Hypothesis / aims of study
The bladder wall is considered as an effective barrier to substantial movements of water and electrolytes, in part to separate underlying tissues from the hostile environment of urine. Important in this barrier function is the low, but nevertheless finite, water and electrolyte permeability of the urothelium. Moreover, the demonstration of urothelial aquaporins [1], and Na ion channels subject to mineralocorticoid control [2] suggests controlled transport of water and salts is possible. This is also consistent with a reduction in the volume of a saline infusion instilled into the rat bladder over several hours, a phenomenon not repeated if soybean oil was used to fill the bladder [3]. However, it is unclear if the human bladder has a significant capacity to transport salt and water from the bladder lumen and thus decrease salt and water excretion. A reason to address this question is to quantify base-line transport capacity as a prelude to understand whether this is diminished in subjects that suffer nocturia. The hypothesis under examination was that increased urine storage time decreases urinary salt and water daily excretion. The aim of the study was to determine, in healthy human volunteers, if increased bladder storage time of urine from three to five hours diminished daily excretion of water, Na and K.
Study design, materials and methods
Eight, healthy undergraduate student participants (six females, two males; mean age 20 years) were recruited after informed consent; the study was given ethical committee approval. Exclusion criteria were: self-reported dysuria or urinary tract infection, or conditions likely to result in polyuria. Participants were asked to maintain a normal life-style, avoiding severe exercise and changes to drinking habits, especially caffeine-containing drinks. Alcohol intake was confined to ≤14 units, spaced equally throughout the week. Participants filled out a bladder diary stating time and volume of each void and a record of fluid intake to calculate total fluid intake (V(in)) and total urine output (V(out)), ml/day. A bladder-training leaflet was provided to facilitate participants with maintaining prolonged urine storage in the bladder - see protocol.
On day-1 and day-2 the interval between voids was set at three-hours (six voids/day, between 8:00 and 23:00): after a rest day, on day-3 and day-4 the interval was five hours (four voids/day). No nocturnal voids were reported. At each void a mid-stream urine sample (20 ml) was stored at 4°C for subsequent urinalysis. Urine samples were analysed for: total osmolality (mOsm/kg; Osmocheck, Advanced Instruments, UK); [Na] and [K] (mmol/l; Cole Palmer, USA) at room temperature (20-22°C). Osmometers were calibrated with mixed NaCl/KCl solutions up to 1000 mOsm/kg) to measure total urine osmolality and used to estimate daily osmolal urinary losses (U(osm,T), mOsm/day). Ion-selective electrodes (ISEs) were calibrated with mixed NaCl/KCl solutions (total osmolality 400 mOsm/kg) to measure urinary [Na] and [K] and used to estimate daily urinary losses of Na (U(Na)) and K (U(K)) as mmol/day. The contribution of urinary NaCl and KCl to total urinary osmolality (U(osm,Na+K)) was calculated as 2*(U(Na)+U(K)), as Cl ions are the major anion in urine. Non-NaCl/KCl urine osmolality (U(osm,*)) is therefore U(osm,T) - (U(osm,Na+K)).
Data from day-1 and day-2 collections and from day-3 and day-4 collections were averaged for each participant and group data expressed as median values [25% and 75% interquartiles]. 3-hour protocol data are always quoted first in the text. Significance between data sets from 3-hour and 5-hour paired samples were analysed by Wilcoxon signed-rank tests. The null hypothesis was rejected at p<0.05. A previous power calculation of U(K) measurements with ISEs indicated reproducible data with 80% power required at least n=6 participants.
Results
Reported fluid intake (V(in)) was not significantly different between the 3-hour and 5-hour storage protocols: these were 1408 [1331,1584] vs 1362 [1225,1530] ml/day, respectively, n=8; p=0.438. However, daily voided volume (V(out)) was significantly less with 5-hour storage in all participants; Figure 1A (1723 [1528,1954] vs 1158 [1095,1329] ml/day, n=8; p=0.008).
Daily losses of urinary Na and K were also significantly less with the 5-hour storage protocol. Data for Na (Figure 1B) were 134 [102,205] vs 88 [53,99] mmol/day respectively; n=8, p=0.008. Data for K (Figure 1C) were 42 [35,49] vs 22 [18,29] mmol/day respectively; n=8, p=0.014. As with voided volume data, the reduction of urinary Na and K excretion with the 5-hour storage protocol was seen in all participants.
Total urine osmolal (U(osm,T)) losses were not significantly different between the 3-hour and 5-hour storage protocols (Figure 1D); values were, respectively 685 [573,713] vs 618 [541,669] mOsm/day; n=8, p=0.195. However, non Na/K urinary losses (U(osm,*)) were greater in the 5-hour storage protocol (seven of eight participants) as determined by greater values of (U(osm,*)) compared with values for the 3-hour storage protocol, Figure 1E (239 [191,354] vs 382 [366,447] mOsm/day; n=8, p=0.047).
Interpretation of results
Data are compatible with the hypothesis that Na, K and water reabsorption is possible from the lower urinary tract, possibly across the bladder wall, as stored urine spends most time here after leaving the kidneys. The magnitude of these movements is large enough to modify urine volume and electrolyte content. There is abundant evidence from other studies that regulation of ionic Na and K transport by mineralocorticoids is possible. Water movement is likely to be by osmosis, but it cannot be ascertained from this study if it is facilitated by aquaporins. Of interest is that total urine osmolality remained unchanged with prolonged storage so that greater Na and K reabsorption is counter-balanced by augmented excretion of other osmolytes, possibly urea.
The data are interesting in the context of nocturia. Salt and water reabsorption during prolonged nocturnal urine storage would lessen bladder volume and consequent perceptions of bladder filling to potentially reduce the chance of a nocturia episode. Attenuation of reabsorption might therefore enhance the likelihood of episodes.