Odor and Spatial Working Memory in an Environmentally-Induced Animal Model of Parkinsonism

1. Odor and Spatial Working Memory in an Environmentally-Induced Animal Model of Parkinsonism Christina M. Parr & Aileen Bailey, Ph.D. Department of Psychology, St. Mary’s College of Maryland Results – Y-maze Task Results – Social Odor Task Introduction Parkinson’s disease (PD) is a neurodegenerative disorder of the central nervous system currently affecting approximately 500,000 people in the United States6. Clinically, PD is characterized by resting tremor, bradykinesia, rigidity, and postural instability1. PD is a chronic and progressive disease characterized neuropathologically by degeneration of the dopaminergic neurons and clumps of the protein α-synuclein2,4. Cycad seed is an environmental neurotoxin known to induce ALS/PDC-like symptoms among the Chomorro population in Guam and PD-like symptoms in rat models3. The cycad seed model induces PD progressively which makes this animal model more similar to the human pathogenesis of the disease than other animal models7. Research has found that cognitive deficits occur before the onset of motor deficits in PD8. The visuo-spatial deficits and working memory deficits seem to be the most studied cognitive deficits in these animal models since they are consistently reported in humans with PD2. The cycad model may allow researchers to see when and which deficits are occurring and where in the brain they cause abnormalities. Previous research has shown spatial information and processing deficits in a mouse 6-OHDA lesion model2 and social odor recognition deficits using an MPTP mouse model of Parkinson’s disease8. The following research investigated spatial and odor working memory deficits occurring in rats fed washed cycad seed. It was hypothesized that cycad fed rats would show spatial working memory deficits as measured by the Y-maze and social memory deficits as measured by a social odor recognition task. Figure 1. Average time (sec) spent locomoting± 1 SD between the first day and the second day of testing between the cycad low dose animals, the cycad high dose animals, and the control animals in the social odor recognition task. Figure 5. Mean proportion time spent in the novel arm ± 1 SD during the 3 min testing period comparing cycad low dose animals, the cycad high dose animals, and the control animals in the Y-maze. Figure 2. Mean proportion time ± 1 SD spent exploring the first novel scent (now no longer novel) and the second novel scent on the second day of testing between the cycad low dose animals, the cycad high dose animals, and the control animals in the social odor recognition task. Figure 6. Mean number of entries ± 1 SD into the novel arm between the low dose cycad animals, the high dose cycad animals, and the control animals during the 3 min testing period in the Y-maze. Methods • Subjects • 32 adult, male, Sprague Dawley rats were used • The high dose cycad rats (n=11) were given washed cycad seed as 20% of their diet which correlated to 5 g of cycad/day/rat • The low dose cycad rats (n=13) were given washed cycad seed as 5% of their diet which correlated to 1.25 g of cycad/day/rat. • The control rats (n=8) were given a daily pellet made from all-purpose white flour and flavored with vanilla or maple extract • Each animal was given 1 pellet per day Monday-Friday only in addition to their regular rodent chow and consumption of the pellet was observed to ensure the pellet was completely eaten • Procedures • Y-maze task. Testing novelty and spatial memory, the Y-maze is in the shape of a “Y” 40 cm (L) x 33 cm (L) x 21 cm (H) x 10 cm (W) and the arms are 10 cm in width and 3 black and white geometric cue signs surrounding it. A total of 16 rats (2 control rats, 3 low dose cycad rats and 11 high dose cycad rats) were used. During the exposure phase the rat was placed in one arm and another arm was blocked. The rats were allowed to explore the two arms for five minutes. Number of entries and time spent in each arm were recorded. After 5 min the animal was removed from the maze and placed in its home cage for a 5 min delay. The rat was placed back into the Y-maze for a two min test phase. All three arms were open and time spent in and number of entries into each arm was recorded. Statistical analyses were performed. • Social odor recognition task. Testing odor and social working memory, this task used 29 rats (8 control, 13 low dose cycad rats, and 8 high dose cycad rats). Three wooden shims were placed in the rat’s home cage for 24 hours prior to testing. On the first day each animal was placed into a 60 x 60 x 28 cm open field for 3 min with a wooden shim of their own (F) and a wooden shim that was with another rat (N1). Time spent exploring each of the shims, time spent locomoting, number of outer squares crossed, number of inner squares crossed, number fecal boli, number of rears, and number of grooming sessions were counted and recorded. Twenty four hours later the same rats were placed in the open field again, but this time with 3 wooden shims; the two from the previous day (F and N1) and another shim saturated with a third rat’s scent (N2). All of the same measures as the previous day were recorded. Conclusions • There was a significant decrease in the amount of time spent locomoting over the first day to the second day, F(1,25) = 26.26, p < 0.001. The significant decrease occurred between the control rats, t(7) = 6.217, p < 0.001, and the cycad low dose rats t(12 )= 3.069, p = 0.01, but not the cycad high dose rats. • This shows that the cycad high dose rats remain highly active. This suggests that the high dose cycad rats are not habituating to the open field suggesting a possible deficit or abnormality. • There are no significant differences in the time spent exploring the first novel scent (N1) on the first day. • There are no significant differences in the time spent exploring the first novel scent or the second novel scent on the second testing day. • There are significantly less entries into the inner squares compared to the outer squares on both the first day, F(1,26) = 189.6, p < 0.001, and the second day of testing, F(1,26) = 167.8, p < 0.001 in all groups of rats. • There was no significant difference between number of outer vs. inner squares crossed between the control rats, the low dose cycad rats, and the high dose cycad rats, F(2,26) = 1.31, p > 0.05. • There are no significant differences in time spent in the novel arm or number of entries into the novel arm during the testing phase in the Y-maze task. • Although not significant, there is a pattern for the low dose cycad rats in both the social odor recognition task (N2) and the Y-maze task to perform at lower levels. • The large variability found in the tasks could mean that there may be neurochemical differences, specifically α-synuclein aggregates also known as Lewy bodies, within the groups in the rats individually that could correlate with the individual behavioral differences. Figure 3. Mean number of entries ± 1 SD into the inner or outer squares in the open field apparatus on the first day of testing between the cycad low dose animals, the cycad high dose animals, and the control animals in the social odor recognition task. Figure 4. Mean number of entries ± 1 SD into the inner or outer squares in the open field apparatus on the second day of testing between the cycad low dose animals, the cycad high dose animals, and the control animals in the social odor recognition task.. References 1 Bisaglia, M., Greggio, E., Maric, D., Miller, D., Cookson, M., & Bubacco, L. (2010). α-Synucleinoverexpression increases dopamine toxicity in BE(2)-M17 cells. BMC Neuroscience, 1141-46. doi:10.1186/1471-2202-11-41. 2 De Leonibus, E., Pascucci, T., Lopez, S., Oliverio, A., Amalric, M., & Mele, A. (2007). Spatial deficits in a mouse model of Parkinson disease. Psychopharmacology, 194(4), 517-525. 3 Ince, P., & Codd, G. (2005). Return of the cycad hypothesis – does the amyotrophic lateral sclerosis/parkinsonism dementia complex (ALS/PDC) of Guam have new implications for global health?. Neuropathology & Applied Neurobiology, 31(4), 345-353. doi:10.1111/j.1365-2990.2005.00686.x. 4 Kövari, E., Gold, G., Herrmann, F., Canuto, A., Hof, P., Bouras, C., et al. (2003). Lewy body densities in the entorhinal and anterior cingulate cortex predict cognitive deficits in Parkinson's disease. ActaNeuropathologica, 106(1), 83-88. Retrieved from Academic Search Premier database. 5 Monaghan, M., Leddy, L., Sung, M., Albinson, K., Kubek, K., Pangalos, M., et al. (2010). Social odor recognition: a novel behavioral model for cognitive dysfunction in Parkinson's disease. Neuro-Degenerative Diseases, 7(1-3), 153-159. 6 Parkinson’s Disease: Hope Through Research. (2010). In National Institutes of Neurological Disorder and Stroke. Retrieved from: http://www.ninds.nih.gov/disorders/parkinsons_disease/detail_parkinsons_disease.htm#toc. 7 Schober, A. (2004). Classic toxin-induced animal models of Parkinson’s disease: 6-OHDA and MPTP. Cell & Tissue Research, 318(1), 215-224. doi:10.1007/s00441-004-0938-y. 8 Shen, W., McDowell, K., Siebert, A., Clark, S., Dugger, N., Valentino, K., et al. (2010). Environmental neurotoxin-induced progressive model of Parkinsonism in rats. Annals of Neurology, 68(1), 70-80.  Acknowledgements I would like to extend a special thank you to Dr. Bailey, Jennifer St. Germain, and Angie Draheim for their help and support throughout my project. I would also like to thank Kimberly McDowell and Dr. Paul Yarowsky from University of Maryland – Baltimore, School of Medicine for their support and their help with learning specific protocols and techniques used in this ongoing project.