Impact of photoperiod and functional clock on male diapause in cryptochrome and pdf mutants in the linden bug Pyrrhocoris apterus.

Numerous insect species living in temperate regions survive adverse conditions, such as winter, in a state of developmental arrest. The most reliable cue for anticipating seasonal changes is the day-to-night ratio, the photoperiod. The molecular mechanism of the photoperiodic timer in insects is mostly unclear. Multiple pieces of evidence suggest the involvement of circadian clock genes, however, their role might be independent of their well-established role in the daily oscillation of the circadian clock. Furthermore, reproductive diapause is preferentially studied in females, whereas males are usually used for circadian clock research. Given the idiosyncrasies of male and female physiology, we decided to test male reproductive diapause in a strongly photoperiodic species, the linden bug Pyrrhocoris apterus. The data indicate that reproduction is not under circadian control, whereas the photoperiod strongly determines males' mating capacity. Clock mutants in pigment dispersing factor and cryptochrome-m genes are reproductive even in short photoperiod. Thus, we provide additional evidence of the participation of circadian clock genes in the photoperiodic time measurement in insects.

Light and Temperature Synchronizes Locomotor Activity in the Linden Bug, Pyrrhocoris apterus.

Circadian clocks are synchronized with the external environment by light and temperature. The effect of these cues on behavior is well-characterized in Drosophila, however, little is known about synchronization in non-model insect species. Therefore, we explored entrainment of locomotor activity by light and temperature in the linden bug Pyrrhocoris apterus (Heteroptera), an insect species with a strong seasonal response (reproductive diapause), which is triggered by both photoperiod and thermoperiod. Our results show that either light or temperature cycles are strong factors entraining P. apterus locomotor activity. Pyrrhocoris is able to be partially synchronized by cycles with temperature amplitude as small as 3°C and more than 50% of bugs is synchronized by 5°C steps. If conflicting zeitgebers are provided, light is the stronger signal. Linden bugs lack light-sensitive (Drosophila-like) cryptochrome. Notably, a high percentage of bugs is rhythmic even in constant light (LL) at intensity ∼400 lux, a condition which induces 100% arrhythmicity in Drosophila. However, the rhythmicity of bugs is still reduced in LL conditions, whereas rhythmicity remains unaffected in constant dark (DD). Interestingly, a similar phenomenon is observed after temperature cycles entrainment. Bugs released to constant thermophase and DD display weak rhythmicity, whereas strong rhythmicity is observed in bugs released to constant cryophase and DD. Our study describes the daily and circadian behavior of the linden bug as a response to photoperiodic and thermoperiodic entraining cues. Although the molecular mechanism of the circadian clock entrainment in the linden bug is virtually unknown, our study contributes to the knowledge of the insect circadian clock features beyond Drosophila research.

Circadian regulation of caterpillar feeding and growth.

Circadian clocks orchestrate many physiological processes in adult organisms. For example, rhythmic feeding behavior is regulated by the central clock in the nervous system in coordination with metabolic rhythms, which in turn depend mostly on peripheral clocks localized in many tissues. Disruption of the circadian clock leads to metabolic dysregulation both in mammals and in the model insect Drosophila melanogaster. Circadian coordination of feeding and metabolism has been studied mainly in adult insects and not in larval stages, which are dramatically different from adults in species with complete full metamorphosis. The goal of this study was to determine whether feeding and metabolism in lepidopteran larvae are subject to circadian regulation. We show that cotton leafworm caterpillars, Spodoptera littoralis, display rhythmic feeding behavior and that circadian clock genes are expressed in two peripheral tissues, the midgut and fat body. Even though both tissues display rhythmic circadian clock gene expression, the main component of the clock, per, is arrhythmic in the gut and rhythmic in the fat body. In both tissues, the presence of rhythmic physiological processes was observed, which suggested that metabolism is already driven by the circadian clock in the insect's juvenile stages.