**2. Biological Rhythms**

Exposure to periodic environmental changes during evolution is thought to have driven the development of adaptive biological rhythms of which the best known are the circadian rhythms, which have a period length of around 24 h. However, there are also ultradian rhythms (>24 h) and infradian rhythms (<24 h) [14,15]. Biological rhythms allow the adaptation to changing environments, from the light-night cycle, to the seasons or feed-fast cycles. However, current 24/7 lifestyles dim the environmental di fferences between day and night, resulting in weak zeitgebers (weak day light, absence of darkness during night, constant environmental temperature, sedentarism and frequent snacking), which may impair the circadian system [16].

The central circadian clock lies in the suprachiasmatic nucleus in the anterior hypothalamus and coordinates peripheral clocks, including the kidney circadian clock which, in turn, coordinate local physiologic functions with patterns of activity and/or feeding [17]. Several signals contribute to coordinate peripheral circadian rhythms, including hormone secretion (e.g., production of the melatonin hormone by the pineal gland during nighttime, circadian production of aldosterone), neuronal activity (including physical activity and feeding) and body temperature. In addition, canonical clock genes (e.g., *Clock*, *Bmal1*, *Rev-erb*<sup>α</sup>, *Cry1*, *Cry2*, *Per1*, *Per2*) are expressed and/or active in a cyclical manner within cells, driving cell autonomous circadian rhythms [14,15]. In the most basic regulatory loop, *Clock* and *Bmal1* are transcription factors that promote *Cry* and *Per* gene expression, and *Cry* and *Per* in turn suppress *Clock*/*Bmal1* induction of their own transcription [18] (Figure 1). On top of this basic regulatory loop, associated elements account for the circadian regulation of 13% of kidney expressed genes. Furthermore, posttranslational modifications (e.g., phosphorylation, acetylation) are also responsible for circadian changes in protein activity. Functional circadian molecular clockwork evolves in the late fetal and early postnatal kidney. During the nursing period, oscillations are entrained by nutritional cues [19].

**Figure 1.** Canonical clock genes and the basic regulatory loop: impact on the kidney of genetic defects. In the most basic regulatory loop, *Clock* and *Bmal1* are transcription factors that promote *Cry* and *Per* gene expression, and Cry and Per proteins, in turn, suppress *Clock*/*Bmal1* induction of their Cry and Per transcription. Genetic disruption of some canonical clock genes has yielded renal-hypertension phenotypes as illustrated above for *Clock*, *Bmal1*, and *Per1* in mice. *Clock* KO mice display loss of water and electrolyte excretion rhythmicity as well as differential responses to induction of kidney fibrosis, which appears specific of the driver of fibrosis (worse unilateral ureteral obstruction (UUO)-induced fibrosis but milder sodium overload-induced fibrosis). *Bmal1* KO mice display accelerated aging, loss of rhythmicity of water excretion as well as non-dipping hypotension (red line) as compared to the normal blood pressure circadian rhythm (green line). *Per1* KO mice display non-dipping hypertension (red line) as compared to the normal blood pressure circadian rhythm (green line).

Kidney function has circadian rhythms (Table 1). The amplitude of circadian oscillations in GFR and renal plasma flow are around 50%, while water and electrolyte (sodium, potassium, calcium, magnesium, and phosphate) excretion may be several fold higher during the active phase and this is paralleled by circadian changes in kidney oxygenation and the corticomedullary interstitial osmolarity gradient and in the expression of genes involved in its regulation (e.g., vasopressin receptors V1aR, V2R, urea transporter UT-A2 and water channel Aqp2) [14]. Changes in kidney oxygenation modulate HIF-1α activation and erythropoietin levels, which display an amplitude of more than 10-fold under constant darkness and normoxia in mice [15]. Blood pressure peaks early in the beginning of the active period of both diurnal and nocturnal animals [20]. Molecular clocks regulate sodium balance, sympathetic function and vascular tone, all contributing to blood pressure regulation. Altered kidney circadian rhythms have been associated with the development of hypertension, chronic kidney disease, and kidney stones (reviewed in [14]).


**Table 1.** Some examples of kidney functions which have circadian rhythms.

Insights into the circadian regulation of kidney functions is derived from genetic defects in clock genes [14] (Figure 1). Thus, *Per1* KO mice develop non-dipping hypertension under conditions of sodium retention while *Clock* KO mice lose the circadian rhythmicity in urinary water and electrolyte excretion and develop more severe kidney fibrosis upon ureteral obstruction but were protected from kidney fibrosis driven by sodium retention conditions [14]. Additionally, *Clock* mutants had some features suggesting increased severity of adenine-induced CKD, such as higher blood pressure and expression as some gelatinase genes, but there were no differences in kidney fibrosis or serum creatinine [21]. *Bmal1* KO mice develop accelerated aging, hypotension and a non-dipping blood pressure pattern and lose the circadian variations in interstitial medullary osmolarity suggesting a role of circadian clocks in the control of urine volume beyond dietary clues [14,22]. Kidneys from conditional nephron-specific *Bmal1* deletion mice exhibited a decrease in NAD+-to-NADH ratio, increase in plasma urea and creatinine and a reduced capacity of the kidney to secrete anionic drugs (furosemide) paralleled by changes in the expression of tubule transporters such as organic anion transporter 3 (SLC22a8) [23]. Na+-H+ exchanger 3 (NHE3) activity also has rhythmic oscillations causing daily fluctuations in Na+ and water transport of the proximal tubule cell.
