Mitochondrial LETM1 drives ionic and molecular clock rhythms in circadian pacemaker neurons

The mechanisms that generate robust ionic oscillation in circadian pacemaker neurons are under investigation. Here, we demonstrate critical functions of the mitochondrial cation antiporter leucine zipper- EF-hand-containing transmembrane protein 1 (LETM1), which exchanges K+/H+ in Drosophila and Ca2+/H+ in mammals, in circadian pacemaker neurons. Letm1 knockdown in Drosophila pacemaker neurons reduced circadian cytosolic H+ rhythms and prolonged nuclear PERIOD/TIMELESS expression rhythms and locomotor activity rhythms. In rat pacemaker neurons in the hypothalamic suprachiasmatic nucleus (SCN), circadian rhythms in cytosolic Ca2+ and Bmal1 transcription were dampened by Letm1 knockdown. Mitochondrial Ca2+ uptake peaks late during the day were also observed in rat SCN neurons following photolytic elevation of cytosolic Ca2+. Since cation transport by LETM1 is coupled to mitochondrial energy synthesis, we propose that LETM1 integrates metabolic, ionic, and molecular clock rhythms in the central clock system in both invertebrates and vertebrates

Histamine Regulates Molecular Clock Oscillations in Human Retinal Pigment Epithelial Cells via H1 Receptors

Vertebrate eyes are known to contain circadian clocks, but their regulatory mechanisms remain largely unknown. To address this, we used a cell line from human retinal pigment epithelium (hRPE-YC) with stable coexpression of reporters for molecular clock oscillations (Bmal1-luciferase) and intracellular Ca2+ concentrations (YC3.6). We observed concentration-dependent increases in cytosolic Ca2+ concentrations after treatment with histamine (1–100 µM) and complete suppression of histamine-induced Ca2+ mobilizations by H1 histamine receptor (H1R) antagonist d-chlorpheniramine (d-CPA) in hRPE-YC cells. Consistently, real-time RT-PCR assays revealed that H1R showed the highest expression among the four subtypes (H1–H4) of histamine receptors in hRPE-YC cells. Stimulation of hRPE-YC cells with histamine transiently increased nuclear localization of phosphorylated Ca2+/cAMP-response element-binding protein that regulates clock gene transcriptions. Administration of histamine also shifted the Bmal1-luciferase rhythms with a type-1 phase-response curve, similar to previous results with carbachol stimulations. Treatment of hRPE-YC cells with d-CPA or with more specific H1R antagonist, ketotifen, blocked the histamine-induced phase shifts. Furthermore, an H2 histamine receptor agonist, amthamine, had little effect on the Bmal1-luciferase rhythms. Although the function of the in vivo histaminergic system within the eye remains obscure, the present results suggest histaminergic control of the molecular clock via H1R in retinal pigment epithelial cells. Also, since d-CPA and ketotifen have been widely used (e.g., to treat allergy and inflammation) in our daily life and thus raise a possible cause for circadian rhythm disorders by improper use of antihistamines