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The Action of Avertin on the Ureter
THE ACTION OF AVERTIN ON THE URETER J. A. WADDELL
From the Department of Pharmacology, University of Virginia, University, Virginia.
In their comprehensive summary of the literature on avertin, Anschultz et al. in 1930 (1) cited clinical and experimental data as to its effects on the kidney and urine but reported no observations as to the urinary passages. Waddell in 1931 (2), however, in the course of an investigation of the effects of the drug on musculature in general, demonstrated a depressant action on the ureters of cats, pigs, and calves. He employed suspended segments for observations on changes in rhythmicity and tone and perfused organs for those on alterations in patency of the lumen. Since the ureter in the work cited above proved to be highly reactive to avertin, further study ,vas undertaken, amplifying the previous work and extending the observations to the living animal. Accordingly, this paper deals with the changes in rhythmic activity, internal pressure, and patency of the ureter, both excised and in situ, as influenced by avertin, when applied directly to its mucous membrane or brought to it through the blood stream. TECHNICAL PROCEDURES
The avertin (tribromethyl alcohol) employed in this investigation was the pure crystalline substance-not avertin-fluid, which contains amylene hydrate. It was prepared from the latter by evaporation in the dark at 20°C., repeatedly washed with distilled water, and air-dried. The long, glassy crystals thus obtained were preserved in glass-stoppered bottles and showed no evidence of deterioration within nine months. The solutions of the drug were always prepared within a fe,v minutes of their intended use. According to the purpose for which they were to be employed, the solvents were sodium chlo707
708
J. A. WADDELL
ride solution or Ringer's solution. The ranges of dilution were from 1: 3000 for excised tissue to 1: 40 for direct application to mucous membranes and for rectal administration. To facilitate dissolving, the crystals were triturated with portions of the solvent at 40°C. until a fine suspension was obtained; this was diluted to the required strength and the preparation was maintained at the above temperature until thoroughly dissolved. Then the material, thoroughly shaken, was brought to the working temperature and retained at that point until used. As mentioned above, both excised and intact ureters were studied. The former, taken from recently dead animals, were either suspended and immersed in saline for a direct lever tracing or perfused with saline; the latter, in situ in anesthetized animals, were always perfused with saline, while the drug was administered either in the perfusate for local action or to the animal, rectally or intravenously. The details as to the technic employed were necessarily different according to the way in which the tissue was manipulated and the drug administered; hence, they can be best presented along with the experimental data. It may be further stated here, however, that kymographic records, showing the changes in ureteral activity, were made in all instances. EXPERIMENTAL DATA
The experimental data can best be presented under captions appropriate to the method of handling the tissue and applying or administering the drug. 1. The excised ureter Freshly excised ureters of dogs, pigs, and calves were stripped of their connective tissue and examined, using (a) suspended sections for observations of changes in rhythm and tone, and (b) perfused whole organs for those of alterations in internal pressure and passage of fluids. The ureter of the pig proved to be best for the former purpose and that of the dog for the latter. a. Suspended sections. Longitudinal sections of about 1 inch in length were suspended in a small reservoir and arranged for a direct lever tracing, according to the Magnus method (3). The
ACTION OF A VERTIN ON URETER
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immersion fluid employed was Ringer's solution containing 1 gram of sodium bicarbonate per liter. In this, at temperatures from 37° to 38°C., the tissue exhibited rhythmic contractions and tone changes, maintaining its vital activities for hours. On introducing avertin into the reservoir, depressant effects were shown almost immediately. Concentrations of 1: 2000 usually produced arrest of movement and profound relaxation. Ones of 1: 3000 decreased rhythmicity and tone, and occasionally produced complete inhibition of all activity. Even after the higher concentrations of the drug, its removal by changing the tissue to a plain saline bath was quickly folowed by a recovery in all the functions of the musculature. Avertin apparently does not permanently damage the tissue.
Fm. 1.
RELAXATION OF THE EXCISED URETER IN A SOLUTION OF AVERTIN
Ureter (pig; suspended; immersed in Ringer's solution at 37°C.), showing depressant effects of avertin.
b. Perfused ureters. Cannulas were inserted in the extremities of whole ureters and perfusion was effected in accordance with the method of Trattner, Wright, and Barlow (4). The perfusate was 0.9 per cent sodium chloride solution. This was admitted through the cannula in the proximal end of the organ at a temperature of 37°C. and a pressure of 20 cm. (water). The outflow was conducted by rubber tubing through a membrane manometer, which recorded intraureteral pressure, to a drop counter of the type described by Biskin and Dan (5). With this arrangement of the apparatus, the ureter proved to be very active and exhibited rhythmic changes in internal pressure and in outflow. Usually these changes were irregular as in figure 2, but occaTHE JOURNAL OF UROLOGY 1 VOL, XXIX, NO.
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sionally they were as regular as the contractions exhibited by theexcised intestine of the rabbit. Perfusion with avertin, dissolved in the above saline, produced a decrease in intraureteral pressure and an increase in the outflow, which demonstrated a relaxation of the_ organ. With concentrations of 1: 1000 and higher, the pressure curve became a straight line and the outflow lost its periodic character, phenomena indicative of profound relaxation . with the production of an inert tube-like condition in the passage-
Fro.
2.
REDUCTION OF SPASM IN THE ExcrsED URETER ON PERFUSION WITH A SOLUTION OF AVERTIN
Ureter (dog; perfused with 0.9 per cent NaCl at 37°C.), showing spas,m produced by barium chloride and its reduction by avertin dissolved in 0.9 per cent NaCl.
way. Removal of the drug by returning to the initial perfusate, was followed by a decrease in the outflow and a resumption of rhythmic activity. The increase in rhythmic activity produced by the nerve stimulants, pilocarpine and arecoline, could readily be antagonized by perfusion with avertin, as could also the spasm following the application of the muscle stimulants, barium and potassium. It was more effective than atropine for the former or benzyl'esters for the latter.
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ACTION OF A VERTIN ON URETER
711
2. The intact ureter In this portion of the investigation, the study was devoted to the effects of avertin on the ureter, in situ, in the living animaL Dogs were used throughout. Amytal was administered, intraperitoneally, sufficient to produce a basal anesthesia and ether was supplemented during the operative procedures. The bladder was exposed through an incision just above the symphysis pubis and a cannula was inserted in the ureter close to its distal extremity. This was secured with a ligature, care being exercised to exclude the visible vessels. Then the wound was sutured, leaving the end of the cannula exposed. The renal extremity of the ureter was exposed extraperitonea1ly at the pelvis of the kidney and the tip of a trochar was placed in its lumen after being thrust through the kidney substance and the pelvis. No ligature was applied to the ureter here, the resistance of the kidney being sufficient to prevent backflow. In this way the most vital portion of the blood supply to the ureter was not disturbed. In general, the method employed was that of Trattner, Wright, and Barlow (4). The trochar, used as an intake cannula, was connected with the perfusion reservoir and the cannula in the distal extremity with the apparatus for recording the outflow and pressure changes. The procedure for perfusing was the same as was employed for the excised organ and has been described above. The perfusate was 0.9 per cent sodium chloride solution. Slightly higher pressures were required than in the case of the excised, about 30 cm. as compared with 20 cm. When the temperature and pressure had been properly adjusted, the in situ ureter exhibited activity like the excised, except that respiratory movements were in evidence on the graphs at times and the pressure changes were not as pronounced. Obviously a very even depth of anesthesia was desirable and non-interference with the circulation imperative. a. In sitn ureter, avertin being administered in the perfusate. The results obtained on perfusing the intact ureter with solutions of avertin in 0.9 per cent sodium chloride were qualitatively
712
J. A. WADDELL
identical with those on the excised perfused organ; i.e., a decrease in internal pressue with a corresponding increase in outflow, which progressively became more continuous under the higher concentrations. Dilutions of 1: 2000 were very effective within a few seconds. Spasm, which had been produced by barium chloride or potassium chloride, was readily reduced by application of avertin, as was also the increase in tone and peristalsis following the intravenous administration of pilocarpine and arecoline. No systemic effects on the animal were shown on perfusing the ureter with avertin. Such would hardly be expected, in view of the small quantity of drug employed and the high resistance of the
FIG.
3.
REDUCTION OF SPASM IN THE INTACT URETER ON PERFUSION WITH A SOLUTION OF AVERTIN
Ureter (dog; in situ; perfused with 0.9 per cent NaCl at 37°C.), showing spasm produced by barium chloride and its reduction by avertin dissolved in 0.9 per cent NaCl.
dog, even were absorption to take place as has been observed in the case of the bladders of rats (6). b. In situ ureter, the drug being administered intravenously. With the apparatus arranged for perfusion under constant conditions, avertin was introduced into the femoral vein of the animal. Doses of from 50 to 200 mgm. per kilogram were employed. Depressant effects on the ureter were produced almost immediately. These with the smallest doses were barely appreciable, while with the largest asphyxia! phenomena disturbed the records. Doses of 100 mgm. per kilogram, which are considerably less than the intravenous dose for basal anesthesia in dogs,
ACTION OF .AVERTIN ON URETER
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were effective in reducing the spasm produced by adding potassium and barium salts to the perfusate and also in counteracting the effects of pilocarpine and arecoline given by vein. c. In situ ureter, the drug being administered rectally. Since avertin is usually administered per rectum, experiments were performed to determine whether on employing this route effects
FIG. 4.
REDUCTION OF SPASM IN THE INTACT URETER ON ADMINISTERING AvERTIN INTRA VENOUSLY
Ureter (dog; in situ; perfused with 0.9 per cent NaCl at 37°C.), showing spasm produced by potassium chloride and its reduction following the intravenous administration of avertin.
FIG. 5.
RELAXATION OF THE INTACT URETER ON ADMINISTERING AVERTIN RECTALLY
Ureter (dog; in situ; perfused with 0.9 per cent NaCl at 37°C.), showing depressant effects of avertin, administered by rectum.
could be produced on the ureter. The animals were prepared and the apparatus was arranged as described above, except that a rectal tube was inserted and secured in position by a ligature placed around the anal canal. The results were in brief the same as in the other perfusion experiments, but quantitatively
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much less pronounced. Doses below 200 mgm. per kilogram were not effective; 400 mgm. per kilogram introduced complications as a result of asphyxia, due doubtless to the prior use of amytal. However, it should be noted in this connection that the rectal dose for the dog is about 500 mgm. per kilogram as a basal anesthetic. Spasms produced by barium and potassium salts were reducible by 200 mgm. per kilogram and this quantity also antagonized effectively the actions of pilocarpine and arecoline. Doubtless, in an animal not already in the state of basal anaesthesia, smaller doses would be effective, since absorption from the alimentary canal would be more active. SUMMARY
1. The effects of avertin on the excised and intact ureter have been described. 2. Avertin very actively depresses the musculature of the excised ureter, producing decrease in tone and rhythmicity and increase in patency. 3. Avertin introduced into the in situ ureter, or administered intravenously or rectally, similarly reduces the tone and rhythmicity and increases the patency. 4. Ureteral spasm in the dog may be reduced by introducing avertin into the lumen of the organ or by intravenous and rectal administration. CONCLUSIONS
1. Avertin does not injure the ureter, the effects being immediately abolished on withdrawing the drug. 2. Avertin may be useful in urological work in reducing ureteral spasm and in other conditions where relaxation is desirable. REFERENCES (I) ANSCHULTz, W., SPECHT, K., AND TIEMAN, F.: Avertin narcosis in surgery, Ergebn. d. Chir. u. Orthop., 1930, xxiii, 406. (2) WADDELL, J. A.: The action of avertin on voluntary and non-voluntary muscle. Jour. Lab. and Olin. Med., 1932, xvii, 1104.. (3) MAGNIS, R.: Experiments on the surviving intestine. Arch. f. d. ges. Physiol., 1905, cviii, 1.
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(4) TRATTNER, H. R., WRIGHT, H. B., AND BARLOW, 0. W.: An experimental study of the action of sodium iodide on the excised and intact ureters of dogs. Jour. Urol., 1930, xxviii, 441. (5) BISKIN, M. S., AND DAN, lVI.: An automatic drop recorder. Proc. Soc. Exper. Biol. and Med.,, 1928, xxvi, 52. (6) WADDELL, J. A.: Absorption of avertin from the bladder. Unpublished data. Department of Pharmacology, University of Virginia, 1932.
From the Department of Pharmacology, University of Virginia, University, Virginia.
In their comprehensive summary of the literature on avertin, Anschultz et al. in 1930 (1) cited clinical and experimental data as to its effects on the kidney and urine but reported no observations as to the urinary passages. Waddell in 1931 (2), however, in the course of an investigation of the effects of the drug on musculature in general, demonstrated a depressant action on the ureters of cats, pigs, and calves. He employed suspended segments for observations on changes in rhythmicity and tone and perfused organs for those on alterations in patency of the lumen. Since the ureter in the work cited above proved to be highly reactive to avertin, further study ,vas undertaken, amplifying the previous work and extending the observations to the living animal. Accordingly, this paper deals with the changes in rhythmic activity, internal pressure, and patency of the ureter, both excised and in situ, as influenced by avertin, when applied directly to its mucous membrane or brought to it through the blood stream. TECHNICAL PROCEDURES
The avertin (tribromethyl alcohol) employed in this investigation was the pure crystalline substance-not avertin-fluid, which contains amylene hydrate. It was prepared from the latter by evaporation in the dark at 20°C., repeatedly washed with distilled water, and air-dried. The long, glassy crystals thus obtained were preserved in glass-stoppered bottles and showed no evidence of deterioration within nine months. The solutions of the drug were always prepared within a fe,v minutes of their intended use. According to the purpose for which they were to be employed, the solvents were sodium chlo707
708
J. A. WADDELL
ride solution or Ringer's solution. The ranges of dilution were from 1: 3000 for excised tissue to 1: 40 for direct application to mucous membranes and for rectal administration. To facilitate dissolving, the crystals were triturated with portions of the solvent at 40°C. until a fine suspension was obtained; this was diluted to the required strength and the preparation was maintained at the above temperature until thoroughly dissolved. Then the material, thoroughly shaken, was brought to the working temperature and retained at that point until used. As mentioned above, both excised and intact ureters were studied. The former, taken from recently dead animals, were either suspended and immersed in saline for a direct lever tracing or perfused with saline; the latter, in situ in anesthetized animals, were always perfused with saline, while the drug was administered either in the perfusate for local action or to the animal, rectally or intravenously. The details as to the technic employed were necessarily different according to the way in which the tissue was manipulated and the drug administered; hence, they can be best presented along with the experimental data. It may be further stated here, however, that kymographic records, showing the changes in ureteral activity, were made in all instances. EXPERIMENTAL DATA
The experimental data can best be presented under captions appropriate to the method of handling the tissue and applying or administering the drug. 1. The excised ureter Freshly excised ureters of dogs, pigs, and calves were stripped of their connective tissue and examined, using (a) suspended sections for observations of changes in rhythm and tone, and (b) perfused whole organs for those of alterations in internal pressure and passage of fluids. The ureter of the pig proved to be best for the former purpose and that of the dog for the latter. a. Suspended sections. Longitudinal sections of about 1 inch in length were suspended in a small reservoir and arranged for a direct lever tracing, according to the Magnus method (3). The
ACTION OF A VERTIN ON URETER
709
immersion fluid employed was Ringer's solution containing 1 gram of sodium bicarbonate per liter. In this, at temperatures from 37° to 38°C., the tissue exhibited rhythmic contractions and tone changes, maintaining its vital activities for hours. On introducing avertin into the reservoir, depressant effects were shown almost immediately. Concentrations of 1: 2000 usually produced arrest of movement and profound relaxation. Ones of 1: 3000 decreased rhythmicity and tone, and occasionally produced complete inhibition of all activity. Even after the higher concentrations of the drug, its removal by changing the tissue to a plain saline bath was quickly folowed by a recovery in all the functions of the musculature. Avertin apparently does not permanently damage the tissue.
Fm. 1.
RELAXATION OF THE EXCISED URETER IN A SOLUTION OF AVERTIN
Ureter (pig; suspended; immersed in Ringer's solution at 37°C.), showing depressant effects of avertin.
b. Perfused ureters. Cannulas were inserted in the extremities of whole ureters and perfusion was effected in accordance with the method of Trattner, Wright, and Barlow (4). The perfusate was 0.9 per cent sodium chloride solution. This was admitted through the cannula in the proximal end of the organ at a temperature of 37°C. and a pressure of 20 cm. (water). The outflow was conducted by rubber tubing through a membrane manometer, which recorded intraureteral pressure, to a drop counter of the type described by Biskin and Dan (5). With this arrangement of the apparatus, the ureter proved to be very active and exhibited rhythmic changes in internal pressure and in outflow. Usually these changes were irregular as in figure 2, but occaTHE JOURNAL OF UROLOGY 1 VOL, XXIX, NO.
6
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J. A. WADDELL
sionally they were as regular as the contractions exhibited by theexcised intestine of the rabbit. Perfusion with avertin, dissolved in the above saline, produced a decrease in intraureteral pressure and an increase in the outflow, which demonstrated a relaxation of the_ organ. With concentrations of 1: 1000 and higher, the pressure curve became a straight line and the outflow lost its periodic character, phenomena indicative of profound relaxation . with the production of an inert tube-like condition in the passage-
Fro.
2.
REDUCTION OF SPASM IN THE ExcrsED URETER ON PERFUSION WITH A SOLUTION OF AVERTIN
Ureter (dog; perfused with 0.9 per cent NaCl at 37°C.), showing spas,m produced by barium chloride and its reduction by avertin dissolved in 0.9 per cent NaCl.
way. Removal of the drug by returning to the initial perfusate, was followed by a decrease in the outflow and a resumption of rhythmic activity. The increase in rhythmic activity produced by the nerve stimulants, pilocarpine and arecoline, could readily be antagonized by perfusion with avertin, as could also the spasm following the application of the muscle stimulants, barium and potassium. It was more effective than atropine for the former or benzyl'esters for the latter.
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711
2. The intact ureter In this portion of the investigation, the study was devoted to the effects of avertin on the ureter, in situ, in the living animaL Dogs were used throughout. Amytal was administered, intraperitoneally, sufficient to produce a basal anesthesia and ether was supplemented during the operative procedures. The bladder was exposed through an incision just above the symphysis pubis and a cannula was inserted in the ureter close to its distal extremity. This was secured with a ligature, care being exercised to exclude the visible vessels. Then the wound was sutured, leaving the end of the cannula exposed. The renal extremity of the ureter was exposed extraperitonea1ly at the pelvis of the kidney and the tip of a trochar was placed in its lumen after being thrust through the kidney substance and the pelvis. No ligature was applied to the ureter here, the resistance of the kidney being sufficient to prevent backflow. In this way the most vital portion of the blood supply to the ureter was not disturbed. In general, the method employed was that of Trattner, Wright, and Barlow (4). The trochar, used as an intake cannula, was connected with the perfusion reservoir and the cannula in the distal extremity with the apparatus for recording the outflow and pressure changes. The procedure for perfusing was the same as was employed for the excised organ and has been described above. The perfusate was 0.9 per cent sodium chloride solution. Slightly higher pressures were required than in the case of the excised, about 30 cm. as compared with 20 cm. When the temperature and pressure had been properly adjusted, the in situ ureter exhibited activity like the excised, except that respiratory movements were in evidence on the graphs at times and the pressure changes were not as pronounced. Obviously a very even depth of anesthesia was desirable and non-interference with the circulation imperative. a. In sitn ureter, avertin being administered in the perfusate. The results obtained on perfusing the intact ureter with solutions of avertin in 0.9 per cent sodium chloride were qualitatively
712
J. A. WADDELL
identical with those on the excised perfused organ; i.e., a decrease in internal pressue with a corresponding increase in outflow, which progressively became more continuous under the higher concentrations. Dilutions of 1: 2000 were very effective within a few seconds. Spasm, which had been produced by barium chloride or potassium chloride, was readily reduced by application of avertin, as was also the increase in tone and peristalsis following the intravenous administration of pilocarpine and arecoline. No systemic effects on the animal were shown on perfusing the ureter with avertin. Such would hardly be expected, in view of the small quantity of drug employed and the high resistance of the
FIG.
3.
REDUCTION OF SPASM IN THE INTACT URETER ON PERFUSION WITH A SOLUTION OF AVERTIN
Ureter (dog; in situ; perfused with 0.9 per cent NaCl at 37°C.), showing spasm produced by barium chloride and its reduction by avertin dissolved in 0.9 per cent NaCl.
dog, even were absorption to take place as has been observed in the case of the bladders of rats (6). b. In situ ureter, the drug being administered intravenously. With the apparatus arranged for perfusion under constant conditions, avertin was introduced into the femoral vein of the animal. Doses of from 50 to 200 mgm. per kilogram were employed. Depressant effects on the ureter were produced almost immediately. These with the smallest doses were barely appreciable, while with the largest asphyxia! phenomena disturbed the records. Doses of 100 mgm. per kilogram, which are considerably less than the intravenous dose for basal anesthesia in dogs,
ACTION OF .AVERTIN ON URETER
713
were effective in reducing the spasm produced by adding potassium and barium salts to the perfusate and also in counteracting the effects of pilocarpine and arecoline given by vein. c. In situ ureter, the drug being administered rectally. Since avertin is usually administered per rectum, experiments were performed to determine whether on employing this route effects
FIG. 4.
REDUCTION OF SPASM IN THE INTACT URETER ON ADMINISTERING AvERTIN INTRA VENOUSLY
Ureter (dog; in situ; perfused with 0.9 per cent NaCl at 37°C.), showing spasm produced by potassium chloride and its reduction following the intravenous administration of avertin.
FIG. 5.
RELAXATION OF THE INTACT URETER ON ADMINISTERING AVERTIN RECTALLY
Ureter (dog; in situ; perfused with 0.9 per cent NaCl at 37°C.), showing depressant effects of avertin, administered by rectum.
could be produced on the ureter. The animals were prepared and the apparatus was arranged as described above, except that a rectal tube was inserted and secured in position by a ligature placed around the anal canal. The results were in brief the same as in the other perfusion experiments, but quantitatively
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much less pronounced. Doses below 200 mgm. per kilogram were not effective; 400 mgm. per kilogram introduced complications as a result of asphyxia, due doubtless to the prior use of amytal. However, it should be noted in this connection that the rectal dose for the dog is about 500 mgm. per kilogram as a basal anesthetic. Spasms produced by barium and potassium salts were reducible by 200 mgm. per kilogram and this quantity also antagonized effectively the actions of pilocarpine and arecoline. Doubtless, in an animal not already in the state of basal anaesthesia, smaller doses would be effective, since absorption from the alimentary canal would be more active. SUMMARY
1. The effects of avertin on the excised and intact ureter have been described. 2. Avertin very actively depresses the musculature of the excised ureter, producing decrease in tone and rhythmicity and increase in patency. 3. Avertin introduced into the in situ ureter, or administered intravenously or rectally, similarly reduces the tone and rhythmicity and increases the patency. 4. Ureteral spasm in the dog may be reduced by introducing avertin into the lumen of the organ or by intravenous and rectal administration. CONCLUSIONS
1. Avertin does not injure the ureter, the effects being immediately abolished on withdrawing the drug. 2. Avertin may be useful in urological work in reducing ureteral spasm and in other conditions where relaxation is desirable. REFERENCES (I) ANSCHULTz, W., SPECHT, K., AND TIEMAN, F.: Avertin narcosis in surgery, Ergebn. d. Chir. u. Orthop., 1930, xxiii, 406. (2) WADDELL, J. A.: The action of avertin on voluntary and non-voluntary muscle. Jour. Lab. and Olin. Med., 1932, xvii, 1104.. (3) MAGNIS, R.: Experiments on the surviving intestine. Arch. f. d. ges. Physiol., 1905, cviii, 1.
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(4) TRATTNER, H. R., WRIGHT, H. B., AND BARLOW, 0. W.: An experimental study of the action of sodium iodide on the excised and intact ureters of dogs. Jour. Urol., 1930, xxviii, 441. (5) BISKIN, M. S., AND DAN, lVI.: An automatic drop recorder. Proc. Soc. Exper. Biol. and Med.,, 1928, xxvi, 52. (6) WADDELL, J. A.: Absorption of avertin from the bladder. Unpublished data. Department of Pharmacology, University of Virginia, 1932.