Total duration of test phase per day | Approx. 5 h (4 animals tested am, 3 tested pm) | Approx. 5 h (3 animals tested am, 3 tested pm) |
Animal under diet restrictions during experimental phase | No | No, except during NAT-1 application and drug administration: water bottles were removed, and cage lids were overturned to prevent NAT devices from accidentally wedging onto cage-lids and food-hoppers |
* Naivety to experimental paradigm and drugs | Naïve to both experimental procedures and drugs | Home cage EEG recording and check, 24 h prior to open field test; naïve to drugs |
Experimental: surgical protocols | ||
* Time period between surgeries and start of experiment | N.A. | 2 weeks |
* Total duration of surgery per day | N.A. | Approx. 7 h/day (commencement at 08.00; surgeries performed only on Mondays and Tuesdays |
* Anaesthetic | N.A. | Gas anaesthesia (isoflurane) 1.0 mL saline (intraperitoneal) and 0.2 mL of 0.1 mg/kg buprenorphine (subcutaneous; Vetergesic®, Cera Animal Health Ltd., High Wycombe, UK). |
* Analgesia | N.A. | Drinking water was supplemented with Rimadyl® (Zoetis UK Ltd., Surrey, UK) for 2 days before and after the surgery |
* Surgical procedure | N.A. | 1 surgeon; 2 assistants |
Experimental: design and methods/data analysis | ||
Drug, and concentration | 0.9% saline; 0.65 mg/kg MK801; 0.85 mg/kg scopolamine. Intra-peritoneal administration of 10 mg/kg of treatment. | 0.9% saline; 0.65 mg/kg MK801; 0.85 mg/kg scopolamine. Intra-peritoneal administration of 10 mg/kg of treatment. |
* Experimenter | 1 female | 2 females and 1 male |
* Frequency of handling | Handling during drug injections, immediately before and after test. | Handling during NAT-1 applications, drug injections, immediately before and after test. |
Randomisation method for allocation of animals to test groups | random.org (https://www.random.org/sequences/, accessed 29 October 2018; Williams square design. | random.org (https://www.random.org/sequences/, accessed 19 February 2018); Williams square design. |
Blinding of experimenter to group allocation | Yes. | Yes. |
Standardisation of animal handling across experimenters (protocol and training) | Yes (1 in randomisation and blinding, 1 in drug administration and experiment) | Yes (2 involved in NAT-1 device application and drug administration and randomisation, 1 in experiment) |
Software used in recording and data extraction | Automatic tracking software, ANY-Maze (v.5.1) | Automatic tracking software, ANY-Maze (v.5.1) |
Inclusion and exclusion criteria | No inclusion criteria. Exclusion criteria based on analysis using Grubb’s method before re-evaluation of animal behaviour (video). | No inclusion criteria. Exclusion criteria based on analysis using Grubb’s method before re-evaluation of animal behaviour (video). |
Statistical analysis | Bootstrapped one-way ANOVA and t-test for conventional analysis; repeated measures ANOVA for binned analysis; and estimation statistics to determine the magnitude of differences. | Bootstrapped one-way ANOVA and t-test for conventional analysis; repeated measures ANOVA for binned analysis; and estimation statistics to determine the magnitude of differences. |
Experimental: complexity of test arena | ||
Test arena | Square; 50 × 50 × 50 cm; white Perspex | Square; 50 × 50 × 50 cm; white Perspex |
Average light intensity in test arena | 320 lux | 320 lux |
Environment in test room (temperature, humidity and decibels relative to the carrier) | 20 ± 2 °C; 48 ± 5%; 30 ± 5 dBc; dedicted sound-attenuated room | 20 ± 2 °C; 48 ± 5%; 30 ± 5 dBc; dedicted sound-attenuated room |
Number of test arenas used at any given time | 1 | 1 |
Cleaning agents | Non-fragrant and alcohol-free wet wipes | Non-fragrant and alcohol-free wet wipes |
Parameters | Drug | Between-Experiment Comparisons | E2 minus E1 | ||
---|---|---|---|---|---|
E1 | E2 | p-Value | Mean Difference [95% CI] | ||
Mean ± SD | Mean ± SD | ||||
Distance moved (m) | Saline | 131.6 ± 20.2 | 139.9 ± 19.2 | 0.347 | 8.4 [−8.1; 24.6] |
MK801 | 70.0 ± 25.6 | 39.3 ± 23.5 | 0.028 | −30.6 [−48.6; −6.0] | |
Scopolamine | 182.2 ± 20.0 | 155.2 ± 26.7 | 0.048 | −26.9 [−52.3; −10.4] | |
Average distance from centre-point (cm) | Saline | 0.218 ± 0.007 | 0.199 ± 0.010 | 0.001 | −0.020 [−0.026; −0.012] |
MK801 | 0.155 ± 0.030 | 0.161 ± 0.050 | 0.782 | 0.006 [−0.041; 0.036] | |
Scopolamine | 0.237 ± 0.013 | 0.230 ± 0.012 | 0.275 | −0.008 [−0.018; 0.003] | |
Thigmotaxis (ratio) | Saline | 0.532 ± 0.052 | 0.463 ± 0.049 | 0.020 | −0.068 [−0.11; −0.021] |
MK801 | 0.223 ± 0.157 | 0.285 ± 0.189 | 0.471 | 0.061 [−0.1; 0.207] | |
Scopolamine | 0.807 ± 0.147 | 0.777 ± 0.128 | 0.667 | −0.030 [−0.131; 0.115] | |
Thigmotaxis (s) | Saline | 1266.1 ± 96.3 | 1050.0 ± 105.7 | 0.002 | −216 [−295; −123] |
MK801 | 545.1 ± 379.3 | 699.2 ± 445.6 | 0.435 | 154 [−199; 527] | |
Scopolamine | 1539.8 ± 184.5 | 1475.8 ± 163.8 | 0.437 | −64 [−192; 107] | |
Line-crossings | Saline | 1060.6 ± 145.3 | 1030.0 ± 132.49 | 0.442 | −28.6 [−148; 86.7] |
MK801 | 866.9 ± 265.1 | 458.3 ± 238.5 | 0.014 | −409 [−600; −158] | |
Scopolamine | 1374.2 ± 154.3 | 1012.0 ± 196.9 | 0.003 | −362 [−541; −242] | |
Meandering (degrees/s) | Saline | 912.5 ± 89.5 | 932.0 ± 107.4 | 0.668 | 19.5 [−63.6; 102] |
MK801 | 2716.4 ± 1008.4 | 3077 ± 1813.4 | 0.603 | 361 [−663.0; 2100] | |
Scopolamine | 705.6 ± 73.5 | 750.4 ± 89.2 | 0.325 | 44.7 [−25.6; 117] | |
Rotation | Saline | 76.5 ± 10.5 | 98.8 ± 12.5 | 0.006 | 16.3 [5.6; 26] |
MK801 | 180.1 ± 142.7 | 158.2 ± 138.3 | 0.758 | −21.9 [−156.0; 87.5] | |
Scopolamine | 99.8 ± 10.6 | 96.1 ± 18.2 | 0.623 | −3.7 [−18.9; 7.16] |
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Lim, C.J.M.; Bray, J.; Janhunen, S.K.; Platt, B.; Riedel, G. Mouse Exploratory Behaviour in the Open Field with and without NAT-1 EEG Device: Effects of MK801 and Scopolamine. Biomolecules 2024, 14, 1008. https://doi.org/10.3390/biom14081008
Lim CJM, Bray J, Janhunen SK, Platt B, Riedel G. Mouse Exploratory Behaviour in the Open Field with and without NAT-1 EEG Device: Effects of MK801 and Scopolamine. Biomolecules. 2024; 14(8):1008. https://doi.org/10.3390/biom14081008
Chicago/Turabian StyleLim, Charmaine J. M., Jack Bray, Sanna K. Janhunen, Bettina Platt, and Gernot Riedel. 2024. "Mouse Exploratory Behaviour in the Open Field with and without NAT-1 EEG Device: Effects of MK801 and Scopolamine" Biomolecules 14, no. 8: 1008. https://doi.org/10.3390/biom14081008