Prokineticin 2 protein is diurnally expressed in PER2-containing clock neurons in the mouse suprachiasmatic nucleus

Abstract

Expression of prokineticin 2 (PK2) mRNA in the suprachiasmatic nucleus (SCN), also known as the brain's clock, exhibits circadian oscillations with peak levels midday, zeitgeber time (ZT) 4, and almost undetectable levels during night. This circadian expression profile has substantially contributed to the suggested role of PK2 as an SCN output molecule involved in transmitting circadian rhythm of behavior and physiology. Due to unreliable specificity of PK2 antibodies, the 81 amino acid protein has primarily been studied at the mRNA level and correlation between circadian oscillating mRNAs and protein products are infrequent. Hence, data on PK2 protein expression in the SCN is lacking. In this study a thorough validation of a commercial PK2 antibody for immunohistochemistry (IHC) was performed followed by fluorescence IHC on SCN mouse brain sections at six consecutive ZTs over a 24-h cycle (12:12 light-dark, ZT0 =light ON whereas ZT12 =light OFF). Data were visualized and processed using confocal microscopy. Results showed that PK2 protein expression diurnally oscillates with calculated peak expression ZT5:40 ± 1:40 h. Opposite than described for PK2 mRNA, PK2 immunoreactivity was detectable at all times during the 24-h cycle. PK2 was primarily located in neurons of the shell compartment and > 80 % of these neurons co-expressed the core clock protein PER2. In conclusion, PK2 protein expression oscillates as the mRNA, supporting the suggested role of PK2 as a SCN molecule involved in circadian rhythm regulation.

Keywords: Diurnal; Immunohistochemistry; Mouse; Oscillation; Period 2; Prokineticin 2; Suprachiasmatic Nucleus.