Individual neurons within the suprachiasmatic nuclei (SCNs) are capable of functioning as autonomous clocks and generating circadian rhythms in the expression of genes that form the molecular clockworks. Limited information is available on how these molecular oscillations in individual clock cells are coordinated to provide for the ensemble rhythmicity that is normally observed from the entire SCN. Because calcium influx via voltage-dependent calcium channels (VDCCs) has been implicated in the regulation of gene expression and synchronization of rhythmicity across the population of SCN clock cells, we first examined the rat SCN and an immortalized line of SCN cells (SCN2.2) for expression and circadian regulation of different VDCC α1 subunits. The rat SCN and SCN2.2 cells exhibited mRNA expression for all major types of VDCC α1 subunits. Relative levels of VDCC expression in the rat SCN and SCN2.2 cells were greatest for L-type channels, moderate for P/Q- and T-type channels, and minimal for R- and N-type channels. Interestingly, both rat SCN and SCN2.2 cells showed rhythmic expression of P/Q- and T-type channels. VDCC involvement in the regulation of molecular rhythmicity in SCN2.2 cells was then examined using the nonselective antagonist, cadmium. The oscillatory patterns ofrPer2andrBmal1expression were abolished in cadmium-treated SCN2.2 cells without affecting cellular morphology and viability. These findings raise the possibility that the circadian regulation of VDCC activity may play an important role in maintaining rhythmic clock gene expression across an ensemble of SCN oscillators.

Ascorbic acid is essential for normal brain development and homeostasis. However, the effect of ascorbic acid on adult brain aging has not been determined. Long-term treatment with high levels of D-galactose (D-gal) induces brain aging by accumulated oxidative stress. In the present study, mice were subcutaneously administered with D-gal (150 mg/kg/day) for 10 weeks; from the seventh week, ascorbic acid (150 mg/kg/day) was orally co-administered for four weeks. Although D-gal administration alone reduced hippocampal neurogenesis and cognitive functions, co-treatment of ascorbic acid with D-gal effectively prevented D-gal-induced reduced hippocampal neurogenesis through improved cellular proliferation, neuronal differentiation, and neuronal maturation. Long-term D-gal treatment also reduced expression levels of synaptic plasticity-related markers, i.e., synaptophysin and phosphorylated Ca²⁺/calmodulin-dependent protein kinase II, while ascorbic acid prevented the reduction in the hippocampus. Furthermore, ascorbic acid ameliorated D-gal-induced downregulation of superoxide dismutase 1 and 2, sirtuin1, caveolin-1, and brain-derived neurotrophic factor and upregulation of interleukin 1 beta and tumor necrosis factor alpha in the hippocampus. Ascorbic acid-mediated hippocampal restoration from D-gal-induced impairment was associated with an enhanced hippocampus-dependent memory function. Therefore, ascorbic acid ameliorates D-gal-induced impairments through anti-oxidative and anti-inflammatory effects, and it could be an effective dietary supplement against adult brain aging.

The leaner (tg^la) mouse mutation occurs in the gene encoding the voltage-activated Ca²⁺channel α_1A subunit, the pore-forming subunit of P/Q-type Ca²⁺ channels. This mutation results in dramatic reductions in P-type Ca²⁺ channel function in cerebellar Purkinje neurons of tg^la/tg^la mice that could affect intracellular Ca²⁺ signaling. We combined whole cell patch-clamp electrophysiology with fura-2 microfluorimetry to examine aspects of Ca²⁺ homeostasis in acutely dissociated tg^la/tg^la Purkinje cells. There was no difference between resting somatic Ca²⁺ concentrations in tg^la/tg^la cells and in wild-type (+/+) cells. However, by quantifying the relationship between intracellular Ca²⁺ elevations and depolarization-induced Ca²⁺influx, we detected marked alterations in rapid calcium buffering between the two genotypes. Calcium buffering values (ratio of bound/free ions) were significantly reduced in tg^la/tg^la (584 ± 52) Purkinje cells relative to +/+ (1,221 ± 80) cells. By blocking the endoplasmic reticulum (ER) Ca²⁺-ATPases with thapsigargin, we observed that the ER had a profound impact on rapid Ca²⁺ buffering that was also differential between tg^la/tg^laand +/+ Purkinje cells. Diminished Ca²⁺ uptake by the ER apparently contributes to the reduced buffering ability of mutant cells. This report constitutes one of the few instances in which the ER has been implicated in rapid Ca²⁺ buffering. Concomitant with this reduced buffering, in situ hybridization with calbindin D28k and parvalbumin antisense oligonucleotides revealed significant reductions in mRNA levels for these Ca²⁺-binding proteins (CaBPs) in tg^la/tg^la Purkinje cells. All of these results suggest that alterations of Ca²⁺ homeostasis in tg^la/tg^la mouse Purkinje cells may serve as a mechanism whereby reduced P-type Ca²⁺ channel function contributes to the mutant phenotype.