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Differentiation of PC12 Cells Results in Enhanced VIP Expression and Prolonged Rhythmic Expression of Clock Genes

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Abstract

To examine for circadian rhythmicity, the messenger RNA (mRNA) amount of the clock genes Per1 and Per2 was measured in undifferentiated and nerve-growth-factor-differentiated PC12 cells harvested every fourth hour. Serum shock was needed to induce circadian oscillations, which in undifferentiated PC12 cultures lasted only one 24-h period, while in differentiated cultures, the rhythms continued for at least 3 days. Thus, neuronal differentiation provided PC12 cells the ability to maintain rhythmicity for an extended period. Both vasoactive intestinal polypeptide (VIP) and its receptor VPAC2 are expressed in the suprachiasmatic nucleus (SCN), and in agreement with VIP signaling being crucial for maintenance of rhythmicity, we found both VIP and VPAC2 mRNA increased after differentiation of PC12 cells. Pituitary adenylate cyclase activating polypeptide (PACAP) exerts time- and concentration-dependent effects on Per gene expression in the SCN. We added 1 nM and 1 μM PACAP to oscillating PC12 cells at times corresponding to midday and early and late night to evaluate whether the effects were similar as in SCN. Induction of Per1 mRNA was found at all three times, which differs from results in SCN. Thus, PC12 cells seem more useful for studying mechanisms behind acquirement of rhythmicity of cell cultures than for resetting of circadian rhythm.

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Abbreviations

HS:

horse serum

NGF:

nerve growth factor

PACAP:

pituitary adenylate cyclase activating polypeptide

PBS:

phosphate-buffered saline

RHT:

retinohypothalamic tract

RT-PCR:

reverse transcription polymerase chain reaction

SCN:

suprachiasmatic nucleus

VIP:

vasoactive intestinal polypeptide

ZT:

zeitgeber time

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Acknowledgments

The skilful technical assistance of Sanne S. Kelly, Lone K. Hellström, and Anita Hansen is gratefully acknowledged. Henrik L. Jørgensen, PhD is acknowledged for his help with the statistical analysis. The study was supported by The Danish Biotechnology Center for Cellular Communication.

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Authors and Affiliations

  1. Department of Clinical Biochemistry, Bispebjerg Hospital, Faculty of Health Sciences, University of Copenhagen, 2400, Copenhagen, NV, Denmark

    Camilla P. Pretzmann, Jan Fahrenkrug & Birgitte Georg

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  1. Camilla P. Pretzmann
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  2. Jan Fahrenkrug
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  3. Birgitte Georg
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Correspondence to Birgitte Georg.

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Pretzmann, C.P., Fahrenkrug, J. & Georg, B. Differentiation of PC12 Cells Results in Enhanced VIP Expression and Prolonged Rhythmic Expression of Clock Genes. J Mol Neurosci 36, 132–140 (2008). https://doi.org/10.1007/s12031-008-9063-9

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  • DOI: https://doi.org/10.1007/s12031-008-9063-9

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