SignificanceShift workers, whose schedules are misaligned relative to their suprachiasmatic nuclei (SCN) circadian pacemaker, are at elevated risk of metabolic disorders. In a study of simulated day- versus night-shift work followed by a constant routine, we separated plasma-circulating metabolites according to whether their 24-h rhythms aligned with the central SCN pacemaker or instead reflected externally imposed behavioral schedules. We found that rhythms in many metabolites implicated in food metabolism dissociated from the SCN pacemaker rhythm, with the vast majority aligning with the preceding sleep/wake and feeding/fasting cycles. Our metabolomics study yields insight into the link between prolonged exposure to shift work and the spectrum of associated metabolic disorders by providing a window into peripheral oscillators and the biobehavioral factors that orchestrate them.

Parkinson’s disease (PD) is a chronic disorder that presents a range of premotor signs, such as sleep disturbances and cognitive decline, which are key non-motor features of the disease. Increasing evidence of a possible association between sleep disruption and the neurodegenerative process suggests that sleep impairment could produce a detectable metabolic signature on the disease. In order to integrate neurocognitive and metabolic parameters, we performed untargeted and targeted metabolic profiling of the rotenone PD model in a chronic sleep restriction (SR) (6 h/day for 21 days) condition. We found that SR combined with PD altered several behavioural (reversal of locomotor activity impairment; cognitive impairment; delay of rest-activity rhythm) and metabolic parameters (branched-chain amino acids, tryptophan pathway, phenylalanine, and lipoproteins, pointing to mitochondrial impairment). If combined, our results bring a plethora of parameters that represents reliable early-phase PD biomarkers which can easily be measured and could be translated to human studies.

Patients with liver cirrhosis can develop hyperammonemia and hepatic encephalopathy (HE), accompanied by pronounced daytime sleepiness. Previous studies with healthy volunteers show that experimental increase in blood ammonium levels increases sleepiness and slows the waking electroencephalogram. As ammonium increases adenosine levels in vitro, and adenosine is a known regulator of sleep/wake homeostasis, we hypothesized that the sleepiness-inducing effect of ammonium is mediated by adenosine. Eight adult male Wistar rats were fed with an ammonium-enriched diet for 4 weeks; eight rats on standard diet served as controls. Each animal was implanted with electroencephalography/electromyography (EEG/EMG) electrodes and a microdialysis probe. Sleep EEG recording and cerebral microdialysis were carried out at baseline and after 6 h of sleep deprivation. Adenosine and metabolite levels were measured by high-performance liquid chromatography (HPLC) and targeted LC/MS metabolomics, respectively. Baseline adenosine and metabolite levels (12 of 16 amino acids, taurine, t4-hydroxy-proline, and acetylcarnitine) were lower in hyperammonemic animals, while putrescine was higher. After sleep deprivation, hyperammonemic animals exhibited a larger increase in adenosine levels, and a number of metabolites showed a different time-course in the two groups. In both groups the recovery period was characterized by a significant decrease in wakefulness/increase in NREM and REM sleep. However, while control animals exhibited a gradual compensatory effect, hyperammonemic animals showed a significantly shorter recovery phase. In conclusion, the adenosine/metabolite/EEG response to sleep deprivation was modulated by hyperammonemia, suggesting that ammonia affects homeostatic sleep regulation and its metabolic correlates.

Metabolic rhythms include rapid, ultradian (hourly) dynamics, but it remains unclear what their relationship to circadian metabolic rhythms is, and what role meal timing plays in coordinating these ultradian rhythms in metabolism. Here, we characterized widespread ultradian rhythms under ad libitum feeding conditions in the plasma metabolome of the vole, the gold standard animal model for behavioral ultradian rhythms, naturally expressing ~2‐h foraging rhythms throughout the day and night. These ultradian metabolite rhythms co‐expressed with diurnal 24‐h rhythms in the same metabolites and did not align with food intake patterns. Specifically, under light–dark entrained conditions we showed twice daily entrainment of phase and period of ultradian behavioral rhythms associated with phase adjustment of the ultradian cycle around the light–dark and dark–light transitions. These ultradian activity patterns also drove an ultradian feeding pattern. We used a unique approach to map this behavioral activity/feeding status to high temporal resolution (every 90 min) measures of plasma metabolite profiles across the 24‐h light–dark cycle. A total of 148 known metabolites were detected in vole plasma. Supervised, discriminant analysis did not group metabolite concentration by feeding status, instead, unsupervised clustering of metabolite time courses revealed clusters of metabolites that exhibited significant ultradian rhythms with periods different from the feeding cycle. Two clusters with dissimilar ultradian dynamics, one lipid‐enriched (period = 3.4 h) and one amino acid‐enriched (period = 4.1 h), both showed co‐expression with diurnal cycles. A third cluster solely comprised of glycerophospholipids (specifically ether‐linked phosphatidylcholines) expressed an 11.9 h ultradian rhythm without co‐expressed diurnal rhythmicity. Our findings show coordinated co‐expression of diurnal metabolic rhythms with rapid dynamics in feeding and metabolism. These findings reveal that ultradian rhythms are integral to biological timing of metabolic regulation, and will be important in interpreting the impact of circadian desynchrony and meal timing on metabolic rhythms.

Purpose: Chondrocytes rely primarily on glycolysis to meet their energy requirements, but possess the metabolic flexibility to support cell survival and matrix synthesis during periods of nutrient stress, by enhancing glycolysis with mitochondrial respiration. Accessing this 'spare respiratory capacity' requires optimal mitochondrial function, but since impaired mitochondrial function is implicated in osteoarthritis (OA), this mechanism may be deficient or attenuated in joint disease. Metabolic adaptation is evident in early-stage OA, however cartilage from late-stage disease does not seem to have this flexibility. A deeper understanding of these complex metabolic pathways may identify new metabolic markers of disease stage, and support therapeutic strategies for treating OA. Metabolomics has the potential to identify underlying metabolic changes, reveal pathological pathways, novel biomarkers and therapeutic targets. The aim of this study was to identify metabolic processes involved in early stage disease by analysis of metabolites and metabolic function in two inflammatory models of cartilage degradation. Methods: Macroscopically normal articular cartilage was obtained from equine and bovine metacarpophalangeal joints. Six-millimetre diameter equine cartilage explants (n¼6) and isolated primary equine chondrocytes (n¼4), seeded at high density (105,000/cm 2), were cultured for 7 days in serum-free low or high glucose Glutamax DMEM (Gibco), respectively, with or without 10ng/ml equine interleukin-1b (IL-1b) and 10ng/ml equine tumour necrosis factor-a (TNF-a). Spent media (secretome) was collected and subjected to metabolomic analysis. Secretome metabolite levels were measured using the AbsoluteIDQ® p180 targeted metabolomics kit (Biocrates), and a Waters Xevo TQ-S mass spectrometer coupled to an Acquity UPLC system. Multivariate analysis was performed by principal component analysis and orthogonalized partial least squares discriminant analysis using SIMCA-P v12.0 software (Umetrics). Metabolic function of primary equine (n¼9) and bovine chondrocytes (n¼3) was determined using Seahorse XFp and XFe24 analyzers (Agilent). Cells were seeded at high density, treated with species-specific 10ng/ml IL-1b and/or 10ng/ml TNF-a for 18 h, and metabolically challenged with the Mito Stress Test during analysis. Metabolite levels, and oxygen consumption rates, were normalised to total cell protein, and differences between means determined by one-way analysis of variance with Tukey's multiple comparison post-tests. Results: Secretome metabolites which decreased with pro-inflammatory cytokine treatment were proline, ornithine and alpha-aminoadipic acid (p<0.0001). Citrulline increased with cytokine treatment (p<0.0001) and glutamate, present in DMEM, was elevated with cytokine treatment (p<0.0001). Metabolomic analysis of the chondrocyte secretome showed that glutamine decreased (p<0.02) with cytokine treatment whereas citrulline was elevated (p<0.003). Metabolic analysis showed that combined cytokine treatment reduced basal re...