Abstract
Throughout history, humans have been constantly driven by the rising and setting of the sun. Being able to anticipate this consistently changing environment is encompassed in our endogenous circadian clock. This system allows for the coordination of our physiology and behaviour to suit the environment around us. Human studies within the last few decades suggest that disruptions to this system increase the prevalence of metabolic disturbance, leading to the development of diseases such as obesity, diabetes, cardiovascular disease and their associated co-morbidities. Little is known about the mechanisms by which disruptions lead to an eventual degradation in the metabolic system.
This thesis first sought to characterise the effect that chronic light cycle disruptions have on metabolism. To emulate the human condition, I used a mouse model exposed to constantly shifting lighting environment, comprised of a 6-hour advance, occurring every 6 days, to chronically disrupt their circadian timing system. This treatment caused a gradual increase in body weight of 3.54 ± 0.34g after 12 phase shifts compared with mice under control lighting conditions gaining 1.97 ± 0.25g. Additionally, following the 5th phase shift, light cycle-disrupted animals showed a reversal in their diurnal pattern of energy homeostasis and locomotor activity followed by a subsequent loss of this rhythm. To investigate potential molecular mechanisms mediating these metabolic alterations, we assessed central leptin and insulin sensitivity. We discovered that light cycle-disrupted mice had a reduction in central leptin signalling sensitivity, as indicated by a reduction in the number of phosphorylated STAT3 immunoreactive cells in the arcuate nucleus of the hypothalamus. Furthermore, light cycle-disrupted animals exhibited a marked increase in fasting blood glucose from 108.8 ± 21.3mg/dl in control animals, to 269.4 ± 21.1mg/dl in animals under light disruption. Peripheral hyperglycaemia was accompanied by alterations in central insulin signalling at the level of pAkt and IRS1, suggesting that light cycle-disruption has a direct effect on metabolic signalling pathways within the brain.
Secondly, I sought to investigate the effect light cycle disruptions have on whole body glucoregulation. Mice were subjected to the same light cycle disruption schedule as previous, and glucose tolerance was assessed throughout the experiment. Glucose tolerance of mice under light cycle disruptions was altered from the 9th light cycle shift. Fasting blood glucose was elevated in animals under light cycle disruption (143.1 ± 4.8 mg/dl) in comparison to controls (110.2 ± 6.6 mg/dl), however, not to the same magnitude as in the previous cohort of mice. The loss in glucose tolerance is likely a direct result of the light cycle disruptions given that there was no increase in body weights above controls, in animals under light disruptive conditions.
Lastly, I aimed to assess the effect of disrupting either the light zeitgeber or the food intake zeitgeber, on metabolism. Additionally, I sought to ameliorate previously observed metabolic disturbances by intervening with strict feeding regimes. Mice were either fed ad libitum, restricted to the dark phase, or had a rotating window of food access, throughout the experiment. Of these, half the animals were subjected to light cycle disruptions, identical to previous experiments. Animals under light cycle disruption fed ad libitum gained 51 ± 5.3% increase in body weight compared to 40.7 ± 4.1% increase in controls, as well as an increase in fat mass relative to body weight. Interestingly, a third group of mice presented themselves during this study, that had a distinct phenotype difference when compared to wildtype animals. Animals deemed ‘NowShift’ were observed to shift their behaviour almost instantaneously following a shift in the lighting environment. These animals were also protected against light disruption induced weight gain, increasing by only 31.3 ± 6.2% across the experiment, in those fed ad libitum. Mice under light cycle disruption that had their food restricted to the dark phase gained only 21 ± 3.5% body weight from baseline, in comparison to controls that gained 27.9 ± 4.3% body weight from baseline. Animals in this group deemed as ‘NowShift’ were also unaffected by the lighting disruptions, gaining only 23.6 ± 3.8% body weight from baseline. Additionally, there were no differences in fat mass across all three groups. Animals with rotating access to food showed a reduced body weight in comparison to those fed ad libitum, likely due to a reduction in total food intake. These animals had a severely disrupted rhythm of food intake; however, little impact was seen in activity rhythms. Our findings suggest that light entrains and controls locomotor activity patterns, whereas food intake is a crucial entrainment cue for maintaining metabolic health. Forcing synchronisation of the food and light zeitgeber through time restricted feeding can ameliorate body weight increases observed in animals fed ad libitum. Finally, the spontaneous phenotype of NowShift animals was a surprise, however, the fact that these animals are protected from light disruption induced weight gain raises the question of the mechanism behind their genetic advantage. Exome sequencing was performed in these animals, however further investigation is required to identify potential genetic variants.