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Behavioral, Neurobiological, and Genetic Analysis of the Circadian Mutant Duper

Abstract
The recently discovered circadian mutant hamster duper has a short period of ~23 hours and exhibits exaggerated phase shifts in response to a 15-min light pulse. To increase the understanding of the duper mutation, I performed behavioral, neurobiological, and genetic experiments. Behavioral studies using photic and non-photic stimuli found that large phase shifts exhibited by duper hamsters are specific to photic cues, but not to phase. Additionally, 2/3 of duper hamsters, but no WTs, displayed transient ultradian wheel-running patterns when transferred from light to dark at CT 18. This suggests that the mutation may weaken coupling among components of the circadian pacemaker. Anatomical and immunocytochemical analysis of the SCN was used to examine the neurobiological mechanisms of large light-induced phase shifts in dupers. Brains were collected from duper and WT hamsters at CT 12 and 15 as well as 1, 2, 3, 6 and 9 hours following a light pulse, or control handling, at CT 15. Surprisingly, the only difference in PER1 (a core clock protein) expression in the SCN between dupers and WTs was seen 2-hours after a light pulse; duper hamsters displayed a significantly greater percentage of retinorecipient VIP cells co-labeled with PER1 compared to WTs. Additional differences between genotypes occurred 9 hours after CT15 (controls). In the SCN, the number of PER1-ir cells was significantly greater in WT than duper hamsters, however this finding was reversed in the PVN. This anatomical mismatch suggests the mutation may affect signaling between the SCN and extra-SCN oscillators. Finally, to identify the genetic basis of the duper phenotype, I crossed dupers with a novel ecotype in order to perform fast homozygosity mapping. Duper transmitted onto the novel ecotype with the predicted Mendelian inheritance of phenotype. I collected DNA from F2 duper hamsters, and expect fast homozygosity mapping will identify candidate genetic regions of the duper mutation. Additional behavioral experiments in F2 dupers demonstrated that duper hamsters are resistant to jet lag. As duper is a unique circadian mutation, understanding of the behavioral phenotype, neurobiological mechanism, and genetic basis of the duper mutation will greatly increase our knowledge of the circadian system.
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