Hypothesis / aims of study
Humans rarely void during sleep, partly because urine production decreases and bladder storage capacity increases at night. Circadian rhythm disruption has been implicated in pediatric nocturnal enuresis and nocturia in older adults(1). We hypothesized that defective circadian regulation of bladder capacity and urine production during development contributes to nocturnal enuresis, and aimed to clarify the developmental timing of clock gene circadian rhythm acquisition in the bladder and kidney and the relationship of this timing with voiding behavior.
Study design, materials and methods
Male and female rats aged 0, 1, 3, and 6 weeks (n=3 per sex, per group) were examined as described below. Bladder, kidney, and blood samples were collected at 0, 4, 8, 12, 16, and 20 h. mRNA level circadian clock (Bmal1, Clock, Per2, Cry1/Cry2), bladder mechanosensor (TRPV1, TRPV4, Piezo1), and vasopressin receptor (V1aR, V2R) gene expression was quantified using qRT-PCR. Plasma vasopressin levels were measured. Urine volume and voiding frequencies of 3- and 6-week-old rats (n=6 per sex per group) were assessed using metabolic cages to evaluate circadian patterns.
Results
In 0- and 1-week-old rats, no circadian periodicity was observed in clock genes, mechanosensors, vasopressin receptors, or plasma vasopressin levels in the bladder or kidney. In contrast, 3- and 6-week-old rats exhibited clear circadian rhythms in clock gene expression and plasma vasopressin levels (Figure1). Circadian variations in bladder mechanosensors and vasopressin receptors were evident at six weeks (Figure2). Metabolic cage analysis showed no circadian rhythm in urine volume or voiding frequency at 3 weeks, whereas 6-week-old mice demonstrated increased voiding during the active phase and reduced voiding during the sleep phase.
Interpretation of results
These findings suggest that Circadian rhythms in clock genes, mechanosensors, and vasopressin-related pathways in the bladder and kidneys are gradually acquired during development. Establishment of these molecular rhythms appears to contribute to development of mature circadian voiding behavior.