Cross-talk between the fat body and brain regulates insect developmental arrest

Abstract

Developmental arrest, a critical component of the life cycle in animals as diverse as nematodes (dauer state), insects (diapause), and vertebrates (hibernation), results in dramatic depression of the metabolic rate and a profound extension in longevity. Although many details of the hormonal systems controlling developmental arrest are well-known, we know little about the interactions between metabolic events and the hormones controlling the arrested state. Here, we show that diapause is regulated by an interplay between blood-borne metabolites and regulatory centers within the brain. Gene expression in the fat body, the insect equivalent of the liver, is strongly suppressed during diapause, resulting in low levels of tricarboxylic acid (TCA) intermediates circulating within the blood, and at diapause termination, the fat body becomes activated, releasing an abundance of TCA intermediates that act on the brain to stimulate synthesis of regulatory peptides that prompt production of the insect growth hormone ecdysone. This model is supported by our success in breaking diapause by injecting a mixture of TCA intermediates and upstream metabolites. The results underscore the importance of cross-talk between the brain and fat body as a regulator of diapause and suggest that the TCA cycle may be a checkpoint for regulating different forms of animal dormancy.

Fig. 1.

COX activity and ATP content in the (A) brain and (B) fat body of nondiapausing pupae (ND), diapausing pupae (D), and diapausing pupae injected with 1 μg 20-hydroxyecdysone (20E). Numbers represent hours after injection of 20E or distilled water (H₂O). Control, no injection. Bars represent mean ± SD of three replicates. *P < 0.05; **P < 0.01 (determined by one-way ANOVA).

Fig. 2.

Differences in metabolic intermediates present in the hemolymph of (A) nondiapausing and diapausing pupae and (B) after diapause termination in response to an injection of 20E. Metabolic intermediates, including trehalose, glucose, pyruvate, and four substances in the TCA cycle, were compared: day 3 nondiapause pupae (ND), day 21 diapausing pupae (D), and day 21 diapausing pupae injected with 1 μg 20E. Relative ratio is the ratio of the peak area of metabolic intermediate to the peak area of the internal standard (sucrose). Hemolymphs from 10 pupae were mixed as one sample; bars represent the mean ± SD of four replicates. *P < 0.05; **P < 0.01 (determined by one-way ANOVA).

Fig. 3.

Manipulations of development with a nonusable glucose derivative and a metabolic mixture and a regulatory scheme defining documented interactions. (A) Developmental delay caused by DOG injection. Nondiapause pupae were kept at 20 °C for 1 d after pupation, injected with a 5 μL solution of DOG, and incubated at 20 °C. The developmental delay was determined by examining the location of the pupal stemmata on different days after injection. No treatment, n = 40; distilled water (H₂O), n = 29; 0.2 mM DOG, n = 31; 2 mM DOG, n = 30. (B) Change in PTTH mRNA expression after injection of a metabolite mixture into 21-d-old diapausing pupae. The 30× mixture is described in Tables 2 and 3. Bars represent the mean ± SD of three replicates. *P < 0.05 (determined by one-way ANOVA). (C) A schematic representation showing the action of metabolites on the brain, prothoracic glands (PGs), and fat body in the regulation of (a) nondiapause and (b) diapause. Black lines indicate demonstrated pathways (pathway 1), and red lines (pathways 2 and 3) indicate results from this study. Arrows and broken arrows indicate activation and no activation, respectively.