J. Physiol. (I958) I40, 23-36

Transcription

1 23 J. Physiol. (I958) I40, EFFECTS OF ADRENALECTOMY ON OXYGEN POISONING IN THE RAT BY D. W. TAYLOR From the Department of Physiology, University of Aberdeen (Received 1 July 1957) Until recent years little work seems to have been done to find out whether or not the adrenal glands are involved in the disturbances arising from exposure to oxygen under pressure. This seems all the more remarkable in view of the volume of work on 'stress' and the importance of the adrenal glands in determining the reactions of the body to it. Campbell (1937) increased the severity of toxic manifestations in rats by administering adrenaline. Bean (1951) discovered that repeated exposure of rats to oxygen under high pressure resulted in hypertrophy of the adrenal cortex. Brief reports of the earlier experiments in this paper have been given to the International Physiological Congress in Montreal, and to the Physiological Society (Taylor, 1953, 1954). While the work described below was in progress, both Bean and Gerschman and their colleagues published a considerable number of interesting papers on the subject; these are dealt with in the discussion. METHODS Animals and diet. Hooded rats of the Rowett strain, from a colony kept in this laboratory and closed for some forty generations, were used in all experiments but two. In these two the reactions of this strain were compared with those of the eighth generation in descent from a wild black x albino mating. All animals were fed throughout on the stock diet which differs from that of Thomson (1936) only in that it contains 14%, not 7%, of skim milk. The animals were aged 3-5 months, and in any given experiment (apart from Expts. 1 and 7) they were of the same age. In Expt. 1 there was a considerable variation in age and Expt. 7 was designed to test the effect of age. In the light of later experiments the age variation may have had a slight influence on the results in Expt. 1, but insufficient to affect the general conclusions. Operation and post-operative maintenance. Bilateral adrenalectomy was carried out, under ether anaesthesia, through a central dorsal skin incision. From the day of operation the rats were given 0.9% NaCl solution to drink, except in Expt. 8. In other respects their maintenance did not differ from that of unoperated control animals. Exposure to oxygen under pressure. The details of exposure have been described (Taylor, 1956). Briefly, the animals were placed in the steel chamber which was then flushed with some Pressure was then raised at a uniform rate to 90 Lb./sq. in. (6 atm approx.). CO2 was absorbed by trays of soda-lime. Room temperature was kept, as far as possible, at 200 C.

2 24 D. W. TAYLOR In some experiments adrenalectomized animals were exposed twice. They were exposed first for a fixed period of time (50 min) at the end of which all those showing any evidence of O2 poisoning were rejected. The others were then divided into treatment groups and re-exposed at a later date. Drug8. Thyroid powder (Armour Laboratories) was administered daily by mouth (10 mg/100 g body wt./day) as a suspension in water. Deoxycortone acetate B.P. (DOCA) and cortisone acetate (Merck and Co.) were each injected intramuscularly twice daily in doses of 0 5 mg/100 g body wt. Hydrocortisone acetate (Roussel Laboratories) was similarly injected twice daily in doses of 0 05 or 0 5 mg/100 g body wt. Cortin (Eucortone, Allen and Hanbury), which is the whole cortical extract, was administered subcutaneously in two daily doses of 0 5 ml./100 g body wt. Adrenaline (0.01 mg/100 g body wt). was given subcutaneously as the hydrochloride in a single dose 15 min before exposure. Duration of treatment, and also the period of time between operation and exposure, varied somewhat from experiment to experiment. AsWe88ment of reaults. The method of assessing general effects has been described already (Taylor, 1956). Severe poisoning is usually indicated by convulsions, but sometimes prostration occurs without motor disturbance. This prostration is often, but not always, accompanied by hyperpnoea. The interval from the initial raising of the 02 pressure to the appearance of such prostration or convulsions, whichever was earlier, was taken as the 'time to the onset of severe poisoning'. In some operated animals neither convulsions nor prostration occurred after exposures varying from 70 to 120 min, the experiments being stopped in most cases when respiratory distress became apparent, and in a very few instances, merely on account of the passage of time. In all these cases, to permit of statistical assessment of the results, the time to the beginning of decompression was taken as the 'time to onset of severe poisoning'. Post-mortem examination was carried out on all animals. Lung damage was assessed on an arbitrarv scale of 0-5. In Expt. 2, which involved much the largest number of rats, a careful histological examination was made of tissues in the field of operation, in an attempt to discover either aberrant cortical tissue or evidence of incomplete removal of a gland. Specimens were preserved in 'Susa' (Heidenhain), blocked in ester-wax (British Drug Houses) and stained witb haematoxylin-eosin (Carleton & Leach, 1947). RESULTS Experiment 1. Two groups of sixteen rats, three male and thirteen female in each, were used. One group was adrenalectomized and the other retained as a control. Two of the operated animals died before exposure, which took place on the fourteenth day after operation. All the sixteen controls had severe convulsions between 26 and 44 min after exposure began; the mean time was min (S.E. of mean). Six of the fourteen operated animals became convulsed, and for the remainder exposure ended with the onset of respiratory distress. The duration of exposure ranged from 45 to 90 min, except for one rat which had an anomalous convulsion at 16 min; this last value was omitted, the mean time for the others being 68x5 + 3x9 min. The time was more variable for the operated group, but the evidence for a longer tolerance by operated animals was overwhelming. Lung damage in the two groups could not be compared because the control rats were saved for further tests. Eighteen days later, fourteen of the erstwhile control rats were divided into two groups of seven, of which one group was adrenalectomized. After 14 days

3 ADRENALECTOMY AND OXYGEN POISONING 25 (i.e. 4-5 weeks after their first exposure) all were exposed again and the time to onset of severe poisoning recorded. Table 1 shows the mean times for first and second exposures. The unoperated animals had essentially the same time on both occasions, but adrenalectomy increased the time for the other group very substantially, bringing it to about the same level as for the other animals, operated on in the first part of the experiment. Indices of lung damage were obtained for the fourteen animals; the difference between the means of for the unoperated and for the operated was statistically significant. TABLE 1. Expt. 1. Mean times (min) to the onset of severe poisoning in rats exposed to oxygen at a pressure of 6 atm. The rats in Part 2 comprise fourteen of the controls used in Part 1 Part 1 Control Adrenalectomized Difference 36* ±4-2* Part 2 Control Adrenalectomized throughout after 1st test Difference 1st test ±2O0* 2nd test ±5t5 * S.E. of mean. Experiment 2. In this experiment adrenalectomized and control rats were compared in respect of the effects of different drugs; animals received 'no drug', cortisone, DOCA or thyroid, and were independently assigned to 'adrenaline' or 'no adrenaline' groups. Equal numbers of male and female rats were used in a balanced manner. On account of the evidence for greater variation in sensitivity amongst adrenalectomized rats shown in Expt. 1, twice as many rats were allotted to the operated group as to the unoperated. In all, 144 rats were used in a 4 x 23 factorial scheme, having threefold replication of unoperated and sixfold replication of operated animals. In order to eliminate any trends with time, 'blocks' of twelve rats were used on each of twelve occasions in accordance with a design confounding each complete replicate of the 4 x 23 scheme into quarters and losing information on three unimportant interactions. Each block of twelve contained four unoperated animals, two of each sex, two with and two without adrenaline, and one on each of the other four drug treatments; it also contained eight operated animals, twice as many from each category. Exposure to oxygen took place 2 weeks after adrenalectomy, treatments being started on the eighth post-operative day and continuing to include the day of exposure. Ages of all animals on their day of exposure were comparable, and the whole experiment extended over 12 weeks. Seven rats died, and appropriate adjustments to the statistical analyses were used to take account of this. A few of the times to onset of poisoning

4 26 D. W. TAYLOR were recorded simply as lower limits (75 +, 100 +, etc.), and these were all arbitrarily taken as 100. The mean times to onset of severe poisoning are given in Table 2. Despite the precautions taken in designing the experiment, the variability in the response to high tension of oxygen was as high as in Expt. 1; there was no correlation between susceptibility of rats and their changes in weight. The operated animals again had significantly longer times to onset of severe poisoning, though the difference was smaller than in Expt. 1. The average time for females was 7-5 min longer than for males, also a statistically significant TABLE 2. Expt. 2. Mean times (min) to the onset of severe poisoning in rats exposed to oxygen at a pressure of 6 atm, and treated with various drugs Control Adrenalectomized Difference Male 35* ±4.0* Female ±4-0 Mean ±2t8 Mean No drug Cortisone DOCA Thyroid (±2-0) No adrenaline *0 42' Adrenaline Mean (±2 8) Difference (±5 6) ± 2.8* Unoperated rats only Mean No drug Cortisone DOCA Thyroid (+1-8) No adrenaline * Adrenaline Mean (±2 6) Difference (+5 2) * * s.e. of mean. difference. No large effects of the drugs appeared, though there was some suggestion that cortisone increased and thyroid decreased the time to onset; adrenaline appeared to decrease the time somewhat iii the absence of other drugs or in the presence of cortisone, but to increase it in the presence of DOCA. None of these drug effects was clearly significant. Table 3 summarizes the records of lung damage. The reduction in damage in the adrenalectomized animals was highly significant, the sex difference being negligible. DOCA and thyroid both significantly increased the lung damage, irrespective of whether or not the animals were adrenalectomized, but cortisone and adrenaline appeared to be practically without effect. In twenty-one animals tissue was present which microscopic examination showed closely to resemble suprarenal cortical tissue. A typical example is illustrated in Pl. 1 b and compared with normal rat tissue in P1. 1 a. The times to the onset of severe poisoning in these twenty-one rats were apparently distributed at random over the whole range; four gave values of 90 min or more.

5 ADRENALECTOMY AND OXYGEN POISONING 27 TABLE 3. Expt. 2. Mean index of lung damage in rats exposed to oxygen at a pressure of 6 atm and treated with various drugs i No drug 2-71 Male Female Mean Control * Cortisone 2*73 Adrenalectomized 2* *45 DOCA 3-60 * ±S.E. of mean. Difference - 1* * - 1*42±0* *21 Thyroid 3*27 (±0-21) Experiment 3. As already stated, despite special precautions in the previous experiment the variability remained high, and it was thought that the effects of drugs, particularly of DOCA, might therefore be masked. As a further test, unoperated rats 70 adrenalectomized rats 30 ns z F1 I I Time (min) to onset of severe poisoning Fig. 1. Block diagram showing distribution of times to the onset of severe poisoning in all adrenalectomized and unoperated rats, exposed for the first time and given no drug treatment. thirty-two rats, sixteen male and sixteen female, were adrenalectomized and, on the fourteenth post-operative day, were exposed for 50 min to oxygen at 6 atm. All showing any evidence of poisoning were discarded, and the others were put at random into two groups, each of four males and three females. One group was given injections of DOCA for 8 days, the other was untreated. Fourteen days after the initial exposure, both groups were again exposed to oxygen at 5 atm, except for one animal of the control group which had died. I I

6 28 D. W. TA YLOR The times to onset of severe poisoning were on average almost equal, min and min for the control and DOCA groups respectively. Contrary to the findings of Expt. 2, the lung damage also was almost the same in the two groups, O57 and respectively; however, with standard errors as large as these, an effect on lung damage could escape detection. Experiment 4. Two series of adrenalectomized rats were used, each consisting of the survivors of thirty-two rats exposed in precisely the same way as in the preliminary part of Expt. 3. The two series were tested on different occasions, but for both experimental conditions were as far as possible the same as in Expt. 3 and the results are therefore presented together. Series 1 consisted of fourteen rats, seven of which were given no drug and seven ' cortin'; one of the control group died. Series 2 consisted of twenty rats, of which five were assigned to each of the groups ' no drug', 'cortin', ' adrenaline', 'cortin + adrenaline'; one in each of the groups receiving adrenaline died. No account was taken of sex. Table 4 summarizes the mean times to onset of severe poisoning. The two series agreed in showing a substantial reduction in time to the onset of poisoning for animals receiving cortin, the difference of min being highly significant. The slight reduction associated with adrenaline in Series 2 was not statistically significant, but agrees with the indication of a similar small effect in Expt. 2 in the absence of other drugs. Lung damage was not assessed. Experiment 5. Forty-eight rats were adrenalectomized and exposed to oxygen as in Expts. 3 and 4, but on the fourth post-operative day. After rejection of any that showed evidence of sensitivity, the sixteen survivors were divided into three groups of six, five and five rats. The first group was retained as a control, and the other two had divided doses of hydrocortisone for 5 days at 0-1 mg/100 g body wt. and 1x0 mg/100 g body wt. daily. On the ninth day after operation, the rats were again exposed to oxygen at 6 atm. Table 5 shows the results. Hydrocortisone markedly reduced the time to convulsion; the difference between the two doses suggested a lesser reduction at the higher dose, but this was not significantly different from the lower dose. Lung damage appeared to be rather greater in the rats that received hydrocortisone, but on these few animals the effect was not sufficiently great to be regarded as clearly demonstrated. Experiment 6. Richter & Uhlenhuth (1954) have suggested that, under repeated inbreeding, laboratory rats become progressively less dependent on the suprarenal cortical tissue than are wild rats, and that the amount of this tissue is much less than in the wild strain. If this were so, the variety in sensitivity observed in the operated animals in this series of experiments might be due, in part, to the strain of rats. In order to examine this point, a group of twelve

7 ADRENALECTOMY AND OXYGEN POISONING 29 male black rats, of the strain described earlier, was compared with a similar group of hooded rats. Six of each group were adrenalectomized and, 6 days later, all were exposed to oxygen at 6 atm. The difference in weight of suprarenal tissue in the two strains was very great, averaging 13*2 mg/100 g body wt. for the hooded and 20-1 mg/100 g body wt. for the black; the standard error of each mean was 05 mg/100 g; so TABLE 4. Expt. 4. Mean times (min) to the onset of severe poisoning in adrenalectomized rats exposed to oxygen at a pressure of 6 atm and treated with various drugs No cortin Cortin Difference Series 1 71* ±6.0* Series 2, no adrenaline ±6X8 Series 2, adrenaline ±7-e Weighted mean difference ±3-9 * ±s.e. of mean. TABLE 5. Expt. 5. Mean times (min) to the onset of severe poisoning and mean index of lung damage in adrenalectomized rats exposed to oxygen at a pressure of 6 atm and treated with hydrocortisone Daily dose of hydrocortisone None 0.1 mg/lo0 g 1.0 mg/100 g Mean time (min) to onset of * 33 0±4* severe poisoning Lung damage 1-0± ±06 2-2i0-6 * S.E. of mean. TABLE 6. Expt. 6. Mean times (min) to the onset of severe poisoning and mean index of lung damage in hooded and black rats exposed to oxygen at a pressure of 6 atm Time (min) to the onset of severe poisoning Hooded Black Mean Control Adrenalectomized Difference 32* ±6.1* *7+6* ±4-3 Lung damage I A Control Adrenalectomized Difference Hooded Black 3-7 1* ± Mean i56 * +S.E. of mean. that the difference was highly significant. The hooded rats were slightly heavier, averaging 415 g against 380 g for the black. Nevertheless, as Table 6 shows, the strains scarcely differed in their reactions to adrenalectomy. The hooded gave rather shorter times to onset of poisoning and rather greater lung damage than the black, whether or not they were adrenalectomized, but differences between operated and control groups were very similar for the two strains. The experiment involved too few rats to detect small differences between strains, but there was certainly no evidence of any large difference.

8 30 D. W. TAYLOR Experinent 7. Hartman & Brownell (1949) have stated that accessory cortical tissue occurs in a high proportion of young rats. If the variability of previous experiments were due to the presence of functioning accessory tissue, young animals might fail to show increased resistance after adrenalectomy more frequently than old. This hypothesis was examined by exposing seven groups of eight rats, four males and four females in each, to oxygen at 6 atm. The groups were aged 1, 2, 3,..., 7 months. Two males and two females in each group were adrenalectomized. Three rats died. TABLE 7. Expt. 7. Effect of age on mean time (min) to the onset of severe poisoning in rats exposed to oxygen at a pressure of 6 atm Age of rats (months) Control Adrenalectomized Difference TABLE 8. Expt. 7. Effect of age and of sex on mean index of lung damage in rats exposed to oxygen at a pressure of 6 atm Age of rats (months) S.E. Control 2-5 3* Adrenalectomized 2-0 0* Male Female Mean (±0 24) Control Adrenalectomized S.E ±8-0 ±8-2 Mean (±0'24) *05 As usual, the times to onset of severe poisoning were consistently longer and much more variable for the adrenalectomized rats. Table 7 shows the mean times. The unoperated rats gave evidence of a small but steady increase in time with increasing age, amounting to about min/month; both the times and the rate of increase were rather greater for females than for males. The operated rats showed a much greater rate of increase, min/ month, but the greater variability prevents any certainty that this rate differs from the other; there was no sign of any sex difference. Although by no means conclusive, the results of the experiment are in accordance with the hypothesis under test. At 1 month of age the difference between adrenalectomized and normal rats in respect of tolerance of oxygen at high pressure appears to be small, but it increases quite markedly at any rate up to 7 months. There was no evidence that age affected lung damage (Table 8), but damage was significantly less for the adrenalectomized rats and significantly less for females than for males.

9 ADRENALECTOMY AND OXYGEN POISONING 31 Experiment 8. If repeated exposure of rats to oxygen under high pressure were to lead to suprarenal cortical hypertrophy, then such hypertrophy should affect any accessory cortical tissue and repeated exposure might reasonably be expected to lead to an increased sensitivity to oxygen. Moreover, if postoperative maintenance on 0-9 % NaCl solution were to give more protection against the full development of cortical insufficiency, thus tending to increase sensitivity to oxygen, then maintenance of the animals on water might have the opposite effect. Expt. 8 was planned as a test of these possibilities. TABLE 9. Expt. 8. Mean times (min) to the onset of severe poisoning in control and saline- and water-maintained, adrenalectomized rats on single or after repeated exposure to oxygen at a pressure of 6 atm Adrenalectomized No. of With 0-9% Mean exposures Control With H20 NaCl solution +S.E. Single ±3-7 Repeated ±3-7 Mean±s.E. 32-0± Six litters of rats (four of black and two of hooded) were used, each consisting of three males and three females. Four of each litter were adrenalectomized, two being thereafter maintained on 0-9% NaCl solution and two on water; the other two were kept as unoperated controls. One animal from each of these pairs was exposed to oxygen at 6 atm for 20 min on 5 of the 7 days preceding final exposure on the fifteenth post-operative day. Four rats, all adrenalectomized and maintained on water, died before the final exposure. Of the eight other rats that were adrenalectomized and maintained on water, seven lost weight between operation and final exposure; on the other hand, of the twelve adrenalectomized rats maintained on NaCl solution, ten gained in weight between operation and final exposure. Adrenalectomy again increased the time to onset of severe poisoning, to an extent similar to that of other experiments (Table 9). There was little difference between the operated rats maintained on water and on NaCl solution. The control rats showed the same time to onset, irrespective of whether or not they had had repeated exposure before the final tests; the adrenalectomized showed somewhat shorter times if they had been exposed repeatedly, though the difference from the unexposed was not statistically significant. Neither sex nor strain of rat appeared to modify these effects. There was no correlation between change in weight of rats and susceptibility to oxygen. Table 10 summarizes results in respect of lung damage. The adrenalectomized rats had less damage, but variability from animal to animal was high and the difference from the controls was not statistically significant. No consistent difference between water and NaCl appeared, nor was there any effect of repeated exposure. Indeed, the only clear result from the lung

10 32 D. W. TAYLOR damage records was the sex difference, 2-1 for males and 1-2 for females with a standard error of for each. Distribution of times to the onset of severe poisoning in unoperated and adrenalectomized rats. The figures for time to onset of severe poisoning for all rats in Expts. 1-8, irrespective of age, strain or sex, but excluding those given any drug treatment, and those not being exposed for the first time, have been used to form a block diagram (Text-fig. 1). TABLE 10. Expt. 8. Mean index of lung damage in control and saline- and water-maintained adrenalectomized rats on single or after repeated exposure to oxygen at a pressure of 6 atm Adrenalectomized No. of With 0.9% Mean exposures Control With H20 NaCl solution ±S.]. Single ±0-3 Repeated Mean +S.E. 2-2± ± ± It is easily seen that of the sixty-six unoperated rats the overwhelming majority gave values lying between 20 and 40 min. In the case of the seventy operated animals the values lie between 20 and 90 min, half of them being between 20 and 50 min, but distributed more widely than are the controls; half are similarly distributed between 50 and 90 min. A very few of the experiments in this latter group were terminated, as explained earlier, before unequivocal manifestations of toxicity were observed. Longer exposures in these cases would have extended the graph still farther to the right, while lowering its peak, and there was no definite evidence of a bimodal distribution. DISCUSSION The experiments which have been described show clearly that unanaesthetized, adrenalectomized rats are more resistant to the effects of oxygen under pressure than are unoperated controls. This result was obtained independently by Bean, Johnson & Smith (1953) and by Gerschman & Nadig (1953). The increase in resistance was much less obvious in terms of lung damage than in the case of the central nervous system effects, and there was no correlation between the two types of effect. Cardiac and respiratory slowing are known to occur in animals exposed to oxygen under high pressure. Unpublished experiments by the author have shown that anaesthetized, adrenalectomized rats are just as liable to the cardiac and respiratory slowing produced by oxygen as are normal animals. The two groups therefore seem to differ mainly in their responses to the action of oxygen under pressure on the central nervous system, the difference between operated and unoperated rats in this respect being very great, despite the variability in the responses of the former. If one is to accept the existence of a 'general adaptation syndrome' as postulated by Selye (1949), one is forced to conclude that, exposed to this

11 ADRENALECTOMY AND OXYGEN POISONING 33 particular 'stress', adrenalectomized animals behave in a way opposite to that in which they behave when exposed to more orthodox 'stressor agents'. Indeed, Gerschman & Fenn (1954) have suggested that oxygen at high pressure is an 'unnatural stress', and that therefore the ordinary rules may not apply. It seems better therefore to leave aside the whole vexed question of 'stress' and to attempt an explanation in terms of factors already known to influence the course of oxygen poisoning. Bergen, Hunt & Hoagland (1953) and Hoagland (1955) showed that the rate of cerebral blood flow in adrenalectomized, saline-maintained rats was considerably less than in normal rats. There was also a reduction of 18 % in brain oxygen consumption, and these changes were accompanied by a reduction in the frequency of the electrocorticogram. The probable importance in oxygen poisoning of any factor which reduces oxygen utilization at the cellular level has been discussed (Taylor, 1956). Lambertsen, Kough, Cooper, Emmel, Loeschchke & Schmidt (1953) found a decrease in cerebral blood flow in human subjects exposed to oxygen at high pressure, and suggested that this, by lowering the average oxygen pressure at which the tissues of the brain were functioning, would have a protective effect. If such a decrease in cerebral blood flow were exaggerated by adrenalectomy, this by itself might go far in explaining the protective effect of the operation. Given that adrenalectomy had a definite protective effect, it seemed probable that replacement therapy by one or other of the substances in common use might reverse the effect, and in so doing perhaps point to a possible cause. The variation in the responses of the operated animals made it necessary, after Expt. 2, to use animals which on previous exposure had been shown to be resistant to the effects of oxygen. DOCA was not effective in reversing the protection given to the central nervous system by adrenalectomy; and this, together with the absence of any significant difference between saline-treated and water-treated animals in Expt. 8, seems to show that alterations in water and electrolyte metabolism resulting from the operation are not responsible for the difference between operated and unoperated animals. Cortisone did not significantly alter the sensitivity of either group. In fact some of the highest times to onset of severe poisoning were recorded in cortisone-treated, adrenalectomized animals. Bergen et al. (1953) and Hoagland (1955) showed that the cerebral, circulatory and metabolic changes occurring after adrenalectomy were abolished by administration of whole cortical extract or cortisone, but not of DOCA. The increase in sensitivity to oxygen brought about by cortin, and the absence of any such increase with DOCA, would support the view that such changes were responsible for the increased resistance of adrenalectomized animals, while the results obtained with cortisone are a stumbling-block. Gerschman, Gilbert, Nye, Nadig & Fenn (1954) found that cortisone in small doses increased 3 PHYSIO. CXL

12 34 D. W. TAYLOR sensitivity to oxygen significantly in adrenalectomized rats only, but in large doses had the opposite effect, decreasing sensitivity in both operated and control groups. The dose of cortisone used in Expt. 2 corresponded to the large dose of Gerschman and her colleagues, and this might possibly explain some of the results. On the other hand, cortisone had no effect whatsoever on control rats. It is interesting, too, that in Expt. 5 both large and small doses of hydrocortisone had the effect of reducing tolerance in adrenalectomized rats. According to Vogt (1954) the differences between the actions of hydrocortisone and cortisone are only quantitative, hydrocortisone being more active in most respects; of the two, the rat secretes mainly cortisone. It is strange therefore that cortisone failed to show any definite effect in Expt. 2. Bean & Johnson (1954) found that adrenaline augmented the adverse action of oxygen at high pressure, confirming the work of Campbell (1937). Gerschman et al. (1954) and Gerschman, Gilbert, Nye, Price & Fenn (1955) thought that adrenaline had some effect but concluded that the adrenal cortex played a more important role in oxygen poisoning than did the medulla; this is supported by the results of Expts. 2 and 4. There was a suggestion that thyroid in Expt. 2 decreased the time to onset of severe poisoning in unoperated animals. Campbell (1937), Gersh & Wagner (1944) and Grossman & Penrod (1949) showed that thyroid had such an effect. On the other hand, thyroid had no significant effect on operated animals and some of the times to onset of severe poisoning were amongst the longest recorded. The general opinion is that administration of thyroid increases the requirements for cortical hormone (Pincus & Thimann, 1955). Presumably any tendency to deficiency could be counteracted in an intact animal, but obviously not after adrenalectomy. In the first case thyroid could exert an independent effect, increasing sensitivity to oxygen under pressure, while after adrenalectomy any effect could well be masked by the degree of protection given by the loss of the adrenals. It is worth noting that administration of thyroid aggravated the effects of adrenalectomy before exposure to oxygen since, in Expt. 2, all deaths but one before exposure to oxygen were in thyroidtreated operated rats. One of the most striking features of these experiments is the wide range of values for the time to onset of severe poisoning in the adrenalectomized animals as a group. This made it difficult to draw conclusions even when relatively large numbers were used. The most likely cause is failure to remove all the cortical tissue in the body, with subsequent hypertrophy of cortical 'rests', not necessarily in the perirenal area. Hartman & Brownell (1949, quoting Jaffe) state that macroscopic amounts of accessory cortical tissue occur in 8% of normal rats and in % after removal of the suprarenal glands, and that microscopic accessories have been detected in 70% of immature rats. In Expt. 2 histological examination

13 ADRENALECTOMY AND OXYGEN POISONING 35 showed that tissue resembling suprarenal cortical tissue was present in about one quarter of the operated animals, but the values for the time to onset of severe poisoning could not be correlated with the presence of such tissue, and indeed four out of the twenty-one animals concerned had very high values. The whole situation is probably so complicated that these results cannot be held to dispose of the hypothesis that adrenal cortical rests are in fact responsible for the variation in tolerance, especially when one remembers the difficulty involved in identifying such tissue at the site of operation and the impracticability of determining just how much may be present at other sites in the body, quite apart trom its functional status. But equally, much more experimentation is required before the hypothesis can be finally accepted. SUMMARY 1. The effects of adrenalectomy and of substitution therapy on the manifestations of oxygen poisoning were studied in rats of two strains, exposed to a pressure of 6 atm of pure oxygen. 2. Adrenalectomy gave very definite protection against the central nervous system manifestations of oxygen poisoning, and gave some protection against lung damage. 3. The variability in time of appearance of nervous system effects in adrenalectomized animals was very great. This variability was not confined to either one of the two strains tested, although these possessed suprarenal glands that differed considerably in weight. It was not correlated with the presence of small amounts of residual cortical tissue in the perirenal region. It may possibly depend on accessory cortical tissue elsewhere in the body. 4. There was an increase with age in the time to onset of severe poisoning; this was greater in adrenalectomized than in unoperated animals, but the greater variability in the former made it impossible to say that this represented a real difference. 5. Of the drugs tested, cortin and hydrocortisone were capable of increasing susceptibility of operated animals to the effects of oxygen on the central nervous system. Cortisone, DOCA and thyroid had no significant effect. There was a possibility that adrenaline slightly increased susceptibility. 6. There was no significant difference between the responses of adrenalectomized animals maintained on 0 9 % saline and those maintained on water. In general there was no relation between the time to onset of severe poisoning and gain or loss in weight. 7. Possible explanations for the difference between adrenalectomized and unoperated animals are discussed. I am indebted to the Statistical Department, University of Aberdeen, and especially to Mr I. McDonald, Mr J. Fraser Scott and Miss A. D. Outhwaite, for advice in the design of these experiments and for the statistical analyses. I wish to thank particularly Dr D. J. Finney for much 3-2

14 36 D. W. TAYLOR assistance and advice in presenting the results. My thanks are also due to Mr M. Hay for technical assistance. The work was assisted by an expenses grant from the Medical Research Council. REFERENCES BEAN, J. W. (1951, Adrenal alteration induced by oxygen at high pressure. Fed. Proc. 10, 11. BEAN, J. W. & JOHNSON, P. (1954). Epinephrine and neurogenic factors in the pulmonary oedema and C.N.S. reactions induced by oxygen at high pressure. Amer. J. Physiol. 180, BEAN, J. W., JOHNSON, P. & SMITH, C. W. (1953). The influence of hypophyseal and adrenocortical factors on the chronic effects, especially the motor disability, induced by exposure to oxygen at high pressure. Abstr. XIX int. physiol. Congr. pp BERGEN, J. R., HUNT, C. A. & HOAGLAND, H. (1953). Effects of adrenalectomy and replacement therapy on brain circulation, oxygen consumption and the electrocorticogram. Amer. J. Physiol. 175, CAMPBELL, J. A. (1937). Oxygen poisoning and the thyroid gland. J. Physiol. 90, 91 P. CARLETON, H. & LEACH, E. H. (1947!. Histolo ical Technique, 2nd ed. London. Oxford University Press. GERSH, I. & WAGNER, C. E. (1944). Metabolic factors in oxygen poisoning. Amer. J. Physiol. 144, GERSCHMAN, R. & FENN, W. 0. (1954). Ascorbic acid content of adrenal glands of rat in oxygen poisoning. Amer. J. Physiol. 176, 6-8. GERSCHMAN, R., GILBERT, D. L., NYE, S. W., NADIG, P. W. & FENN, W. 0. (1954). Role of adrenalectomy and adrenal-cortical hormones in oxygen poisoning. Amer. J. Physiol. 178, GERSCHMAN, R., GILBERT, D. L., NYE, S. W., PRICE, W. E. & FENN, W. 0. (1955). Effects of autonomic drugs and of adrenal glands on oxygen poisoning. Proc. Soc. exp. Biol., N.Y., 88, GERSCHMAN, R. & NADIG, P. W. (1953). Stress and oxygen poisoning. Fed. Proc. 12, 159. GROSSMAN, M. S. & PENROD, K. E. (1949). The thyroid and high oxygen poisoning in rats. Amer. J. Physiol. 156, HARTMAN, F. A. & BROWNELL, K. A. (1949). The Adrenal Gland. London: Henry Kimpton. HOAGLAND, H. (1955). In Neurochemistry, pp , ed. ELLIOTT, K. A. C., PAGE, I. H. & QUASTEL, J. H. Springfield, U.S.A.: C. C. Thomas. LAMBERTSEN, C. J., KOUGH, R. H., COOPER, D. Y., EMMEL, G. L., LOESCHCHKE, H. H. & SCEMIDT, C. F. (1953). Oxygen toxicity. Effects in man of oxygen inhalation at 1 and 3-5 atmospheres upon blood gas transport, cerebral circulation and cerebral metabolism. J. appl. Physiol. 5, PiNCUS, G. & THIMANN, K. V. (1955). The Hormones, Vol. III. New York: Academic Press Inc. RICHTER, C. P. & UHLENHUTH, E. H. (1954). Comparison of the effects of gonadectomy on spontaneous activity of wild and domesticated Norway rats. Endocrinology, 54, SELYE, H. (1949). Textbook of Endocrinology, 2nd ed. Montreal: Acta Endocrinologica Inc. TAYLOR, D. W. (1953). Modifying effects of adrenalectomy on 02 poisoning in the rat. Abstr. XIX int. physiol. Congr. pp TAYLOR, D. W. (1954). Effects of high oxygen pressures on adrenalectomized, treated and untreated rats. J. Physiol. 125, P. TAYLOR, D. W. (1956). The effects of vitamin E and of methylene blue on the manifestations of oxygen poisoning in the rat. J. Physiol. 131, THOMSON, W. (1936). Stock diet for rats. J. Hyg., Camb., 36, VOGT, M. (1954). The role of the adrenal gland in homeostasis. Quart. J. exp. Physiol. 39, EXPLANATION OF PLATE Plate 1. (a) Normal adrenal cortical tissue from rat; (b) tissue removed from perirenal region of rat whose adrenals had been excised 14 days previously.

15 THE JOURNAL OF PHYSIOLOGY, VOL. 140, No. 1 PELAE 1 L mm 0.1 mm (Fl+'iciny p- 36;)