drinking through release of renin. Since most of the known physiological

Transcription

1 J. Physiol. (1969), 23, pp With 5 text-figure8 Printed in Great Britain THE EFFECT ON DRINKING IN THE RAT OF INTRAVENOUS INFUSION OF ANGIOTENSIN, GIVEN ALONE OR IN COMBINATION WITH OTHER STIMULI OF THIRST By J. T. FITZSIMONS AND BARBARA J. SIMONS* From the Physiological Laboratory, University of Cambridge (Received 9 December 1968) SUMMARY 1. Intravenous infusion of angiotensin causes rats which are in water balance to drink water. 2. The mean amount of angiotensin needed to initiate drinking was jtg/kg (S.E. of mean) in twenty normal rats, and ,g/kg in thirty-four nephrectomized rats. 3. The nephrectomized rat is therefore more sensitive to this action of angiotensin than the rat with intact kidneys. 4. The rates of infusion (.5-3. jtg/kg-l min-) which cause drinking are comparable to those used to produce other effects in rats. 5. Angiotensin restores the drinking response of the nephrectomized rat subjected to caval ligation to a value similar to that obtained in the uninfused normal rat subjected to caval ligation. 6. The effects of angiotensin and hypertonic saline on drinking are additive when both substances are administered to nephrectomized rats. 7. These experiments provide further support for the view that the renin-angiotensin system is concerned in extracellular thirst. INTRODUCTION Certain procedures which interfere with the circulation of the blood such as ligation of the abdominal inferior vena cava or constriction of the abdominal aorta above the renal arteries cause a rat in normal fluid balance to drink water (Fitzsimons, 1969b). The facts that this effect is much reduced by previous nephrectomy and that there is a potent dipsogen in the renal cortex, probably renin, may mean that these procedures cause drinking through release of renin. Since most of the known physiological effects of renin are mediated through angiotensin (Peart, 1965), the action * Thouron Scholar, University of Pennsylvania.

2 46 J. T. FITZSIMONS AND BARBARA J. SIMONS of angiotensin on drinking was tested. Angiotensin was found to cause drinking in the rat and this was reported to the Physiological Society (Fitzsimons & Simons, 1968). This effect of angiotensin is described in detail here together with further experiments in which the role of angiotensin in combination with other stimuli of drinking is considered. METHODS Care of animals. Male albino rats weighing g had food and water available ad libitum until the start of the experiment at between 9 a.m. and 12 noon. During the experiment which lasted 6 hr, the rats were housed in individual metabolism cages where they had no food but had access to water from a drinking meter which enabled the time course of drinking to be recorded (Fitzsimons, 1958). Infusion of angiotensin into normal and nephrectomized rats. Each animal was anaesthetized with ether and the external jugular vein was catheterized with a soft polyvinyl chloride tube, outside diameter 1 mm. The catheter was filled with NaCl solution (.9 %, w/v) and its distal end was closed with a small plug. In some rats bilateral nephrectomy was carried out through a mid line dorsal incision at the same operation as the jugular catheterization. After the operation the rats were weighed to the nearest -1 g and placed in the individual metabolism cages. Before the rats had fully recovered from the anaesthesia the jugular catheter was connected to a 15 cm length of stainless-steel tubing, outside diameter -75 mm, which projected through the top of the cage. This prevented the rat from chewing through the tubing and eliminated the necessity of restraining the rats during the experiment. This metal tube was connected by 1-1J m of polyvinyl chloride tubing to an infusion pump. The technique of intravenous infusion into unrestrained rats has been described previously (Fitzsimons, 1963). When the rats had recovered from the anaesthetic, angiotensin (val5 angiotensin II amide, Hypertensin CIBA) dissolved in -9 % NaCl was infused at rates of ug kg-' min-, for periods of 3-31 min. The concentration of angiotensin varied between -75 and 2 jg/ml.; the rate at which solution was infused was yl./1 g body wt. min-; and the final volume of the infusate lay between -23 and 3-94 ml./1 g body wt. Control rats were infused with a similar range of volumes of -9 % NaCl. Drinking was recorded continuously throughout the experimental period of 6 hr. Urine was collected in graduated centrifuge tubes and was tested for protein by a standard biuret procedure, the urinary bladder being emptied by suprapubic pressure at the start and finish of the experimental period. At the end of the experimental period all rats were again weighed to the nearest - 1 g. Autopsy was routinely performed on all animals in order to determine whether or not fluid had accumulated in the pleural or peritoneal cavities. Measurement of haematocrit and plasma volume. Similar infusion experiments were carried out on nine rats with intact kidneys and nine nephrectomized rats, with the object of ascertaining the effect of angiotensin on the blood volume. The rats were unrestrained and conscious but they were not allowed to drink. An intravenous injection of -3 ml. -8 % Evans blue in -9 % NaCl was followed by 3 ml. -9% NaCl. Twenty minutes later -5 ml. blood was collected from the tail and the haematocrit and Evans blue space were measured. Immediately after this, infusion of angiotensin or -9 % saline was started. The rates of infusion of angiotensin and the doses given were comparable to the higher ones used in the behavioural part of the experiment. A second sample of -5 ml. blood was taken for haematocrit and Evans blue determination at the end of the infusion. In some instances a third sample of blood was taken an hour or so after cessation of the infusion. Angiotensin in combination with other stimuli of drinking. In some nephrectomized rats

3 INTRA VENOUS ANGIOTENSIN AND THIRST the additivity of the drinking effect of angiotensin with either a cellular thirst stimulus, hypertonic saline, or an extracellular stimulus, caval ligation, was tested as follows. At the time of the operations described above some animals were given a single injection of 2 M-NaCl intravenously in amounts sufficient to increase the osmotic pressure of the body fluids by about 5 or 1 %. It was assumed that the initial osmolality of the body fluid is 29 ju-osmole/g H2 and that body water is 69 g/1 g initial body wt. (Fitzsimons, 1961 a). In another group of rats, immediately after the removal of the kidneys, the inferior vena cava was completely ligated above the entry of the renal veins and below the hepatic veins. In these experiments drinking water was withheld until the effects of the anaesthetic had worn off completely. Then approximately 45 min after the operation infusion of angiotensin was begun and drinking water was made freely available. The rest of the experimental procedure for these two groups of rats was the same as that described for animals given angiotensin alone. RESULTS Drinking by normal and nephrectomized rats during the infusion of angiotensin The intravenous infusion of angiotensin causes rats which are in water balance at the start of the experiment to drink considerable quantities of water, the effect being greater in nephrectomized rats than in rats with intact kidneys. In Fig. 1 are shown typical records of drinking by nephrectomized rats during the infusion of angiotensin or -9 % saline. Once started, angiotensin-induced drinking usually continued throughout the infusion and stopped as soon as the infusion ended. There was, however, individual variation in the drinking patterns. For instance, some rats drank in bursts of several ml. at a time with pauses of 5-1 min between bursts. Also, in a few instances when the infusion lasted a long time, or when the amount of water drunk was large, the rate of drinking decreased towards the end of the infusion. Similar variations in drinking pattern have been seen in animals infused with hypertonic saline. In both normal and nephrectomized rats drinking appeared to depend more on the total amount of angiotensin infused than on any other of the parameters of infusion. The amount of water drunk by rats with intact kidneys was positively correlated with the dose of angiotenmin administered (r = *58, degrees of freedom (d.f.) = 2, P <.1); the formula of the regression line, excluding the controls, being y = x, where y is water drunk in ml./1 g body wt. and x is angiotensin infused in,ug/kg body wt. The same holds in nephrectomized rats (r = 7, d.f. = 33, P < -1; y = x; Fig. 2). When the controls are also taken into account the correlation coefficient for the nephrectomized group is 75 (d.f. 42, P < -1) and the regression formula is y = x. There was no significant relation between the rate of infusion of angiotensin and the amount of water drunk (Fig. 2); at rates of less than 25,ug kg-'min-' the amount drunk after a given dose of angiotensin tended 47

4 48 J. T. FITZSIMONS AND BARBARA J. SIMONS to be slightly less than after the same dose administered at higher rates, though not significantly so. Nor was there any statistically significant effect on drinking attributable either to the rate of infusion of fluid or to the total volume of fluid infused. Similar amounts of angiotensin contained in either a small or in a large volume of fluid caused drinking of similar amounts of water. 162 pg/kg body wt. 68#g kg-' min-' 24mm 455,ug/kg body wt. LT I1 ml. 1-52,ug kg-' min- ' -* 3 min Control t.9% NaCl 24 mm Fig. 1. Typical records of drinking during intravenous infusion of angiotensin into body wt., nephrectomized rats. The total amount of angiotensin infused in /kg/kg and the rate of infusion in Aug kg-' min- are given on the left of the Fig. The duration of the infusion is represented by the double ended arrow. The interval between the first and last mark on each record is 6 hr. The vertical calibration on the right of the Fig. refers to all records. The total amount of water drunk by nephrectomized rats in response to a given dose of angiotensin was generally considerably greater than the intake of normal rats after the same dose (Table 1). After the larger doses, the amounts drunk by nephrectomized rats were sometimes more than half the average daily intake of a normal rat which had been allowed food as well as water ad libitum throughout the 24 hr. The rat drinks at night, and normally takes very little fluid during the daytime, when the infusions were made. It is also apparent from Table 1 that the larger the dose of angiotensin administered to nephrectomized rats the greater the final net positive fluid balance and therefore the more hypotonic the body fluids had become. The positive fluid balance and hypotonicity did not however appear to impair drinking in response to angiotensin. In the normal animal angiotensin caused a brisk diuresis, and since urine flow was greater than the rate of drinking the rat went into negative fluid balance. The loss of

5 INTRAVENOUS ANGIOTENSIN AND THIRST 49 weight in the experimental period was roughly the same for all groups with intact kidneys, including the controls, so that urine flow correlated with the sum of the volumes of the infusate and the water drunk. Despite the lack of fluid retention, with its presumed inhibitory effect on drinking, normal rats drank less than nephrectomized rats after the same dose of angiotensin. 6 r -_ 4 bo -3 (b) >1Op-g kg-' min-'. y=*82+2lx fc) All rates. y=67+21x '(a) <1-ug kg-' min-'. y= x d 2 9 o 11,,,,,,,,1, Angiotensin infused (jug/kg body wt.) Fig. 2. The regressions of water drunk by nephrectomized rats in 6 hr on total amounts of angiotensin infused at (a) <. jug kg-' min-', (b) > 1- /.jg kg-' min-', and (c) all rates combined. The left-hand and right-hand limits of the regressions indicate the range of total doses of angiotensin infused. The mean value + S.D. for the control rats is given on the left with the number of observations in parentheses. The control values were not used in calculating the regressions. The two regressions (a) and (b) do not differ in slope (t = -23, d.f. = 31) and are not displaced from each other (t = -99, d.f. = 32). Angiotensin infused (/zg/kg initial body wt.) (controls) < TABLE 1. Drinking by normal and by nephrectomized rats infused with angiotensin. All experiments lasted 6 hr Rats with intact kidneys Nephrectomized rats A- Change in Urine Change in Water drunk body wt. volume Water drunk body wt. (ml./1 g (g/1 g (ml./1 g (ml./1 g (g/1 g initial body initial initial initial initial wt.) body wt.) body wt.) body wt.) body wt.) (6) (9) (2) (8) (4) (9) (5) (7) (5) (5) (3) (6) (Mean value + S.E. of mean with number of observations in parentheses)

6 5 J. T. FITZSIMONS AND BARBARA J. SIMONS When nephrectomized rats were infused with angiotensin at very low rates, where perforce the doses administered were small, the total intake of water by the experimental animals was very little greater than that of the controls, but the rats receiving these infusions started drinking somewhat earlier in the 6 hr period than did those receiving only a saline infusion. Taking all animals into consideration, the mean delay ± s.e. of the mean before the onset of drinking of thirty-four out of thirty-five nephrectomized rats (i.e. excluding one animal which did not drink) infused with angiotensin was min, contrasted with min for the six out of nine nephrectomized controls (i.e. excluding three animals which did not drink). When the kidneys were left in situ, it was easy to distinguish experimental from control animals, since none of the controls drank during the experimental period of 36 min, whereas twenty out of twenty-two animals infused with angiotensin drank, with a mean delay of min. There is therefore no doubt that angiotensin caused the rats to start drinking. As with total intake of water, there was no consistent relationship between the rate of infusion of angiotensin and the onset of drinking (Fig. 3), nor did the volume of fluid infused affect the onset of drinking. Despite large individual differences it is simplest therefore to take the total amount of angiotensin infused at the onset of drinking as a measure of the minimum amount of angiotensin needed to produce an effect on drinking in animals in water balance. Taking all experimental animals into consideration, the amount of angiotensin + S.E. of the mean infused at the start of drinking was *1 jtg/kg in thirty-four nephrectomized rats, and ug/kg in twenty normal rats. Initiation of drinking is therefore a more sensitive index of the dipsogenic effect of angiotensin than is the total amount of water drunk, and it confirms the enhanced responsiveness of nephrectomized rats to this substance as compared with normal animals. In view of the known effects of angiotensin on capillary permeability the urines of the normal animals were tested for protein using the standard biuret test, a careful search was made for free fluid in the pleural and peritoneal cavities at autopsy, and some measurements of haematocrit and Evans blue space were made. With doses of angiotensin greater than 8,ug/kg the urine biuret test became positive. Free fluid was found when the largest doses of angiotensin were combined with injections of hypertonic saline into nephrectomized rats, and very occasionally when high doses of angiotensin were given alone to nephrectomized rats; free fluid was never found in animals with intact kidneys. Neither the haematocrit, nor the Evans blue space showed any significant effect attributable to angiotensin, though the rate of increase of haematocrit and Evans blue

7 INTRAVENOUS ANGIOTENSIN AND THIRST 51 space after the largest doses of angiotensin was sometimes higher than in the control animals. The dipsogenic effect of angiotensin was manifest with doses of angiotensin far below those which occasionally produced marked changes in blood volume. 1 9 >, Fig. 3.! 6 5 ~~~~~~~ -:~4 43 Normal mean 3 * ~~~. ~~~~~~.. 2 Nephrectomized mean * * 1.o.s.. I f111,li is II I Rate of infusion of angiotensin (jtg kg-' min-') The amounts of angiotensin needed to initiate drinking at different rates of infusion in normal () and nephrectomized () rats. The effect of angiotensin on drinking does not appear to be mediated through aldosterone since adrenalectomized nephrectomized rats drank similar amounts of water to the amounts taken by nephrectomized rats in response to a given dose of angiotensin. In both groups there was a significant (P <.5) positive correlation between the amount of water drunk and the dose of angiotensin. The equation of the regression line of the nephrectomized group of forty-four rats including nine controls was y = *79 + *2x, and that of the adrenalectomized nephrectomized group of twenty-one rats including four controls, y = x where y is

8 52 J. T. FITZSIMONS AND BARBARA J. SIMONS the amount of water drunk in ml./1 g body wt. in 6 hr and x the dose of angiotensin in,tg/kg body wt. The slopes of the two regression lines were not significantly different (t = 15, d.f. = 61) nor were the regressions displaced significantly one from the other (t = O81, d.f. = 62). Nor is the effect of angiotensin on drinking simply a consequence of its pressor action. Vasopressin (vasopressin injection B.P., Pitressin, Parke Davis & Co.) was infused at rates ranging from 1 to 4 u. kg-' min-' into twenty-three normal and twenty-eight nephrectomized rats under the same conditions as for angiotensin. These amounts of vasopressin cause similar rises in blood pressure to those caused by the doses of angiotensin used here; they did not, however, lead to any increase in intake of water (B. J. Simons, unpublished observations). The effect of angiotensin and caval ligation on drinking in the nephrectomized rat Caval ligation in intact rats may cause increased drinking partly through release of renin because drinking is much less when caval ligation is preceded by bilateral nephrectomy (Fitzsimons, 1966). In view of the results of the previous section of this paper it seemed likely that the renin effect was mediated through angiotensin. It was therefore of interest to determine whether angiotensin would restore the response of the nephrectomized rat subjected to caval ligation to a value similar to that of an animal with intact kidneys subjected to the same procedure. Angiotensin was therefore given by intravenous infusion to nephrectomized rats which had had the inferior vena cava ligated at the level of entry of the renal veins immediately after nephrectomy. Increasing doses augmented the rate of drinking in these animals and after the larger doses of angiotensin the response of the nephrectomized rat subjected to caval ligation was the same as that of normal animals subjected to caval ligation alone (Fig. 4). Increasing the dose of angiotensin still further, however, had only a small additional effect on water intake. It is worth noting that the normal animal subjected to caval ligation above the renal veins is almost completely anuric so that its net fluid balance is similar to that of the cavalry ligated nephrectomized groups which received the higher dose of angiotensin. The nephrectomized rat with caval ligation proved to be more sensitive to angiotensin than the rat subjected to nephrectomy alone. The effect of angiotensin and hypertonic saline on drinking in the nephrectomized rat Angiotensin was infused into nephrectomized rats which had been given sufficient 2 M-NaCl intravenously to increase the osmotic pressure of the

9 INTRA VENOUS ANGIOTENSIN AND THIRST 53 Water drunk (ml.11 g body wt.) Angiotensin infused 6 (,/g/kg body wt.) Kidneys present Nephrectomized L,,, I I I I A I I I Fig. 4. The effect of infusion of angiotensin on the amounts of water drunk by nephrectomized rats subjected to caval ligation '- 1_ la 9_ g 8 A,o Ir - n ~ + ++ (a) y=-79+-2x of + +s 4~~~~~~~4 I p1 I I I I I I I I G Angiotensin infused (,ug/kg body wt.) Fig. 5. Regressions of the amount of water drunk in 6 hr on the amount of angiotensin infused in nephrectomized rats after intravenous injection of hypertonic NaCl producing increases in osmotic pressure of (a), (b) 5 % and (c) 1 %. The three regression lines were parallel but significantly separated one from the other, thus supporting the hypothesis of simple additivity of thirst stimuli.

10 54 J. T. FITZSIMONS AND BARBARA J. SIMONS body fluids by about 5 or 1 %. The experiments were carried out on nephrectomized rats to avoid the complication of the renal response to the hypertonic solution which has been shown to affect the amount of water finally drunk (Fitzsimons, 1961 a), and also to eliminate endogenous renin secretion, and possible destruction of exogenous angiotensin by the kidney. An infusion of angiotensin combined with a single injection of hypertonic saline caused nephrectomized rats to drink more water and go into greater positive fluid balance than did injection of hypertonic saline alone (Fig. 5). The effects on drinking of the two stimuli were additive with doses of angiotensin of 4,ig/kg body wt. or greater. This is illustrated by the fact that the regressions of the amount of water drunk on the dose of angiotensin are parallel and separated (Fig. 5); t tests showed no significant difference in slope between any two lines (P > -5 on two tailed tests); the regression of the 5 % group, however, was significantly above that of the % group (t = 13'77, d.f. = 58, P < 1), and the regression of the 1 % group was significantly above that of the 5 % group (t = 3 82, d.f. = 31, P <.1). DISCUSSION Synthetic val5 angiotensin II amide causes rats in normal fluid balance to drink water, therefore making it likely that the dipsogenic effect of renin is mediated through generation of angiotensin. Nephrectomized rats drank more in response to a given dose of angiotensin, despite the retention of water and increasing hypotonicity of body fluids thereby resulting, than did rats with their kidneys present. Nephrectomized rats also show a greater pressor response to angiotensin, so the kidney is important in modifying the action of angiotensin. The enhanced sensitivity of nephrectomized animals to angiotensin may be due to the failure of excretion of angiotensin or to the absence of an angiotensin inhibitor (Page & Helmer, 194) or of angiotensinase which Regoli, Riniker & Brunner (1963) have found to be present in kidney homogenates. Akinkugbe, Brown & Cranston (1966) have also found evidence for renal inactivation of angiotensin in the rabbit. In both normal and nephrectomized rats the onset of drinking and the total amount of water finally drunk depend on the dose of angiotensin and do not appear to be directly determined by the rate of infusion. Because of the rapid and transient nature of the vascular and renal responses to angiotensin most workers have not found it necessary to make dose and rate independent variables and they have described the dose-response relationships in terms of rates of infusion only. Drinking, however, is such a leisurely process compared with other physiological effects of angiotensin

11 INTRAVENOUS ANGIOTENSIN AND THIRST 55 that it would be misleading to regard the rate of infusion as giving an indication of the degree of stimulation of thirst receptor sites; the rate of destruction and the time needed for drinking to be completed are also factors which affect the stimulus-response relationship. Despite the very different time relations of the vascular and renal responses to angiotensin on the one hand, and drinking on the other, the rates of infusion which cause drinking (5-3.,ug kg-'min-') are of the same order as those which cause some other effects in the rat: Malvin & Vander (1967), for example, found that infusion of angiotensin at rates between -8 and 1,ug kg-' min-' caused natriuresis together with a diminution in glomerular filtration rate; Bonjour, Regoli, Roch-Ramel & Peters (1968) used rates of -2-1-,tg kg-1 min- to study the natriuretic response; and Barraclough, Jones & Marsden (1967) used a rather wider range, 5-5 fig kg-1 min-, to study other aspects of renal function. Although the rates at which we administered angiotensin are comparable to those used by other workers, endogenous angiotensin formation probably does not often reach these rates. However, the demonstration that an infusion of angiotensin causes a rat in normal water balance to drink indicates that angiotensin is a potent dipsogenic substance, since animals in water balance do not normally drink, and an increase in endogenous angiotensin does not occur in isolation, i.e. without some precipitating disturbance within the animal, and these same disturbances have all been recognized as causes of thirst. Thus, in a number of species including the rat, haemorrhage, sodium depletion, obstruction of the inferior vena cava, constriction of the abdominal aorta above the renal arteries, and constriction of the renal artery have been shown to stimulate secretion of renin (Peart, 1965). These procedures also cause osmotically inappropriate drinking in the rat, and, most importantly, they have less effect on drinking after nephrectomy (Fitzsimons 1961b, 1969 a, b). It therefore was of particular interest to find that infusion of angiotensin restored the drinking response of the nephrectomized rat subjected to one of these procedures, ligation of the inferior vena cava, to the prenephrectomy value. Caval ligation leads to the release of renin, renin is a potent dipsogen and drinking is very considerably diminished when caval ligation is preceded by bilateral nephrectomy. The cavalry ligated nephrectomized preparation was more sensitive to the dipsogenic action of angiotensin than the rat subjected simply to nephrectomy. It seems reasonable then to suppose that exogenous angiotensin is simply replacing endogenous angiotensin in restoring the response to caval ligation to its prenephrectomy value. The heightened sensitivity to angiotensin in this experiment also suggests that physiological increases in angiotensin levels may have an

12 56 J. T. FITZSIMONS AND BARBARA J. SIMONS effect on water intake if there is an actual or subthreshold thirst stimulus present. The mechanism of angiotensin-induced drinking remains to be elucidated though there is now evidence that part of the effect may be exerted through a direct stimulating action of angiotensin on hypothalamic drinking centres since single intracranial doses as small as 1 ng cause rats in normal water balance to drink (Epstein, Fitzsimons & Simons, 1969). The fact that there is a residual drinking response in nephrectomized rats to extracellular thirst stimuli makes plausible the hypothesis that the function of angiotensin is to sensitize the central drinking centres to thirst stimuli arising from a variety of receptors elsewhere in the body. Another possibility is that the receptors themselves are directly affected by angiotensin. The third possibility is that angiotensin might stimulate drinking by causing a fall in plasma volume (a known stimulus of thirst in the rat, Fitzsimons, 1961 b), through its action in increasing vascular permeability. Angiotensin in moderate doses certainly causes proteinuria, and Geise (1963) has shown that infusions of 4-57 jug/rat given over a period of 2-24 hr cause pancreatic oedema and vascular damage in nephrectomized rats. However, he found only negligible amounts of free fluid in the peritoneal and pleural cavities. The damage caused by 3,g/rat given over a 5 hr period, 19 hr after nephrectomy, was much less; two out of five rats showed pancreatic oedema, and one had moderate to severe vascular lesions. These doses are considerably greater and the duration of the infusions generally much longer than in the present experiments. Geise did not measure plasma volume; in the present experiments angiotensin was found to cause a fall in plasma volume only after the largest doses were administered to nephrectomized rats. It therefore seems unlikely that the effect of 4ngiotensin on water intake reported here is attributable to a fall in plasma volume except perhaps after the largest doses. Infusions of angiotensin cause additional intake of water in rats injected with hypertonic saline, a cellular stimulus of thirst. Angiotensin therefore does not differ from a number of other extracellular stimuli of drinking which have been shown to add in simple algebraic fashion with cellular stimuli (Fitzsimons & Oatley, 1968; Fitzsimons, 1969a). The adrenal cortex does not appear to be involved in drinking caused by angiotensin. It has been suggested that hyperaldosteronism may, through the K depletion that it causes, account for the polydipsia which frequently occurs in Conn's syndrome, and there are also reports that aldosterone may elicit sodium appetite in rats (Wolf & Handal, 1966), though the doses used in these experiments were enormous. The possibility that angiotensin may exert its effect on drinking through aldosterone seems to be excluded, however, by the fact that infusion of angiotensin

13 INTRAVENOUS ANGIOTENSIN AND THIRST 57 into adrenalectomized nephrectomized rats had the same effect on drinking as infusions into nephrectomized rats. Furthermore, angiotensin seems to have little effect on aldosterone secretion in the rat (Eilers & Peterson, 1964); nor could an explanation along these lines account for the intracranial effect of angiotensin. REFERENCES NiKINKuGBE,. O., BROWN, W. C. B. & CRANSTON, W. I. (1966). Pressor effects of angiotensin infusions into different vascular beds in the rabbit. Clin. Sci. 3, BARRACLOUGH, M. A., JONES, N. F. & MARSDEN, C. D. (1967). Effect of angiotensin on renal function in the rat. Am. J. Phygiol. 212, BoNjouR, J. P., REGOLI, D., ROcH-RAMEL, F. & PETERS, G. (1968). Prerequisites for the natriuretic effect of val5-angiotensin II amide in the rat. Am. J. Phy8iol. 214, EILERS, E. A. & PETERSON, R. E. (1964). Aldosterone secretion in the rat. In Aldo8terone, a 8ympo8'um, ed. BAULIEU, E. E. & ROBEL, P., pp Oxford: Blackwell. EPSTEIN, A. N., FITZSIMONS, J. T. & SImoNs, BARBARA J. (1969). Drinking caused by the intracranial injection of angiotensin into the rat. J. Phy8iol. 2, 9-l1P. FITZSIMONS, J. T. (1958). Apparatus for recording drinking and feeding. J. Phy8iol. 143, 31-32P. FITZSIMONS, J. T. (1961 a). Drinking by nephrectomized rats injected with various substances. J. Phyliol. 155, FITZSIMONS, J. T. (1961 b). Drinking by rats depleted of body fluid without increase in osmotic pressure. J. Physiol. 159, FITZSIMONS, J. T. (1963). The effects of slow infusions of hypertonic solutions on drinking and drinking thresholds in rats. J. Phy8iol. 167, FITZSIMONS, J. T. (1966). Hypovolaemic drinking and renin. J. Physiol. 186, P. FITZSIMONS, J. T. (1969a). The effect of nephrectomy on additivity of drinking stimuli. J. comp. physiol. P8ychol. (In the Press.) FITZSIMONS, J. T. (1969 b). The role of a renal thirst factor in drinking induced by extracellular stimuli. J. Phy8iol. 21, FITZSIMONS, J. T. & OATLEY, K. (1968). Additivity of stimuli for drinking in rats. J. comp. physiol. P8ychol. 66, FITZSIMONS, J. T. & SIMONS, BARBARA J. (1968). The effect of angiotensin on drinking in the rat. J. Phy8iol. 196, P. GEISE, J. (1963). Pathogenesis of vascular disease caused by acute renal ischaemia. Acta path. microbial. 8cand. 59, MALVIN, R. L. & VANDER, A. J. (1967). Effects of angiotensin infusion on renal function in the unanesthetized rat. Am. J. Phy8iol. 213, PAGE, I. H. & HELMER,. M. (194). Angiotonin activator, renin and angiotonin inhibitor and the mechanism of angiotonin tachyphylaxis in normal, hypertensive and nephrectomized animals. J. exp. Med. 71, PEART, W. S. (1965). The renin-angiotensin system. Pharmac. Rev. 17, REGOLI, D., RINIKER, B. & BRUNNER, H. (1963). The enzymatic degradation of various angiotensin II derivatives by serum, plasma, or kidney homogenate. Biochem. Pharmac. 12, WOLF, G. & HANDAL, P. J. (1966). Aldosterone-induced sodium appetite: dose-response and specificity. Endocrinology 78,