The aim of this study was to examine the occurrence of endogenous oscillations of Per1, Per2, Bmal1 and Rev-erb α genes in rat pineal explants and to investigate their regulation by adrenergic ligands. Our results show a significant and sustained rhythm of Per2,Bmal1 and Rev-erb α gene expression for up to 48 h in cultured pineal gland with a pattern similar to that observed in vivo. By contrast, the rhythms of Per1 and Aa-nat, the rate-limiting enzyme for melatonin synthesis, were strongly attenuated after 24 h in culture. Addition of the exogenous adrenergic agonist isoproterenol on cultured pineal glands induced a short-term increase in mRNA levels of Per1 and Aa-nat, but not those of Per2,Bmal1 and Rev-erb α. This study demonstrates that the rat pineal gland hosts a circadian oscillator as evidenced by the sustained, noradrenergic-independent, endogenous oscillations of Per2, Bmal1 and Rev-erb α mRNA levels in cultured tissues. Only expression of Per1 was stimulated by adrenergic ligands suggesting that, in vivo, the adrenergic input could synchronize the pineal clock by acting selectively on Per1.

1.
Ko CH, Takahashi JS: Molecular components of the mammalian circadian clock. Hum Mol Genet 2006;15(spec No. 2):R271–R277.
2.
Dardente H, Cermakian N: Molecular circadian rhythms in central and peripheral clocks in mammals. Chronobiol Int 2007;24:195–213.
3.
Harms E, Kivimae S, Young MW, Saez L: Posttranscriptional and posttranslational regulation of clock genes. J Biol Rhythms 2004;19:361–373.
4.
Cuninkova L, Brown SA: Peripheral circadian oscillators: interesting mechanisms and powerful tools. Ann NY Acad Sci 2008;1129:358–370.
5.
Fukuhara C, Tosini G: Peripheral circadian oscillators and their rhythmic regulation. Front Biosci 2003;8:d642–d651.
6.
Yamazaki S, Numano R, Abe M, Hida A, Takahashi R, Ueda M, Block GD, Sakaki Y, Menaker M, Tei H: Resetting central and peripheral circadian oscillators in transgenic rats. Science 2000;288:682–685.
7.
Yamazaki S, Yoshikawa T, Biscoe EW, Numano R, Gallaspy LM, Soulsby S, Papadimas E, Pezuk P, Doyle SE, Tei H, Sakaki Y, Block GD, Menaker M: Ontogeny of circadian organization in the rat. J Biol Rhythms 2009;24:55–63.
8.
Yoo SH, Yamazaki S, Lowrey PL, Shimomura K, Ko CH, Buhr ED, Siepka SM, Hong HK, Oh WJ, Yoo OJ, Menaker M, Takahashi JS: PERIOD2::LUCIFERASE real-time reporting of circadian dynamics reveals persistent circadian oscillations in mouse peripheral tissues. Proc Natl Acad Sci USA 2004;101:5339–5346.
9.
Nishide SY, Honma S, Nakajima Y, Ikeda M, Baba K, Ohmiya Y, Honma K: New reporter system for Per1 and Bmal1 expressions revealed self-sustained circadian rhythms in peripheral tissues. Genes Cells 2006;11:1173–1182.
10.
Namihira M, Honma S, Abe H, Tanahashi Y, Ikeda M, Honma K: Daily variation and light responsiveness of mammalian clock gene, Clock and BMAL1, transcripts in the pineal body and different areas of brain in rats. Neurosci Lett 1999;267:69–72.
11.
Fukuhara C, Dirden JC, Tosini G: Circadian expression of period 1, period 2, and arylalkylamine N-acetyltransferase mRNA in the rat pineal gland under different light conditions. Neurosci Lett 2000;286:167–170.
12.
Takekida S, Yan L, Maywood ES, Hastings MH, Okamura H: Differential adrenergic regulation of the circadian expression of the clock genes Period1 and Period2 in the rat pineal gland. Eur J Neurosci 2000;12:4557–4561.
13.
Von Gall C, Schneider-Huther I, Pfeffer M, Dehghani F, Korf HW, Stehle JH: Clock gene protein mPER1 is rhythmically synthesized and under cAMP control in the mouse pineal organ. J Neuroendocrinol 2001;13:313–316.
14.
Nakamura TJ, Shinohara K, Funabashi T, Mitsushima D, Kimura F: Circadian and photic regulation of cryptochrome mRNAs in the rat pineal gland. Neurosci Res 2001;41:25–32.
15.
Karolczak M, Burbach GJ, Sties G, Korf HW, Stehle JH: Clock gene mRNA and protein rhythms in the pineal gland of mice. Eur J Neurosci 2004;19:3382–3388.
16.
Simonneaux V, Poirel VJ, Garidou ML, Nguyen D, Diaz-Rodriguez E, Pevet P: Daily rhythm and regulation of clock gene expression in the rat pineal gland. Brain Res Mol Brain Res 2004;120:164–172.
17.
Engel L, Lorenzkowski V, Langer C, Rohleder N, Spessert R: The photoperiod entrains the molecular clock of the rat pineal. Eur J Neurosci 2005;21:2297–2304.
18.
Wongchitrat P, Felder-Schmittbuhl MP, Phansuwan-Pujito P, Pevet P, Simonneaux V: Endogenous rhythmicity of Bmal1 and Rev-erbα in the hamster pineal gland is not driven by norepinephrine. Eur J Neurosci 2009;29:2009–2016.
19.
Klein DC, Coon SL, Roseboom PH, Weller JL, Bernard M, Gastel JA, Zatz M, Iuvone PM, Rodriguez IR, Begay V, Falcon J, Cahill GM, Cassone VM, Baler R: The melatonin rhythm-generating enzyme: molecular regulation of serotonin N-acetyltransferase in the pineal gland. Recent Prog Horm Res 1997;52:307–357.
20.
Simonneaux V, Ribelayga C: Generation of the melatonin endocrine message in mammals: a review of the complex regulation of melatonin synthesis by norepinephrine, peptides, and other pineal transmitters. Pharmacol Rev 2003;55:325–395.
21.
Vanecek J, Sugden D, Weller J, Klein DC: Atypical synergistic α1- and β-adrenergic regulation of adenosine 3′,5′-monophosphate and guanosine 3′,5′-monophosphate in rat pinealocytes. Endocrinology 1985;116:2167–2173.
22.
Nelson W, Tong YL, Lee JK, Halberg F: Methods for cosinor-rhythmometry. Chronobiologia 1979;6:305–323.
23.
Oishi K, Sakamoto K, Okada T, Nagase T, Ishida N: Antiphase circadian expression between BMAL1 and period homologue mRNA in the suprachiasmatic nucleus and peripheral tissues of rats. Biochem Biophys Res Commun 1998;253:199–203.
24.
Abe M, Herzog ED, Yamazaki S, Straume M, Tei H, Sakaki Y, Menaker M, Block GD: Circadian rhythms in isolated brain regions. J Neurosci 2002;22:350–356.
25.
Fukuhara C, Dirden JC, Tosini G: Regulation of period 1 expression in cultured rat pineal. Neurosignals 2002;11:103–114.
26.
Simonneaux V, Ouichou A, Pevet P, Masson-Pevet M, Vivien-Roels B, Vaudry H: Kinetic study of melatonin release from rat pineal glands using a perifusion technique. J Pineal Res 1989;7:63–83.
27.
Yoshikawa T, Yamazaki S, Menaker M: Effects of preparation time on phase of cultured tissues reveal complexity of circadian organization. J Biol Rhythms 2005;20:500–512.
28.
Christ E, Pfeffer M, Korf HW, von Gall C: Pineal melatonin synthesis is altered in period1-deficient mice. Neuroscience 2010;171:398–406.
29.
Baggs JE, Price TS, DiTacchio L, Panda S, Fitzgerald GA, Hogenesch JB: Network features of the mammalian circadian clock. PLoS Biol 2009;7:e52.
30.
Meng QJ, McMaster A, Beesley S, Lu WQ, Gibbs J, Parks D, Collins J, Farrow S, Donn R, Ray D, Loudon A: Ligand modulation of REV-ERBα function resets the peripheral circadian clock in a phasic manner. J Cell Sci 2008;121:3629–3635.
31.
Welsh DK, Yoo SH, Liu AC, Takahashi JS, Kay SA: Bioluminescence imaging of individual fibroblasts reveals persistent, independently phased circadian rhythms of clock gene expression. Curr Biol 2004;14:2289–2295.
32.
Abraham U, Prior JL, Granados-Fuentes D, Piwnica-Worms DR, Herzog ED: Independent circadian oscillations of Period1 in specific brain areas in vivo and in vitro. J Neurosci 2005;25:8620–8626.
33.
Stratmann M, Schibler U: Properties, entrainment, and physiological functions of mammalian peripheral oscillators. J Biol Rhythms 2006;21:494–506.
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