THE HISTORY AND IMPORTANCE OF CIRCADIAN RHYTHMS

The term “circadian” was properly defined in the 1970s as follows: “circadian: relating to biologic variations or rhythms with a frequency of 1 cycle every 24 hours (± 4 hours) (21).” Because life on Earth adapted to constant changes in the environment, this 24-h cycle correlates with the Earth’s rotation (63). All living organisms have circadian rhythms that dictate feeding schedules, locomotor activity, time of rest, and gene transcription that directly coincides to the rise and fall of the sun (63). The first known study of circadian rhythms came from an astronomer named Jean-Jacques d’Ortous de Mairan, who, in 1729, observed that leaves of a mimosa plant moved in a rhythmic pattern every 24 h. The 24-h rhythm of leaf movement still occurred after the plants were moved to a dark basement with no light (64). This led to the eventual discovery that an organism’s internal clock must be “entrainable” by the changing solar cycles throughout different seasons (i.e., an organism must be able to adapt to environmental cues) (64). These initial observations have spurred decades of research into how circadian rhythm is regulated, and, recently, how it becomes dysregulated with age and disease. In 2017, the Nobel Prize in Medicine was awarded to Jeffery C. Hall, Michael Rosbash, and Michael W. Young for their work on how circadian rhythms impact human biology. They discovered that 24-h biological rhythms were synchronized from metabolism and behavior to light-dark cycles. Even with current advances in our knowledge surrounding circadian rhythms, scientists are still uncovering the complex world of what leads to disrupted synchronization and how changes in a person’s internal clock can trigger or accelerate certain human diseases.

MOLECULAR MECHANISMS IN CIRCADIAN RHYTHM CONTROL

The mammalian circadian clock operates on a 24-h cycle and regulates responses and adaptations to daily changes in the surrounding environment. Physiological, endocrine, and metabolic responses are coordinated in a rhythmic pattern by the suprachiasmatic nucleus (SCN) located within the anterior hypothalamus (27). The SCN is known as the central, master clock controller that drives several regulatory mechanisms, including positive and negative feedback loop systems. The core feedback loop is regulated by the activator genes circadian locomotor output cycles kaput (Clock) and brain and muscle aryl hydrocarbon receptor nuclear translocator-like protein (ARNT)-like 1 (Bmal1), which are subunits in basic helix-loop-helix-PER-ARNT-SIM (PAS) transcription factors. These two genes regulate a number of accessory genes in the molecular clock pathway (43), including Period1 (Per1), Period2 (Per2), Period3 (Per3), Crytochrome1 (Cry1), and Cryptochrome 2 (Cry2), which together gather feedback to suppress the CLOCK/BMAL heterodimer (23). Additionally, a second feedback loop known as the Rev-Erbαβ complex coordinates circadian timing. These feedback loops fluctuate up and down over a 24-h time period and are reset every morning when light enters the SCN via the retinohypothalamic tract. In other words, the SCN circadian oscillator establishes its functional effectiveness because it can be entrained from the Earth’s rotation due to the day-night light cycles (53). Other factors besides light, albeit at a lesser intensity, can drive SCN activity in mammals, such as feeding (22, 52, 55) and voluntary or forced exercise (22, 55, 60). Expression of key circadian rhythm genes is linked to physiological functions in all peripheral tissues. Thus, disruption of the SCN is important, with aberrant function caused by increasing age, jetlag, rotating night-shift work, increased light due to urban sprawl, and others. The impact of these factors on the central clock has been reviewed elsewhere (57, 65); thus, the focus of our review will be circadian misalignment specifically in the kidney, with a focus on age as a driver of misalignment and physical activity as a means to restore clock function.

CIRCADIAN RHYTHMS IN THE KIDNEY

The SCN, in addition to centrally controlling circadian rhythms, also dictates circadian rhythms in peripheral tissues like the kidney (36). Most CLOCK genes are expressed in peripheral tissues; however, there is approximately a 4-h delay in peripheral expression compared with that within the SCN. Peripheral clocks are tissue specific, and all have their own endogenous circadian rhythms yet require the central clock within the SCN to maintain synchronization (67). The kidney has several important physiological functions that are governed by the SCN, including removing waste and metabolites from the blood (38), regulating extracellular fluid volume (40), and producing several hormones that help to regulate blood pressure (54) and control Ca²⁺ metabolism (13) (Fig. 1). Peripheral endogenous circadian rhythms differ from each other and are influenced by the SCN via a combination of neural and hormonal signals (67). Arginine vasopressin (AVP), also known as antidiuretic hormone, is a hormone that is synthesized by neurons that have their cell bodies located in the hypothalamus [SCN, supraoptic nucleus (SON), and paraventricular nucleus (PVN)]. AVP is then cleaved, stored in secretory vesicles, and transported to the posterior pituitary before being released into the bloodstream to act on peripheral tissues, including the kidney (7, 66). Thus, AVP neurons play a critical role in maintaining appropriate circadian function of the SCN network and peripherally (37). AVP is released primarily in response to stimulation of osmoreceptors within the hypothalamus due to increased osmolality of blood. However, other stimuli for AVP secretion include pain, nausea, hypoglycemia, reduced blood pressure, and exercise (6, 7). Once released, AVP acts on peripheral tissues through its binding to three unique receptors, V2, V1a (or V1), and V1b (or V3). Although many studies have sought to control the influence of circadian rhythms on AVP concentration by standardizing time of day for sample collection, this relationship is still underdescribed. For example, it is understood that AVP concentrations increase at night due to the disinhibition of neurons stretching between the organum vasculosum lamina terminalis, SCN, and SON (19, 58). However, the implications of these fluctuations need further investigation, especially as they are related to health outcomes.

Alignment of circadian rhythms is critical for renal function, as misalignment of circadian patterns of water and electrolyte excretion has been proposed to cause abnormal blood pressure leading to a nondipping pattern in blood pressures at night (47). Nondipping blood pressure is generally defined as a nocturnal fall in blood pressure that is <10% from daytime pressure (46) and is associated with unfavorable patterns of blood pressure regulation that contribute to enhanced stroke risk (39), end-stage regnal disease (31), and even organ damage (44). Studies have linked specific renal genes to circadian changes in secretion/reabsorption capacities of the distal nephron and collecting ducts (10, 26, 45). For example, researchers have found that in the kidney, PER1 expression regulates genes related to Na⁺ transport and PER1 has been shown to regulate the NaCl cotransporter, which is important to renal Na⁺ retention (10, 45). The importance of circadian rhythm in the renal system is also evident in clock^−/− mutant mice (70). These mice demonstrate decreased urine osmolality, increased water intake, a modified Na⁺ excretion pattern, and increased plasma osmolality, all resulting in nondipping patterns of blood pressure at night (70). Together, this was the first study to demonstrate that altered circadian rhythms in the kidney could lead to unfavorable changes in blood pressure and increase risk for adverse events (70). The authors were also able to correlate the significant alterations in circadian rhythms in the kidney to changes in the mouse distal nephron segments and collecting ducts, uncovering a significant role of the molecular clock in renal function (70). Taken together, these results indicate that a functional renal molecular clock is essential for regulation of kidney function, including physiological and pathological control of blood pressure.

PHYSIOLOGY OF THE AGING KIDNEY

Aging is generally defined as a deterioration of tissue structure and function. From the perspective of the renal system, older humans and animals have diminished renal function. For example, glomerular filtration rate (GFR) declines with increasing age, such that by the age of 90, GFR is ~50% of young individuals (4). Some have argued that this marked decline in GFR is due to incorrect estimates from established calculators, such as the Modification of Diet in Renal Disease Study equation, which are not validated for older individuals (18), or that the decreased renal function with age is compounded by age-associated disease like diabetes and hypertension (34). However, although it is likely true that age-associated comorbidities and poorly validated equations for older populations exacerbate decreases in GFR with age, it is generally accepted that kidney function is diminished with increased age.

Several important physiological functions in the renal system change with age, which may explain the observed changes in GFR. Renal plasma flow decreases with age due to increased renal vascular resistance (15). The reduction in renal flow is greater than changes in GFR, resulting in an enhanced filtration fraction. Together, these changes in renal function result in greater resistance in the efferent arterioles and increased glomerular pressure, creating unfavorable hyperfiltration and damaging glomerular capillaries. Coupled with a reduction in the number of superficial nephrons due to age, homeostatic regulation in the kidney also becomes dysfunctional. For example, proximal reabsorption of Na⁺ is enhanced, whereas distal reabsorption is reduced, resulting in a narrowed control range and reduced response to altered Na⁺ load (15) and higher risk of volume depletion and salt retention. Also, the aged kidney demonstrates dysfunctional renal K⁺ handling, most likely due to decreased Na⁺-K⁺-ATPase activity (29) and reduced functional reserve in acid-base balance (16). The mechanisms responsible for these changes remain largely unknown, although urea transport and water channel expression (aquaporin 2 and 3, which are driven by AVP) change with age and likely play a role (48). Together, these observations support significant age-associated disruptions in renal function and warrant future investigation into the mechanisms responsible for age-related changes in renal function.

CIRCADIAN RHYTHMS IN THE AGING KIDNEY

Normal aging itself has a profound influence on central and peripheral circadian rhythms. In fact, one of the most common health complaints among older adults is sleep disturbances (56). Decreased function of the circadian clock has been recognized for several decades, with documented decreased amplitude, phase modifications, and reduced sensitivity to photic and nonphotic stimuli (11). Because of changes in gene expression of the molecular clock, older adults sleep fewer hours and have altered physiological responses, including in the kidney. A study of older adults compared with young healthy adults clearly demonstrated disturbances in the phase of electrolyte excretory patterns and reduction in peak to trough range, suggesting that the intrinsic mechanisms that control human renal diurnal rhythms undergo deterioration in old age (33). Similar changes have also been noted in kidney circadian gene expression (68), although the changes in the clock expression in the kidney are not as well documented as central changes. Our group has studied the aging kidney and found aberrant expression of Per2 in the morning, a change driven by female sex, identifying significant sex differences in age-related circadian misalignment (Fig. 2). Our results show that Per2 expression in the kidney remains the same in men across increasing age (6–24 mo) but significantly declines after middle age (12 mo) in women. Similar sex differences in the aging kidney have been demonstrated in humans with significant age-related increases in the morning of cortisol acrophase in women but not in men (61). Additionally, sex hormones have been found to play a role in the pathogenesis of kidney disease in men who develop decreased plasma testosterone levels (50) and in women who undergo premature menopause (2). Interestingly, in our animal study, Bmal expression did not change throughout the lifetime in the kidney. Our preliminary results demonstrate that male and female subjects differ in their circadian gene expression, particularly at mid and old age. This preliminary work warrants further research into the sex-dependent differences in circadian gene expression as well as what drives these changes over the lifespan.

Functionally, perhaps one of the best examples of deviant circadian rhythm in the aging kidney is nocturia. Defined as nighttime awaking of two or more times to urinate, nocturia is prevalent in >50% of men and women over the age of 60 yr (32). Individuals with nocturia excrete half of their total 24-h urinary Na⁺ at night compared with healthy young individuals, who secrete 1.5–2 times more Na⁺ during the day than at night (30). This reversal of circadian regulation of urine output is significant because nocturia is associated with poor sleep, increased fall risk, and increased daytime arterial blood pressure. Circadian biology may also impact older patient dialysis outcomes. In a retrospective study of close to 7,000 patients from the United States Renal Data System Dialysis Morbidity and Mortality Waves III/IV database, the time of day at which a patient received dialysis significantly impacted survival, but only in patients older than 60 yr (1). Although the relative risk for time of day was lower than the risk of mortality from preexisting conditions like diabetes, patients undergoing evening shift dialysis (6:00 PM–12:00 AM) had significantly better survival rates than those in the morning shift. This suggests that circadian regulation of blood pressure/volume as well as higher morning rates of myocardial infarction and hemorrhage may contribute to better survival outcomes in older patients.

PHYSICAL ACTIVITY AND THE KIDNEY

It is well understood that exercise is positively associated with decreases in chronic diseases such as cardiovascular disease, obesity, type II diabetes, and some forms of cancer (59). However, it is also becoming clearer that exercise directly improves or attenuates declining kidney function (5, 8, 9, 15). The benefits of exercise for the prevention of kidney function decline is mainly thought to be a function of a reduction in the comorbidities hyperglycemia/type II diabetes and hypertension. Elevated blood sugar is a cause of impaired kidney function because glucose itself activates proinflammatory/fibrotic signaling, eventually leading to diabetic nephropathy (17). Hypertension causes deterioration in kidney function through the renin-angiotensin-aldosterone system, oxidative stress, endothelial dysfunction, and genetic and epigenetic determinants (35). Chronic exercise participation reduced risk of both hyperglycemia (5) and hypertension (8), thus reducing risk for kidney dysfunction. Although these indirect effects of exercise are beneficial for kidney function, physical activity can also directly improve kidney function through attenuation of renal sympathetic nerve activity (RSNA). For example, Meredith et al. (35a) demonstrated that 1 mo of cycle ergometry performed by healthy but sedentary individuals was successful in reducing resting norepinephrine spillover (a measure of total peripheral sympathetic nerve activity) by 24%, which was primarily due to a 41% drop in renal norepinephrine spillover (9). The importance of these reductions in RSNA at rest and during exercise is that RSNA is positively associated with renal Na⁺ reabsorption. Therefore, reduced RSNA would result in decreased Na⁺ retention and, consequently, reduced blood pressure. Furthermore, decreased RSNA due to exercise training could attenuate renal vasoconstriction during exercise and reduce risk for acute renal failure during exercise (14).

Additionally, the benefits of exercise on kidney function also extend to individuals with compromised kidney function. For example, in patients with chronic kidney disease, low-intensity aerobic and resistance training is recommended to attenuate progression of the disease (28). Even low-intensity exercise performed during dialysis has been shown to increase the efficacy of and improve the 6-min walk test of patients undergoing hemodialysis (42). It was hypothesized that these patients had increased muscle blood flow resulting in an the increase flux of urea from the tissue to the vascular compartment, leading to an increase in serum clearance and eventual dialysis efficiency improvement (42). Furthermore, as discussed above, older individuals with compromised renal function likely also have increased sympathetic nervous system activity (51) and therefore would benefit from exercise training. Although experimental data has yet to demonstrate this, it is not impetuous to assume that the reduced RSNA demonstrated as a result of exercise training in younger adults (above) could also help to attenuate the reduced kidney function classically attributed to age. Although the known benefits of exercise in an individual with compromised kidney function have been documented, researchers are still determining optimal exercise timing (i.e., the time of day to exercise) as well as the ideal combination of resistance and aerobic exercise; however, both are recommended (28).

PHYSICAL ACTIVITY AND THE KIDNEY MOLECULAR CLOCK

Exercise-mediated entrainment of peripheral clocks as well as the underlying mechanisms that control molecular clock function in peripheral tissues (including the kidney) have yet to be well established. Most data to support the hypothesis that exercise can shift the molecular clock or that time of day of exercise can be manipulated for physiological outcomes, stem from studies in skeletal muscle (62). However, many of these conclusions remain contentious, with some groups reporting that scheduled exercise can shift the circadian clock in skeletal muscle (62) and other researchers finding no differences in the time of day of exercise (49). Thus, although we know that exercise timing plays a role in the entrainment of peripheral clocks, the molecular framework needs to be better defined.

Even less data exist with respect to the exercise-trained kidney. However, a survey of circadian signals responsible for enforcing rhythmicity across peripheral tissues suggests that the liver and kidneys are more sensitive to hormonal signals than skeletal muscle, heart, and spleen (20). Exercise releases the hormones renin, aldosterone, and AVP that can affect metabolic status, which could strengthen circadian alignment in the kidney, thereby maintaining and/or restoring healthy renal physiology (3). Mice given free access to a running wheel had more pronounced changes in metabolic parameters like lipid metabolism and energy balance. This suggests that signals that regulate peripheral circadian rhythms differ and that the kidney might be more responsive to blood-borne hormones than the skeletal muscle, thereby resulting in significant regulation of circadian rhythms (69). Although the mechanism(s) regulating exercise-mediated changes in renal circadian rhythm remain completely unstudied, our group is interested in the hormone AVP as a potential molecular target. AVP is under circadian control, as explained above. As individuals age, the diurnal rhythm of AVP expression and secretion are altered. Importantly, this change in rhythm of AVP expression also correlates with nocturic episodes (25). Endurance exercise is a well-established regulator of AVP (24); therefore, if both AVP regulation and water/Na⁺ balance are disrupted in the day/night cycles, perhaps AVP regulation by exercise may be a molecular target to restore rhythm. This novel hypothesis will allow researchers to better establish the role of AVP as a potential molecular target for resetting disrupted circadian rhythms in the kidney.

Exercise is a viable therapeutic intervention to reset the kidney molecular clock in an aging population because 1) there are no known negative effects of exercise, 2) there are several other benefits to exercise outside of preventing renal issues (i.e., better sleep, improved heart function, and improved muscle structure and function), and 3) exercise is a cost-effective tool that many older adults can easily partake in. If our hypothesis is true that aging contributes to misalignment in circadian rhythms, perhaps contributing to a decline in renal function, and exercise helps maintain a normal molecular clock, then exercise should improve kidney function in older populations (Fig. 3). In fact, in a cross-sectional study of ~1,300 men (78.5 ± 4.6 yr), physical activity reduced the odds ratio for lower GFR. Each additional 30 min of light physical activity or each 10-min moderate/vigorous physical activity was associated with lower odds of having a low GFR (41). Interestingly, the amplitude of temperature rhythm is dependent on the level of aerobic capacity in older participants (12). Older adults who engage in regular physical activity had improved circadian fluctuations of core temperature compared with those who did not, suggesting that better circadian regulation can lead to higher aerobic capacity and a better quality of life (12).

CONCLUSIONS AND FUTURE RESEARCH

The kidneys regulate several homeostatic mechanisms such as fluid balance, pH, and total body electrolyte concentration and do so dynamically under the regulation of the molecular clock. Aging not only impairs renal function but also dysregulates circadian control of these homeostatic mechanisms. Disrupted circadian rhythms in the kidney likely contribute to adverse health outcomes such as nocturia, hypertension, and increased risk for stroke, cardiovascular disease, and end-organ failure. Regular endurance exercise improves circadian alignment in both young and old individuals. Recent advances in the heart and skeletal muscle literature, although limited, suggest that regular endurance exercise entrains peripheral clocks, and we propose that similar beneficial adaptations occur in the kidney as well through regulation of renal blood flow, fluid balance, and blood pressure (Fig. 3). Future studies should address the importance of disrupted circadian rhythms in both young (e.g., as influenced by shift work) and old adults (aging) and identify mechanisms in which exercise can realign circadian rhythms. Furthermore, the molecular mechanisms remain poorly described for how exercise entrains peripheral clocks such as the kidney, and there remains a need to identify the mechanism(s) by which exercise helps the aging kidney. A proper functioning molecular clock will likely promote overall health, including in the kidneys, where blood pressure control and fluid and electrolyte balance are of utmost importance to health. Whether exercise will improve circadian function in these populations through central or peripheral mechanisms has yet to be determined, but we hypothesize that old mice will regain circadian rhythm response in the kidney due to positive physiological associated changes through exercise. The available evidence points to the potential that aiding in maintenance of renal circadian function may yet be added to the long list of benefits of lifelong physical activity.

GRANTS

The research reported in this publication was supported by the Jackson Aging Center: Nathan Shock Centers of Excellence in the Basic Biology of Aging Pilot Award P30-AG-038070 (to E. E. Schmitt and D. R. Bruns). In addition, this publication was made possible by an Institutional Development Award from the National Institute of General Medical Sciences under Grant 2-P20-GM-103432 as well as by funds from the Division of Kinesiology and Health from the University of Wyoming.

DISCLOSURES

No conflicts of interest, financial or otherwise, are declared by the authors.

AUTHOR CONTRIBUTIONS

E.E.S., E.C.J., and D.R.B. conceived and designed research; M.Y. performed experiments; E.E.S., M.Y., and D.R.B. analyzed data; E.E.S. and D.R.B. interpreted results of experiments; E.E.S. and D.R.B. prepared figures; E.E.S. drafted manuscript; E.E.S., E.C.J., M.Y., and D.R.B. edited and revised manuscript; E.E.S., E.C.J., M.Y., and D.R.B. approved final version of manuscript.