GSK3 activity regulates rhythms in hippocampal clock gene expression and synaptic plasticity.

Hippocampal rhythms in clock gene expression, enzymatic activity, and long-term potentiation (LTP) are thought to underlie day-night differences in memory acquisition and recall. Glycogen synthase kinase 3-beta (GSK3ß) is a known regulator of hippocampal function, and inhibitory phosphorylation of GSK3ß exhibits region-specific differences over the light-dark cycle. Here, we sought to determine whether phosphorylation of both GSK3α and GSK3ß isoforms has an endogenous circadian rhythm in specific areas of the hippocampus and whether chronic inhibition or activation alters the molecular clock and hippocampal plasticity (LTP). Results indicated a significant endogenous circadian rhythm in phosphorylation of GSK3ß, but not GSK3α, in hippocampal CA1 extracts from mice housed in constant darkness for at least 2 weeks. To examine the importance of this rhythm, genetic and pharmacological strategies were used to disrupt the GSK3 activity rhythm by chronically activating or inhibiting GSK3. Chronic activation of both GSK3 isoforms in transgenic mice (GSK3-KI mice) diminished rhythmic BMAL1 expression. On the other hand, chronic treatment with a GSK3 inhibitor significantly shortened the molecular clock period of organotypic hippocampal PER2::LUC cultures. While WT mice exhibited higher LTP magnitude at night compared to day, the day-night difference in LTP magnitude remained with greater magnitude at both times of day in mice with chronic GSK3 activity. On the other hand, pharmacological GSK3 inhibition impaired day-night differences in LTP by blocking LTP selectively at night. Taken together, these results support the model that circadian rhythmicity of hippocampal GSK3ß activation state regulates day/night differences in molecular clock periodicity and a major form of synaptic plasticity (LTP).

Circadian rhythmicity of active GSK3 isoforms modulates molecular clock gene rhythms in the suprachiasmatic nucleus.

The suprachiasmatic nucleus (SCN) drives and synchronizes daily rhythms at the cellular level via transcriptional-translational feedback loops comprising clock genes such as Bmal1 and Period (Per). Glycogen synthase kinase 3 (GSK3), a serine/threonine kinase, phosphorylates at least 5 core clock proteins and shows diurnal variation in phosphorylation state (inactivation) of the GSK3ß isoform. Whether phosphorylation of the other primary isoform (GSK3α) varies across the subjective day-night cycle is unknown. The purpose of this study was to determine if the endogenous rhythm of GSK3 (α and ß) phosphorylation is critical for rhythmic BMAL1 expression and normal amplitude and periodicity of the molecular clock in the SCN. Significant circadian rhythmicity of phosphorylated GSK3 (α and ß) was observed in the SCN from wild-type mice housed in constant darkness for 2 weeks. Importantly, chronic activation of both GSK3 isoforms impaired rhythmicity of the GSK3 target BMAL1. Furthermore, chronic pharmacological inhibition of GSK3 with 20 µM CHIR-99021 enhanced the amplitude and shortened the period of PER2::luciferase rhythms in organotypic SCN slice cultures. These results support the model that GSK3 activity status is regulated by the circadian clock and that GSK3 feeds back to regulate the molecular clock amplitude in the SCN.