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Clock Gene Expression in the Submandibular Glands

¹ Department of Dental and Medical Biochemistry, and
² Department of Prosthetic Dentistry, Hiroshima University Graduate School of Biomedical Sciences, Hiroshima 734-8553, Japan; and
³ Department of Physiology, Hokkaido University Graduate School of Medicine, Sapporo 060-8638, Japan;

View larger version (25K): [in this window] [in a new window]   Figure 1. Circadian profiles of clock-related genes in the submandibular glands. Six-week-old male C57/BL6 mice were housed under 12:12-hour LD conditions for 19 days, and were killed at the indicated zeitgeber time (ZT: lights on at ZT0, lights off at ZT12)) on day 20. Total RNA was isolated from the submandibular glands and subjected to real-time RT-PCR analysis. Per1, Per2, Cry1, Bmal1, Dec1, Dec2, Dbp, and Reverb mRNA levels (mean ± SEM, n = 3) were determined. P-values were calculated by one-way ANOVA analysis. Expression profiles of Per1, Per2, Cry1, Bmal1, Dec1, Dec2, Dbp, and Reverb were rhythmic. To examine the effect of starvation on Cry1 expression, we withheld foods for 26–46 hrs from ZT0 on day 19 (a dotted line).

View larger version (36K): [in this window] [in a new window]   Figure 2. Daily expression profiles of the salivary-gland-characteristic genes in the submandibular glands. After mice were housed under 12:12-hour LD conditions for 19 days, expression levels (mean ± SEM, n = 3) of mRNAs for amylase 1 (Amy1), mucin 10 (Muc10), cystatin C (Cst3), and epidermal growth factor (Egf) were determined in the presence or absence (dotted lines) of foods. Foods were withheld at ZT0 on day 19, and the mice were killed at the indicated zeitgeber time (ZT—recorded in hours) on day 20. P-values without or within parentheses (the starved group) are shown in the panel. Fasting altered the expression profile of Amy1 mRNA, but it did not induce circadian rhythms of other salivary-gland-characteristic genes.

View larger version (30K): [in this window] [in a new window]   Figure 3. Expression of clock-related genes in the submandibular glands in Clock/Clock mice. Wild-type (WT) and Clock/Clock (Ck/Ck) mice were housed under LD conditions and killed at ZT6 or ZT18. Per2, Bmal1, Dbp, Dec1, Dec2, and Amy1 mRNA levels (mean ± SEM, n = 3) were determined. * P < 0.05 (standard t test).

View larger version (31K): [in this window] [in a new window]   Figure 4. Expression of clock-related genes in the submandibular glands in Cry1–/– and Cry1–/–Cry2–/– mice. Cry1–/– and Cry1–/–Cry2–/– mice were housed under LD conditions and killed at ZT6 or ZT18. Per2, Bmal1, Dbp, Dec1, Dec2, and Amy1 mRNA levels (mean ± SEM, n = 3) were determined. * P < 0.05 (standard t test).

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