Intrinsic circadian timekeeping properties of the thalamic lateral geniculate nucleus.

Circadian rhythmicity in mammals is sustained by the central brain clock-the suprachiasmatic nucleus of the hypothalamus (SCN), entrained to the ambient light-dark conditions through a dense retinal input. However, recent discoveries of autonomous clock gene expression cast doubt on the supremacy of the SCN and suggest circadian timekeeping mechanisms devolve to local brain clocks. Here, we use a combination of molecular, electrophysiological, and optogenetic tools to evaluate intrinsic clock properties of the main retinorecipient thalamic center-the lateral geniculate nucleus (LGN) in male rats and mice. We identify the dorsolateral geniculate nucleus as a slave oscillator, which exhibits core clock gene expression exclusively in vivo. Additionally, we provide compelling evidence for intrinsic clock gene expression accompanied by circadian variation in neuronal activity in the intergeniculate leaflet and ventrolateral geniculate nucleus (VLG). Finally, our optogenetic experiments propose the VLG as a light-entrainable oscillator, whose phase may be advanced by retinal input at the beginning of the projected night. Altogether, this study for the first time demonstrates autonomous timekeeping mechanisms shaping circadian physiology of the LGN.

Studies on primary uveal and cutaneous melanoma cell interaction with vitronectin.

We have examined the diversity between primary uveal (92-1 and Mel202) and cutaneous (FM55P and IGR-39) melanoma cells in their interaction with vitronectin, and established the effect of integrins and ß1,6-branched N-oligosaccharides on this process. The adhesion level of uveal melanoma cells to vitronectin was at least twice lower than that of cutaneous ones, but all cells tested repaired scratch wounds on vitronectin-coated surfaces with similar speed. Swainsonine treatment, by reducing the amount of ß1,6-branches, significantly decreased cell attachment in all cases, but reduction of wound healing efficiency was compromised only in cutaneous melanoma cell. Functional blocking antibodies used in adhesion and migration assays revealed that integrin αvß3 was strongly involved in adhesion and migration only in cutaneous melanoma cells, but its role here was less pronounced than that of integrin αvß5. However, in uveal melanoma the specific anti-αvß5 integrin antibody had no impact on migration speed. Therefore, the anti-α3ß1 integrin antibody was used in order to explain the nature of uveal melanoma interaction with vitronectin, which caused a mild decrease in adhesion efficiency and reduced their motility. Expression of αvß5 integrin differed between the cell lines, but there was no distinct pattern to distinguish uveal melanoma from cutaneous melanoma. In conclusion, αvß5, but not αvß3 integrin is heavily involved in uveal melanoma cell interaction with vitronectin. The role of ß1,6-branched N-glycans in the adhesion, but not during migration, of all cells to vitronectin has been confirmed.

Diversified ß-2-adrenergic Receptor Expression and Action in Melanoma Cells.

BACKGROUND/AIM: Growing evidence links stress hormones with development and progression of various cancer types. The aim of this study was to assess susceptibility of cutaneous and uveal melanoma cells to adrenaline (AD). MATERIALS AND METHODS: The expression of ß-2-adrenergic receptor in primary cutaneous (FM-55-P), primary uveal (92-1, Mel202) and metastatic cutaneous (A375) melanoma cells was estimated at mRNA, protein and cell surface levels. The impact of AD on cell proliferation and migration was also studied. RESULTS: The expression of ß-2-adrenergic receptor was cell line-dependent. Adrenaline treatment caused a slight stimulation of melanoma cell proliferation and activation of matrix metalloproteinases. Adrenaline-treated uveal melanoma cells showed an increased migration rate, whereas, in cutaneous melanoma cells, no changes or even lower migration speed were observed. CONCLUSION: Melanoma cell susceptibility to AD varies depending on origin and progression stage. Metastatic cutaneous melanoma cells were found to be less responsive to AD than primary cutaneous and uveal melanoma cells.