Aurea Blancas

Postdoc, Rath Lab

Title: Time-giving cues in cell culture to preserve circadian biological functions of pinealocytes in-vitro

Abstract: Circadian function is vital to prepare the organism to anticipate 24h cyclic changes in the environment. This circadian system has a hierarchical organization containing self-oscillating autonomous nuclei which are able to self-sustain their rhythmic functions even when they are isolated from the whole organism, for example, by maintaining the rhythmic expression of clock-genes, a group of genes that constitute the molecular circadian clock and display auto-regulatory feedback loops with a 24h period. Virtually, all cells in the body contain this molecular clock, but in most of the tissues when isolated, the rhythmic expression of clock genes is lost.

The pineal gland is a non-autonomous oscillator composed mainly of pinealocytes, a cell type, which synthesizes and releases melatonin during nighttime. Oscillating clock genes are expressed in the pineal gland, but their role in the pinealocyte is unknown. Thus, our aim is to demonstrate the possible relationship between molecular clock of the pinealocyte and circadian melatonin synthesis. Our current approach is to use siRNAs targeting clock genes in combination with a “synchronization protocol” where the cultured cells are simulated with norepinephrine at night during repeated 24h cycles to mimic the in-vivo situation. Using the synchronization protocol, we have been able to re-establish not only rhythmic clock-gene expression but also oscillations in other processes and we show functional relationship between clock-genes and transcription factors that drive circadian melatonin synthesis.