Clock gene expression and locomotor activity predict death in the last days of life in Drosophila melanogaster

Abstract

The importance of the circadian clock for the regulation of behaviour and physiology, and the molecular control of these rhythms by a set of clock genes are well defined. The circadian clock deteriorates with advancing age but the mechanism underlying is unclear. Here we recorded the expression of two key clock genes in young, middle-aged and old Drosophila using transgenic luciferase lines reporting period and timeless in vivo. We report a novel marker of imminent death in the expression of TIMELESS. In the days immediately preceding death TIMELESS expression increased to at least 150% of previous acrophase values (88.0% of n = 217) and lost circadian rhythmicity, which predicted death equally well in flies of different ages and under light and temperature cycles. We suggest this transient aberrant clock-gene expression is central to the mechanism of the disturbance in circadian behaviour before death (82.7% of n = 342). We also find that PERIOD expression in central-clock neurons remained robust with age, however PERIOD and TIMELESS in peripheral clocks showed a reduction in both expression level and rhythmicity. In conclusion, as flies age the molecular clock gradually declines at the peripheral level but continues to function at the central until days before death.

The Development and Decay of the Circadian Clock in Drosophila melanogaster

“…From the existing results of Drosophila studies, a general impression is that the molecular machinery in the central clock is less or later damaged, if at all, by the aging process. We find that the expression of clock genes in the central clock neurons of aged Drosophila stays as robust as in young ones [ 49 ]. As a consequence, causal links between clock gene expression patterns in the central clock and age-associated disturbance in circadian rhythms cannot be conclusively demonstrated.…”

“…Further, the changes in peripheral oscillator behavior may not only result from intrinsic alterations within the tissues themselves, but could also be a consequence of the reduction of their responsiveness to internal or external temporal cues. Our result shows the inefficiency of strong light and temperature entrainments on rescuing the age-related deterioration of the peripheral oscillators [ 49 ]. This indicates that the impairment of the peripheral input pathway may be a possible mechanism involved in the aging process.…”

“…For instance, it has been reported that PER and TIM oscillations in almost all central clock neuron groups (s-LN v , l-LN v , DN 1 , DN 2 , and LN d ) deteriorate with advancing age [ 44 ]. However, this is challenged by the finding that PER is robustly expressed in synchrony among different groups of clock neurons (s-LN v , DN 1 and LN d [ 48 ], and DN 1-3 and LN d [ 49 ]) in aged flies.…”

“…Reduction in amplitude of circadian clock gene expression has been found throughout the body in aged animals [ 50 ]. Likewise, per and tim luciferase reporters have shown a decline in expression of peripheral clocks throughout the fly body with advancing age [ 48 , 49 ]. Further, per mRNA oscillations [ 51 ], as well as per and tim at both mRNA and protein levels [ 48 , 52 ], have been reported to decrease in the heads of old flies.…”

“…Importantly, this marker in the expression of tim was not age dependent and predicted death equally well in fruit flies of different ages and under light and temperature cycles. It is considered the central mechanism of the disturbance in circadian behavior before death [ 49 ].…”