Deficient Central Leptin Signaling “In Vivo” Alters Hypothalamic Levels of Circadian Clock Proteins Changing the Diurnal Eating Behavior in Rats

Leptin, acting at CNS, decreases food intake (FI), body weight (BW) and adiposity, while circadian dysfunction induces hypothalamic leptin resistance associated to increased adiposity. Hence, we hypothesized that partial inhibition of hypothalamic leptin signaling "in vivo" would alter circadian rhythms resulting in changes in diurnal eating behaviors leading to increased adiposity and peripheral insulin resistance.3-months-old Wistar rats were randomly assigned in two different groups: rats infused "in vivo" with saline (PBS) (n=12) or rats infused with a leptin antagonist (SLA) (0.2 micrograms/day) (n=12) via icv minipump insertion for 21 days. On day 0, rats were individually housed in metabolic cages and maintained for 21 days measuring daily BW, FI and locomotor activity. Hypothalamic proteomic analysis (n=6 per group) was carried out to determine if hypothalamic circadian proteins could be altered upon chronic SLA infusion. On day 21 and before sacrifice, one group of rats infused with PBS or SLA (n=6 per group) were injected icv with 100 ng of rat leptin in a volume of 2 microliters. 30 min later rats were euthanatized.SLA infusion decreased leptin-induced phosphorylation of pY705 -STAT3 at the hypothalamus sustaining the induction of central leptin resistance upon chronic SLA administration. As a result, daily FI, final BW and visceral adiposity increased in SLA-infused rats. Remarkedly, the diurnal eating behavior was altered in the SLA group of rats. In fact, daily FI was significantly increased by 48% only during the light phase in SLA infused vs PBS treated control rats, while in the dark phase FI increased less than 10% in the SLA group compared with control PBS-infused rats. Moreover, SLA-infused rats develop peripheral insulin resistance assessed by HOMA-IR at the end of the treatment. Hypothalamic proteomic analysis revealed significant changes in 446 proteins, 266 were up-regulated and 180 down-regulated upon SLA infusion. Interestingly, proteins involved in the transcriptional activity of BMAL1/CLOCK were altered in SLA-infused rats. Paraspeckle component 1, involved in the negative regulation of BMAL1/CLOCK, was over-represented, while vasopressin-neurophysin 2 copeptin, a positive regulator of circadian gene expression, was under-represented.Our results suggest that deficient hypothalamic leptin signaling is associated to circadian dysfunction which leads to changes in the diurnal eating behavior in rats resulting in increased adiposity and peripheral insulin resistance.