J. Physiol. (1976), 256, pp. 93-102 With 4 text-figure Printed in Great Britain
93
THE EXCRETION OF UREA BY DOGS FOLLOWING A MEAT MEAL
BY W. J. O'CONNOR AND R. A. SUMMERILL* From the Department of Physiology, University of Leeds, Leeds LS2 9JT
(Received 29 May 1975) SUMMARY
1. After a meal of meat (10 g/kg), urea excretion in dogs increased by about 200%, plasma urea by 60% and exogenous creatinine clearance by 40 % in comparison with control experiments. 2. Urea, given by stomach tube in doses producing the same increase in plasma urea, caused urea excretion to increase by only 90 %, with no increase in creatinine clearance. With the increased glomerular filtration rate after meat there was added excretion of urea. 3. In control experiments and after urea, the rate of excretion of urea was directly proportional to plasma urea. The ratio urea clearance/creatinine clearance was 0 45. 4. After meat, urea clearance increased more than creatinine clearance, the ratio increasing to 0 55, i.e. a smaller fraction of the filtered urea was re-absorbed after meat. 5. After meat, 10 g/kg, the rate of urea production rose to 230600 ,tmole/min for 4-6 hr.
INTRODUCTION
In the preceding paper, O'Connor & Summerill (1976) showed that following a normal meat meal eaten by dogs, glomerular filtration rate increased by about 40 %. With the fact of increased glomerular filtration thus established, this paper examines the contribution of increased glomerular filtration to the increased excretion of urea which follows the meat meal. Two groups of experiments are compared: firstly after meat there was increased plasma urea, increased glomerular filtration and increased * Present address: Department of Physiology, University of Dar es Tanzania.
Salaam,
94 W. J. O'CONNOR AND R. A. SUMMERILL excretion of urea; secondly, experiments are described in which urea given by stomach tube caused increased plasma urea and increased excretion of urea, without increase in glomerular filtration. Comparison of the two sets of experiments should reveal any contribution of increased glomerular filtration to the increased excretion of urea. Some of the results have been communicated to the Physiological
Society (Summerill, 1974). METHODS
The results reported in this paper are the determinations of urea in the same or similar experiments to those of O'Connor & Summerill (1976); the methods are described in that paper. One hour before the experiment began, 1 g creatinine in 50 ml. water was given by stomach to allow measurement of exogenous creatinine clearance as discussed by O'Connor & Summerill (1976). After initial collections of urine and blood, the dog ate 10 g/kg lean beef and urea excretion was followed for 6-S8 hr. Blood samples were drawn from the carotid artery approximately 2, 4, 6 and 8 hr after the meat (Fig. 1). Determination of creatinine in urine and plasma allowed calculation of exogenous creatinine clearance at these times; this paper reports the changes in urea excretion and in plasma urea. In experiments where urea was given, the first dose was given instead of the meat and further doses were given 2 and 4 hr later, as in Fig. 2. Urea. This was determined in plasma and urine by the Autoanalyser method N-1C (Technicon, 1968), based on the direct reaction of urea and diacetyl monoxime in the presence of thiosemicarbazide under acid conditions giving a pink coloured complex measured at 520 nm. The 95 % confidence limits by the method of Kimball (1952) were 1 %. The method has been fully examined in this laboratory by Golob (1974); it is apparently specific for urea and not subject to interference by other substances in the urine of dogs. RESULTS
Urea excretion after meat, 10 g/kg Fig. 1 A shows the increase in urea excretion after meat in one dog; 10 g/kg is about the ordinary daily meat ration of our dogs and therefore this dose was chosen for detailed investigation. In Fig. 1 A, urea excretion was at its highest 4-6 hr after the meat was eaten, and at this time the plasma urea was increased by 5 m-mole/l. above the corresponding value in control experiments. In control experiments where no meat was given there was a small fall in plasma urea and urea excretion. In a previous paper, O'Connor & Summerill (1976) described the increase in exogenous creatinine clearance following meat, and this is shown at the top of Fig. 1 B. The data in Fig. 1 A allow the calculation of urea clearance (U V/P) where: U V is the rate of excretion and P plasma concentration of urea. The urea clearance is shown at the bottom of Fig. 1 B. Urea and creatinine clearance increased together after the meat.
UREA EXCRETION AFTER MEAT 95 The results in Fig. 1 are typical of thirty-six experiments in which meat (10 g/kg) was given to seven dogs. Most of the experiments were on three dogs as summarized in Table 1 which shows urea excretion 3j-4 hr after the meat in comparison with control experiments. In all cases plasma urea, urea excretion, urea and creatinine clearances increased. The ratio
E L
E
5_0
= IV
CL
-L
E 0 c
E
L
-
to
E
.
_
,, 20 10
0
2
4
6
8 0 Time after meat (hr)
2
4
6
8
Fig. 1. Lassie, 17-18 kg. Urea excretion after meat 10 g/kg. Abscissa time after meat, in hr. A, and *, urea excretion and plasma urea in three experiments in which meat was given: ----- and 0 in five control experiments. Mean and 5.E. of mean are shown. B, creatinine clearance (above) and urea clearance (below) in the same experiments. TABLE 1. Urea excretion of three dogs
3j74 hr after meat, 10 g/kg.
For each
meter the upper figure is the value after meat, and the lower the value at the
time in control experiments: stated
Plasma
urea
mean, s.E.
(m-molefl.)
Urea excretion
(jumole/min)
Urea clearance (ml./min) Creatinine clearance
(ml./min)
of
Lassie 17-18 kg 10*2 + 0 3 (10) 53 ± 0-3 (8) 342 + 26 92+ 9
33-3+2±1 17-2 + 1-5 54-6 ± 2-0
37-7±1-3 Ratio: urea clearance/creatinine clearance Urine volume (ml./min)
mean
0-61 + 0-02 0.46 ± 0.04 0-25 ± 0-02 013 + 0.01
parasame
and number of experiments is
Cathy 13-5-14-5 kg 11.1+ 0*5 (9) 7*5± 0 3 (7) 239+ 15
83±12 21-6± 1-2 11*0+± 1-3 455±2*5 31-0± 1*2 0*48 ± 0-02 0*35 0*03 0.19 0.01 0.10 + 0*01
Cleo 14-15 kg 7-1 0 4 (10) 4-4 ± 0 3 (9) 238 24 93+ 8 33-4 ± 2-6 21-2+1 1
54-5+3±1 42-9 ± 2*3 61 ± 0-02 0.50 ± 0.01 0*24 + 0-02 0-12 + 0-02 0
PH Y 256
96 W. J. O'CONNOR AND R. A. SUMMERILL urea clearance/creatinine clearance also increased after meat, i.e. a greater fraction of the filtered urea appeared in the urine. The urine volume remained low (0.25 ml./min in Table 1) in contrast with most experimental work of this kind where by administration of water or saline urine volumes of 2 5 ml./min or much more were maintained throughout the experiments.
Urea excretion after doses of urea by stomach tube Fig. 2 shows two examples of the effect of urea given by stomach tube. The urea was given in three doses to mimic the slow release of urea after meat and by trial with each dog a dose was found which gave increase in plasma urea similar to that after meat. In the examples of Fig. 2A this
:0=
A
_
50 _ B
Creatinine Fig.eatinine 2 a
6 0
200
,EiO10
A
.
,
A
10
wasablA
2 4 0 6 8 6 8 2 4 0 ateac -ro.B raiieadue Time laacs (hr) Fig. 2. Lassie. Urea excretion when urea was given by stomach tube; , A and.,--- A are urea excretion plotting similar to Fig. 1. A, and plasma urea in two experiments. Doses of 33 in-mole urea were given at each arrow. B, creatinine and urea clearances.
achieved by three doses of 33mi-mole dissolved in 20 ml. water. Despite the similar increases in plasma urea, the rate of excretion of urea after doses of urea (Fig. 2A) was less than after meat (Fig. 1A). There was no increase in either creatinine or urea clearance after the doses of urea (Fig. 2B). Urea was given to Lassie in seven experiments providing the data plotted in Fig. 3. After doses of urea (0) and in control experiments ( x ) urea excretion was proportional to plasma urea, these points in Fig. 3 all falling close to a linear relationship projecting through the origin. The slope of the line (UV/P) in Fig. 3 is 17-4 ml./min in comparison with a mean creatinine clearance in this animal of 39 ml/min (Tables 1, 2). Also
was
UREA EXCRETION AFTER MEAT 97 shown in Fig. 3 (0) is the data 3j-4 and 5j-6 hr after meat, demonstrating the raised urea clearance. Similar results were obtained when urea was given to Cathy and Cleo; the data in Table 2 are the data for each animal at 31-4 hr in the experimental plan of Fig. 2 after two doses of urea. The same conclusions are reached by examination of the data after the first and third doses. Urea excretion after doses of urea was less than after meat, despite the similar S
400
's2 300 0
E
.o4. 200 D1C100
x
U
2
4
6 10 12 8 Plasma urea (m-mole/l.)
14
16
Fig. 3. Lassie. Urea excretion (ordinate, fmole/min) plotted against plasma urea (abscissa, m-molefl.) Points at 10.10 hr each day and in eight control experiments shown by x; 0 in seven experiments after the administration of urea by stomach tube. The regression line is that calculated from these values. Points, 0, are 4-6 hr after meat (ten experiments).
TABi 2. Urea excretion of three dogs after two doses of urea, at the time 3j-4 hr in the plan of Fig. 2: mean, 5.E. of mean and number of experiments Lassie Cathy Cleo 11-2 ±0 4 (7) 10-9 ± 0*7 (4) Plasma urea (m-mole/l.) 8-2± 0-5 (4) 212 ± 14 146 + 22 Urea excretion (m-mole/min) 189 + 21 190+ 1-6 13-4± 1-8 Urea clearance (ml./min) 23-2 ± 2-2 40-3 + 1-7 36-1 + 2-5 41-9 ± 2-5 Creatinine clearance (ml./min) Ratio: urea clearance/creatinine clearance Urine volume (ml./min)
0-47 ± 0-03
0-37 ± 0-03
0-50 ± 0-03
0-17 + 0-03
0-14+ 0-02
0-21 ± 0-01 4-2
98 W. J. O'CONNOR AND R. A. SUMMERILL changes in plasma urea (compare Table 1 and 2). Following urea there was no significant change in urea or creatinine clearance (compare control experiments in Table 1), and no significant increase in the ratio urea clearance/creatinine clearance. In the urea as in the meat experiments the urine volume remained low.
Excretion of electrolytes Table 3 lists the excretion of electrolytes 3 5-4 hr after meat, at the corresponding times in control experiments and in the experiments where urea was given. After meat there were increases; on complete metabolism 170 g meat would yield about 10 m-mole PO0, 12 m-mole K, 6 m-mole Na, 3 m-mole Cl and about one-third of each of these was excreted during the TABLE 3. Electrolyte excretion at 3j-4 hr after meat, after two does of urea and in control experiments: mean, S.E. of mean and n the number of experiments on three on three dogs listed in Tables 1 and 2 Meat Urea Control n= 28 n= 13 n= 22 9.1 + 1.9 6-0 + 0 9 Na excretion (#ctmole/min) 18-5 + 2-2 6 1 + 0.7 K excretion (#mole/min) 28-8 + 1-8 8-4 ± 1.5 4-4 + 0-6 12-1 + 1 7 5.5 + 1-2 Cl excretion (,umole/min) 5.9 + 1.0 3.6 + 0 6 17.6 ± 1-4 PO4 excretion (,umole/min)
6 hr after the meat. After urea, apparent increases in the rate of excretion of electrolytes are of doubtful significance and were certainly small in comparison with the increases after meat.
Production of urea after a meat meal Urea produced in the body is either excreted in the urine or retained dissolved in the total body water. The amount excreted was directly measured; e.g. in the average of Fig. 1A the amount excreted in the first 2 hr after the meal of meat was 19 m-mole. At this time plasma urea had increased from 6-12 to 8-28, i.e. by 2-16 m-mole/l. If it be assumed that urea had equilibrated throughout total body water of 111., then 24 m-mole urea had been retained and the total urea production in the 2 hr after the meal was 43 m-mole. By 4 hr after the meal 48 m-mole had been excreted and plasma urea was increased by 3-95 m-mole/l. indicating a retention of 43 m-mole and a total production in the 4 hr of 91 m-mole. This calculation has been applied to all stages of the experiments of Fig. 1 A to give Fig. 4 where the estimated total urea production is plotted against time. In control experiments with Lassie the production in 8 hr was 23 m-mole. In Fig. 4 in the first 4 hr after the meat the average rate of production was 380 ,smole/min and in control experiments 54 ,umole/min.
UREA EXCRETION AFTER MEAT 99 The rate of urea production decreased after 6 hr and in the period 6-8 hr after the meat only 13 m-mole was produced, 110 /zmole/min, still a little more than in control experiments. With other dogs similar rates of urea production were estimated, the analysis for Cleo and Cathy being included in Fig. 4. In seven dogs the estimated urea production during the first 4 hr after meat (10 g/kg) was 56-150 m-mole, equal to 230-600 iumole/min. 150 I,
0
E C
100
0
e
U
0
0~~~~~~~~~0
4,~~~~0 A
0
0
0
2
4 6 8 Time after meat (hr) Fig. 4. The estimated production of urea after meat, and in control experiments. Abscissa, time after meat: ordinate, accumulated production of urea, in-mole. * and 0 are meat and control experiments in Lassie; A and A are meat and control experiments in Cathy or Cleo, the results being indistinguishable. Mean and s.E. of mean is shown by the bars or by the width of the symbols in control experiments.
After 175 g meat given to Lassie the estimated urea production in 6 hr was 132 m-mole (Fig. 4) and in control experiments 21 m-mole. The total N2 (urea equivalence) of the protein in the meat was 245 m-mole so that the increased urea production in the 6 hr represented 45 % of the total N2 in the meat. In the case of Cleo and Cathy increased urea production in 6 hr after meat amounted to 30 % of its N2. The calculation of urea production depends on the assumption that at the times of blood sampling urea was evenly distributed through the body water and the total body water of Lassie, weight 17-18 kg, was correctly assumed to be 111. (Gaudino & Levitt, 1949; Edelman, 1952). That the method is essentially correct was substantiated when the same calculations were applied to the experiments where known amounts of urea were given by stomach tube (e.g. Fig. 2); the estimated urea 'production' 2 hr after ingestion of doses of urea was 102 % + 2 (s.E. of mean, n = 20), 102 % ± 3 (11), 91 % ± 5 (12) of the administered amount in Lassie, Cathy and Cleo
respectively.
100
W. J. O'CONNOR AND R. A. SUMMERILL DISCUSSION
From the analysis in Fig. 4 it seems that following ingestion of meat the sequence of digestion, absorption of amino acids, deamination and formation of urea reached a rate of urea formation of 230-600 #tmole/min. Somewhere along the sequence: absorption of amino acids; deamination; urea formation; unidentified chemical substances released into the circulation (or other mechanisms) have the effect of increasing glomerular filtration rate (for discussion see O'Connor & Summerill, 1976). Increased glomerular filtration rate itself causes increased urea excretion, with increased urea clearance. However, in our experiments the filtration rate did not increase sufficiently to cause urea excretion to match the increased urea production; urea therefore accumulated in the body with rising plasma urea further increasing urea excretion until production was matched. The ultimate mechanism by which the rate of production of urea was equalled by excretion was increasing plasma urea; the necessary increase in plasma urea was however less by the fact that glomerular filtration rate increased at the time of protein catabolism. Before the work of this and the preceding paper only very large meat meals have been shown to increase glomerular filtration rate. Now that a substantial increase has been demonstrated with moderate sized meat meals, it seems that in the dog, increased glomerular filtration after a meat meal may be of importance in normal function. However, the dog is unique amongst species commonly used for investigation of renal physiology in that it is usually maintained on a single daily meal of high protein content. On a weight basis the similar routine for man would be a single daily meal including 650 g lean beef. Even if the protein intake of man were of this order (140 g/day) it would be spread over three meals a day. There seems to have been no deliberate investigation in man of the possibility that the meals may be an important cause of the higher clearances during the day as compared to night (Wesson, 1964) or that the changes in clearances with changes in dietary protein may contribute differences in renal excretion of urea and electrolytes. Schmidt-Nielson (1958) has reviewed evidence relating glomerular filtration rate and urea excretion in several species. The experiments in which urea was given add to an extensive literature on urea excretion, reviewed by Schmidt-Nielsen (1958). Our experiments differ from almost all of the published work in that they were carried out at the normal rates of urine flow of dogs, i.e. 0-1-0-3 ml./min as defined by O'Connor & Potts (1969) and the doses of urea have been small enough for urea excretion to remain within the normal range and this leads to some differences between our results and the usual findings.
101 UREA EXCRETION AFTER MEAT With large doses of urea, Mudge, Foulkes & Gilman (1949) and Anslow & Wesson (1955) produced osmotic diuresis and increased excretion of electrolytes whereas with our small doses any such changes were small. Urine volume after urea remained below 0-25 ml./min (Table 2) and there was no significant change in electrolyte excretion in comparison with control experiments (Table 3). The only effect with small doses of urea was that urea excretion increased. With regard to urea excretion our results with small doses of urea emphasize that the rate of excretion of urea is predominantly determined by plasma urea concentration (Fig. 3) and by glomerular filtration rate when this changes after meat. Since the urine volume remained low, it does not form a major determinant of urea excretion as has been suggested from the earlier literature. The only investigation of urea excretion at normal low urine flows is contained in the paper of Shannon (1936). At urine flows of about 0-15 ml/ min in his experiments, urea clearance/creatinine clearance was 0 3 and increased progressively to 0-6 as the urine volume was increased by the administration of water. The interpretation of Shannon (1936) and also Mudge, Berndt & Valtin (1973) is that at higher urine flows there is less back diffusion of urea; this must be an over-simplification in view of the supposed roles of different parts of the nephron indicated by the micropuncture work reviewed by Mudge et at. (1973). In our experiments, with the urine volume 0 1-0 3 ml./min both in control experiments and after urea, urea clearance/creatinine clearance was 0 35-0-55 which is higher than in Shannon's (1936) results at similar low urine flow. We have recorded the effects of water on only a few occasions and when urine volume was thus raised above 1 ml./min the ratio increased to 0-6 as in Shannon's experiments. After meat with the urine flow only 0-25 ml./min, the ratio rose to 0-5-0-6 (Table 1), whereas at similar urine flow, Shannon (1936) observed ratios of 0'3-0 4. The increased ratio after meat does not seem to be due solely to the small increase in urine volume; it could be associated with the increased excretion of electrolytes. It seems that decreased re-absorption of urea is a secondary effect of the increased glomerular filtration, which adds to urea excretion after meat, as compared to after doses of urea. This aspect of urea excretion has been particularly considered by Schmidt-Nielsen (1958). The authors are greatly indebted to Mr J. Brook and Mrs S. Snack for their technical assistance.
102
W. J. O'CONNOR AND R. A. SUMMERILL
REFERENCES ANSLOW, W. P. & WEssoN, L. G. (1955). Effect of sustained, graded urea diuresis on water and electrolyte excretion. Am. J. Phy8iol. 180, 605-611. EDELMAN, I. S. (1952). Exchange of water between blood and tissues. Characteristics of deuterium oxide equilibration in body water. Am. J. Phy8iol. 171, 279-296. GAUDrNO, M. & LEVITT, M. F. (1949). Inulin space as a measure of extracellular fluid. Am. J. Phy8iol. 157, 387-393. GOLOB, P. (1974). The effect on water, electrolyte and nitrogen balance of overfeeding dogs. Ph.D. Thesis, Leeds University. KIMBALL, A. W. (1952). Answer to query. Biometric8 8, 85-86. MUDGE, G. H., BERNDT, W. 0. & VALTIN, H. (1973). Tubular transport of urea, glucose, phosphate, uric acid, sulphate and thiosulphate. In Handbook of Phy8iology, section 8: Renal Physiology, chap. 19, pp. 587-652. Washington: American Physiological Society. MUDGE, G. H., FOULEms, J. & GILMAN, A. (1949). Effect of urea diuresis on renal excretion of electrolytes. Am. J. Phy8iol. 158, 218-230. O'CONNOR, W. J. & PoTTs, D. J. (1969). The external water exchanges of normal
laboratory dogs. Qt JZ exp. Phy8iol. 54, 244-265. O'CONNOR, W. J. & SUMMERILL, R. A. (1976). The effect of a meal of meat on glomerular filtration rate in dogs at normal urine flows. J. Phy8iol. 256, 81-91., SHANNON, J. A. (1936). Glomerular filtration and urea excretion in relation to urine flow in the dog. Am. J. Phyaiol. 117, 206-225. SCHMIDT-NIELSEN, B. (1958). Urea excretion in mammals. Phy8iol. Rev. 38, 139-169. SUMMERILL, R. A. (1974). The effects of a meat meal on glomerular filtration rate and urea excretion in the conscious dog. J. Physiol. 238, 64P. TECHNICON (1968). Auto-analy8er Handbook. Houndmills, Basingstoke, Hants: Technicon Instruments. WESSON, L. G. (1964). Electrolyte excretion in relation to diurnal cycles of renal function. Medicine, Baltimore 43, 547-592.
93
THE EXCRETION OF UREA BY DOGS FOLLOWING A MEAT MEAL
BY W. J. O'CONNOR AND R. A. SUMMERILL* From the Department of Physiology, University of Leeds, Leeds LS2 9JT
(Received 29 May 1975) SUMMARY
1. After a meal of meat (10 g/kg), urea excretion in dogs increased by about 200%, plasma urea by 60% and exogenous creatinine clearance by 40 % in comparison with control experiments. 2. Urea, given by stomach tube in doses producing the same increase in plasma urea, caused urea excretion to increase by only 90 %, with no increase in creatinine clearance. With the increased glomerular filtration rate after meat there was added excretion of urea. 3. In control experiments and after urea, the rate of excretion of urea was directly proportional to plasma urea. The ratio urea clearance/creatinine clearance was 0 45. 4. After meat, urea clearance increased more than creatinine clearance, the ratio increasing to 0 55, i.e. a smaller fraction of the filtered urea was re-absorbed after meat. 5. After meat, 10 g/kg, the rate of urea production rose to 230600 ,tmole/min for 4-6 hr.
INTRODUCTION
In the preceding paper, O'Connor & Summerill (1976) showed that following a normal meat meal eaten by dogs, glomerular filtration rate increased by about 40 %. With the fact of increased glomerular filtration thus established, this paper examines the contribution of increased glomerular filtration to the increased excretion of urea which follows the meat meal. Two groups of experiments are compared: firstly after meat there was increased plasma urea, increased glomerular filtration and increased * Present address: Department of Physiology, University of Dar es Tanzania.
Salaam,
94 W. J. O'CONNOR AND R. A. SUMMERILL excretion of urea; secondly, experiments are described in which urea given by stomach tube caused increased plasma urea and increased excretion of urea, without increase in glomerular filtration. Comparison of the two sets of experiments should reveal any contribution of increased glomerular filtration to the increased excretion of urea. Some of the results have been communicated to the Physiological
Society (Summerill, 1974). METHODS
The results reported in this paper are the determinations of urea in the same or similar experiments to those of O'Connor & Summerill (1976); the methods are described in that paper. One hour before the experiment began, 1 g creatinine in 50 ml. water was given by stomach to allow measurement of exogenous creatinine clearance as discussed by O'Connor & Summerill (1976). After initial collections of urine and blood, the dog ate 10 g/kg lean beef and urea excretion was followed for 6-S8 hr. Blood samples were drawn from the carotid artery approximately 2, 4, 6 and 8 hr after the meat (Fig. 1). Determination of creatinine in urine and plasma allowed calculation of exogenous creatinine clearance at these times; this paper reports the changes in urea excretion and in plasma urea. In experiments where urea was given, the first dose was given instead of the meat and further doses were given 2 and 4 hr later, as in Fig. 2. Urea. This was determined in plasma and urine by the Autoanalyser method N-1C (Technicon, 1968), based on the direct reaction of urea and diacetyl monoxime in the presence of thiosemicarbazide under acid conditions giving a pink coloured complex measured at 520 nm. The 95 % confidence limits by the method of Kimball (1952) were 1 %. The method has been fully examined in this laboratory by Golob (1974); it is apparently specific for urea and not subject to interference by other substances in the urine of dogs. RESULTS
Urea excretion after meat, 10 g/kg Fig. 1 A shows the increase in urea excretion after meat in one dog; 10 g/kg is about the ordinary daily meat ration of our dogs and therefore this dose was chosen for detailed investigation. In Fig. 1 A, urea excretion was at its highest 4-6 hr after the meat was eaten, and at this time the plasma urea was increased by 5 m-mole/l. above the corresponding value in control experiments. In control experiments where no meat was given there was a small fall in plasma urea and urea excretion. In a previous paper, O'Connor & Summerill (1976) described the increase in exogenous creatinine clearance following meat, and this is shown at the top of Fig. 1 B. The data in Fig. 1 A allow the calculation of urea clearance (U V/P) where: U V is the rate of excretion and P plasma concentration of urea. The urea clearance is shown at the bottom of Fig. 1 B. Urea and creatinine clearance increased together after the meat.
UREA EXCRETION AFTER MEAT 95 The results in Fig. 1 are typical of thirty-six experiments in which meat (10 g/kg) was given to seven dogs. Most of the experiments were on three dogs as summarized in Table 1 which shows urea excretion 3j-4 hr after the meat in comparison with control experiments. In all cases plasma urea, urea excretion, urea and creatinine clearances increased. The ratio
E L
E
5_0
= IV
CL
-L
E 0 c
E
L
-
to
E
.
_
,, 20 10
0
2
4
6
8 0 Time after meat (hr)
2
4
6
8
Fig. 1. Lassie, 17-18 kg. Urea excretion after meat 10 g/kg. Abscissa time after meat, in hr. A, and *, urea excretion and plasma urea in three experiments in which meat was given: ----- and 0 in five control experiments. Mean and 5.E. of mean are shown. B, creatinine clearance (above) and urea clearance (below) in the same experiments. TABLE 1. Urea excretion of three dogs
3j74 hr after meat, 10 g/kg.
For each
meter the upper figure is the value after meat, and the lower the value at the
time in control experiments: stated
Plasma
urea
mean, s.E.
(m-molefl.)
Urea excretion
(jumole/min)
Urea clearance (ml./min) Creatinine clearance
(ml./min)
of
Lassie 17-18 kg 10*2 + 0 3 (10) 53 ± 0-3 (8) 342 + 26 92+ 9
33-3+2±1 17-2 + 1-5 54-6 ± 2-0
37-7±1-3 Ratio: urea clearance/creatinine clearance Urine volume (ml./min)
mean
0-61 + 0-02 0.46 ± 0.04 0-25 ± 0-02 013 + 0.01
parasame
and number of experiments is
Cathy 13-5-14-5 kg 11.1+ 0*5 (9) 7*5± 0 3 (7) 239+ 15
83±12 21-6± 1-2 11*0+± 1-3 455±2*5 31-0± 1*2 0*48 ± 0-02 0*35 0*03 0.19 0.01 0.10 + 0*01
Cleo 14-15 kg 7-1 0 4 (10) 4-4 ± 0 3 (9) 238 24 93+ 8 33-4 ± 2-6 21-2+1 1
54-5+3±1 42-9 ± 2*3 61 ± 0-02 0.50 ± 0.01 0*24 + 0-02 0-12 + 0-02 0
PH Y 256
96 W. J. O'CONNOR AND R. A. SUMMERILL urea clearance/creatinine clearance also increased after meat, i.e. a greater fraction of the filtered urea appeared in the urine. The urine volume remained low (0.25 ml./min in Table 1) in contrast with most experimental work of this kind where by administration of water or saline urine volumes of 2 5 ml./min or much more were maintained throughout the experiments.
Urea excretion after doses of urea by stomach tube Fig. 2 shows two examples of the effect of urea given by stomach tube. The urea was given in three doses to mimic the slow release of urea after meat and by trial with each dog a dose was found which gave increase in plasma urea similar to that after meat. In the examples of Fig. 2A this
:0=
A
_
50 _ B
Creatinine Fig.eatinine 2 a
6 0
200
,EiO10
A
.
,
A
10
wasablA
2 4 0 6 8 6 8 2 4 0 ateac -ro.B raiieadue Time laacs (hr) Fig. 2. Lassie. Urea excretion when urea was given by stomach tube; , A and.,--- A are urea excretion plotting similar to Fig. 1. A, and plasma urea in two experiments. Doses of 33 in-mole urea were given at each arrow. B, creatinine and urea clearances.
achieved by three doses of 33mi-mole dissolved in 20 ml. water. Despite the similar increases in plasma urea, the rate of excretion of urea after doses of urea (Fig. 2A) was less than after meat (Fig. 1A). There was no increase in either creatinine or urea clearance after the doses of urea (Fig. 2B). Urea was given to Lassie in seven experiments providing the data plotted in Fig. 3. After doses of urea (0) and in control experiments ( x ) urea excretion was proportional to plasma urea, these points in Fig. 3 all falling close to a linear relationship projecting through the origin. The slope of the line (UV/P) in Fig. 3 is 17-4 ml./min in comparison with a mean creatinine clearance in this animal of 39 ml/min (Tables 1, 2). Also
was
UREA EXCRETION AFTER MEAT 97 shown in Fig. 3 (0) is the data 3j-4 and 5j-6 hr after meat, demonstrating the raised urea clearance. Similar results were obtained when urea was given to Cathy and Cleo; the data in Table 2 are the data for each animal at 31-4 hr in the experimental plan of Fig. 2 after two doses of urea. The same conclusions are reached by examination of the data after the first and third doses. Urea excretion after doses of urea was less than after meat, despite the similar S
400
's2 300 0
E
.o4. 200 D1C100
x
U
2
4
6 10 12 8 Plasma urea (m-mole/l.)
14
16
Fig. 3. Lassie. Urea excretion (ordinate, fmole/min) plotted against plasma urea (abscissa, m-molefl.) Points at 10.10 hr each day and in eight control experiments shown by x; 0 in seven experiments after the administration of urea by stomach tube. The regression line is that calculated from these values. Points, 0, are 4-6 hr after meat (ten experiments).
TABi 2. Urea excretion of three dogs after two doses of urea, at the time 3j-4 hr in the plan of Fig. 2: mean, 5.E. of mean and number of experiments Lassie Cathy Cleo 11-2 ±0 4 (7) 10-9 ± 0*7 (4) Plasma urea (m-mole/l.) 8-2± 0-5 (4) 212 ± 14 146 + 22 Urea excretion (m-mole/min) 189 + 21 190+ 1-6 13-4± 1-8 Urea clearance (ml./min) 23-2 ± 2-2 40-3 + 1-7 36-1 + 2-5 41-9 ± 2-5 Creatinine clearance (ml./min) Ratio: urea clearance/creatinine clearance Urine volume (ml./min)
0-47 ± 0-03
0-37 ± 0-03
0-50 ± 0-03
0-17 + 0-03
0-14+ 0-02
0-21 ± 0-01 4-2
98 W. J. O'CONNOR AND R. A. SUMMERILL changes in plasma urea (compare Table 1 and 2). Following urea there was no significant change in urea or creatinine clearance (compare control experiments in Table 1), and no significant increase in the ratio urea clearance/creatinine clearance. In the urea as in the meat experiments the urine volume remained low.
Excretion of electrolytes Table 3 lists the excretion of electrolytes 3 5-4 hr after meat, at the corresponding times in control experiments and in the experiments where urea was given. After meat there were increases; on complete metabolism 170 g meat would yield about 10 m-mole PO0, 12 m-mole K, 6 m-mole Na, 3 m-mole Cl and about one-third of each of these was excreted during the TABLE 3. Electrolyte excretion at 3j-4 hr after meat, after two does of urea and in control experiments: mean, S.E. of mean and n the number of experiments on three on three dogs listed in Tables 1 and 2 Meat Urea Control n= 28 n= 13 n= 22 9.1 + 1.9 6-0 + 0 9 Na excretion (#ctmole/min) 18-5 + 2-2 6 1 + 0.7 K excretion (#mole/min) 28-8 + 1-8 8-4 ± 1.5 4-4 + 0-6 12-1 + 1 7 5.5 + 1-2 Cl excretion (,umole/min) 5.9 + 1.0 3.6 + 0 6 17.6 ± 1-4 PO4 excretion (,umole/min)
6 hr after the meat. After urea, apparent increases in the rate of excretion of electrolytes are of doubtful significance and were certainly small in comparison with the increases after meat.
Production of urea after a meat meal Urea produced in the body is either excreted in the urine or retained dissolved in the total body water. The amount excreted was directly measured; e.g. in the average of Fig. 1A the amount excreted in the first 2 hr after the meal of meat was 19 m-mole. At this time plasma urea had increased from 6-12 to 8-28, i.e. by 2-16 m-mole/l. If it be assumed that urea had equilibrated throughout total body water of 111., then 24 m-mole urea had been retained and the total urea production in the 2 hr after the meal was 43 m-mole. By 4 hr after the meal 48 m-mole had been excreted and plasma urea was increased by 3-95 m-mole/l. indicating a retention of 43 m-mole and a total production in the 4 hr of 91 m-mole. This calculation has been applied to all stages of the experiments of Fig. 1 A to give Fig. 4 where the estimated total urea production is plotted against time. In control experiments with Lassie the production in 8 hr was 23 m-mole. In Fig. 4 in the first 4 hr after the meat the average rate of production was 380 ,smole/min and in control experiments 54 ,umole/min.
UREA EXCRETION AFTER MEAT 99 The rate of urea production decreased after 6 hr and in the period 6-8 hr after the meat only 13 m-mole was produced, 110 /zmole/min, still a little more than in control experiments. With other dogs similar rates of urea production were estimated, the analysis for Cleo and Cathy being included in Fig. 4. In seven dogs the estimated urea production during the first 4 hr after meat (10 g/kg) was 56-150 m-mole, equal to 230-600 iumole/min. 150 I,
0
E C
100
0
e
U
0
0~~~~~~~~~0
4,~~~~0 A
0
0
0
2
4 6 8 Time after meat (hr) Fig. 4. The estimated production of urea after meat, and in control experiments. Abscissa, time after meat: ordinate, accumulated production of urea, in-mole. * and 0 are meat and control experiments in Lassie; A and A are meat and control experiments in Cathy or Cleo, the results being indistinguishable. Mean and s.E. of mean is shown by the bars or by the width of the symbols in control experiments.
After 175 g meat given to Lassie the estimated urea production in 6 hr was 132 m-mole (Fig. 4) and in control experiments 21 m-mole. The total N2 (urea equivalence) of the protein in the meat was 245 m-mole so that the increased urea production in the 6 hr represented 45 % of the total N2 in the meat. In the case of Cleo and Cathy increased urea production in 6 hr after meat amounted to 30 % of its N2. The calculation of urea production depends on the assumption that at the times of blood sampling urea was evenly distributed through the body water and the total body water of Lassie, weight 17-18 kg, was correctly assumed to be 111. (Gaudino & Levitt, 1949; Edelman, 1952). That the method is essentially correct was substantiated when the same calculations were applied to the experiments where known amounts of urea were given by stomach tube (e.g. Fig. 2); the estimated urea 'production' 2 hr after ingestion of doses of urea was 102 % + 2 (s.E. of mean, n = 20), 102 % ± 3 (11), 91 % ± 5 (12) of the administered amount in Lassie, Cathy and Cleo
respectively.
100
W. J. O'CONNOR AND R. A. SUMMERILL DISCUSSION
From the analysis in Fig. 4 it seems that following ingestion of meat the sequence of digestion, absorption of amino acids, deamination and formation of urea reached a rate of urea formation of 230-600 #tmole/min. Somewhere along the sequence: absorption of amino acids; deamination; urea formation; unidentified chemical substances released into the circulation (or other mechanisms) have the effect of increasing glomerular filtration rate (for discussion see O'Connor & Summerill, 1976). Increased glomerular filtration rate itself causes increased urea excretion, with increased urea clearance. However, in our experiments the filtration rate did not increase sufficiently to cause urea excretion to match the increased urea production; urea therefore accumulated in the body with rising plasma urea further increasing urea excretion until production was matched. The ultimate mechanism by which the rate of production of urea was equalled by excretion was increasing plasma urea; the necessary increase in plasma urea was however less by the fact that glomerular filtration rate increased at the time of protein catabolism. Before the work of this and the preceding paper only very large meat meals have been shown to increase glomerular filtration rate. Now that a substantial increase has been demonstrated with moderate sized meat meals, it seems that in the dog, increased glomerular filtration after a meat meal may be of importance in normal function. However, the dog is unique amongst species commonly used for investigation of renal physiology in that it is usually maintained on a single daily meal of high protein content. On a weight basis the similar routine for man would be a single daily meal including 650 g lean beef. Even if the protein intake of man were of this order (140 g/day) it would be spread over three meals a day. There seems to have been no deliberate investigation in man of the possibility that the meals may be an important cause of the higher clearances during the day as compared to night (Wesson, 1964) or that the changes in clearances with changes in dietary protein may contribute differences in renal excretion of urea and electrolytes. Schmidt-Nielson (1958) has reviewed evidence relating glomerular filtration rate and urea excretion in several species. The experiments in which urea was given add to an extensive literature on urea excretion, reviewed by Schmidt-Nielsen (1958). Our experiments differ from almost all of the published work in that they were carried out at the normal rates of urine flow of dogs, i.e. 0-1-0-3 ml./min as defined by O'Connor & Potts (1969) and the doses of urea have been small enough for urea excretion to remain within the normal range and this leads to some differences between our results and the usual findings.
101 UREA EXCRETION AFTER MEAT With large doses of urea, Mudge, Foulkes & Gilman (1949) and Anslow & Wesson (1955) produced osmotic diuresis and increased excretion of electrolytes whereas with our small doses any such changes were small. Urine volume after urea remained below 0-25 ml./min (Table 2) and there was no significant change in electrolyte excretion in comparison with control experiments (Table 3). The only effect with small doses of urea was that urea excretion increased. With regard to urea excretion our results with small doses of urea emphasize that the rate of excretion of urea is predominantly determined by plasma urea concentration (Fig. 3) and by glomerular filtration rate when this changes after meat. Since the urine volume remained low, it does not form a major determinant of urea excretion as has been suggested from the earlier literature. The only investigation of urea excretion at normal low urine flows is contained in the paper of Shannon (1936). At urine flows of about 0-15 ml/ min in his experiments, urea clearance/creatinine clearance was 0 3 and increased progressively to 0-6 as the urine volume was increased by the administration of water. The interpretation of Shannon (1936) and also Mudge, Berndt & Valtin (1973) is that at higher urine flows there is less back diffusion of urea; this must be an over-simplification in view of the supposed roles of different parts of the nephron indicated by the micropuncture work reviewed by Mudge et at. (1973). In our experiments, with the urine volume 0 1-0 3 ml./min both in control experiments and after urea, urea clearance/creatinine clearance was 0 35-0-55 which is higher than in Shannon's (1936) results at similar low urine flow. We have recorded the effects of water on only a few occasions and when urine volume was thus raised above 1 ml./min the ratio increased to 0-6 as in Shannon's experiments. After meat with the urine flow only 0-25 ml./min, the ratio rose to 0-5-0-6 (Table 1), whereas at similar urine flow, Shannon (1936) observed ratios of 0'3-0 4. The increased ratio after meat does not seem to be due solely to the small increase in urine volume; it could be associated with the increased excretion of electrolytes. It seems that decreased re-absorption of urea is a secondary effect of the increased glomerular filtration, which adds to urea excretion after meat, as compared to after doses of urea. This aspect of urea excretion has been particularly considered by Schmidt-Nielsen (1958). The authors are greatly indebted to Mr J. Brook and Mrs S. Snack for their technical assistance.
102
W. J. O'CONNOR AND R. A. SUMMERILL
REFERENCES ANSLOW, W. P. & WEssoN, L. G. (1955). Effect of sustained, graded urea diuresis on water and electrolyte excretion. Am. J. Phy8iol. 180, 605-611. EDELMAN, I. S. (1952). Exchange of water between blood and tissues. Characteristics of deuterium oxide equilibration in body water. Am. J. Phy8iol. 171, 279-296. GAUDrNO, M. & LEVITT, M. F. (1949). Inulin space as a measure of extracellular fluid. Am. J. Phy8iol. 157, 387-393. GOLOB, P. (1974). The effect on water, electrolyte and nitrogen balance of overfeeding dogs. Ph.D. Thesis, Leeds University. KIMBALL, A. W. (1952). Answer to query. Biometric8 8, 85-86. MUDGE, G. H., BERNDT, W. 0. & VALTIN, H. (1973). Tubular transport of urea, glucose, phosphate, uric acid, sulphate and thiosulphate. In Handbook of Phy8iology, section 8: Renal Physiology, chap. 19, pp. 587-652. Washington: American Physiological Society. MUDGE, G. H., FOULEms, J. & GILMAN, A. (1949). Effect of urea diuresis on renal excretion of electrolytes. Am. J. Phy8iol. 158, 218-230. O'CONNOR, W. J. & PoTTs, D. J. (1969). The external water exchanges of normal
laboratory dogs. Qt JZ exp. Phy8iol. 54, 244-265. O'CONNOR, W. J. & SUMMERILL, R. A. (1976). The effect of a meal of meat on glomerular filtration rate in dogs at normal urine flows. J. Phy8iol. 256, 81-91., SHANNON, J. A. (1936). Glomerular filtration and urea excretion in relation to urine flow in the dog. Am. J. Phyaiol. 117, 206-225. SCHMIDT-NIELSEN, B. (1958). Urea excretion in mammals. Phy8iol. Rev. 38, 139-169. SUMMERILL, R. A. (1974). The effects of a meat meal on glomerular filtration rate and urea excretion in the conscious dog. J. Physiol. 238, 64P. TECHNICON (1968). Auto-analy8er Handbook. Houndmills, Basingstoke, Hants: Technicon Instruments. WESSON, L. G. (1964). Electrolyte excretion in relation to diurnal cycles of renal function. Medicine, Baltimore 43, 547-592.
