CARBON DIOXIDE ADM!NIS?MTDN ' 'E‘C CALVfiS' Thu-is for flu: Dow's-c mi: M. S. MICHIGAN STATE UNIVERSITY Seref Nemihi Ersoz 1962 IRESIS LIBRARY ' Michigan State University ABSTRACT CARBON DIOXIDE ADMINISTRATION TO CALVES by Seref Nezihi Ersoz An investigational study has been made to determine the possible surgical anesthetic effect of carbon dioxide (002). A review of literature did not reveal satisfactory references in veterinary medicine with the exception of administration of 002 to slaughter hogs and poultry. In human medicine, CO2 has been used during child delivery, in the attempted treatment of alcoholism, muscular rigidity and psychoneurotic cases. The experimental equipment consisted of two 002 tanks, one oxygen (02) tank, a 002 gas analyzer (Ranarex), an electrocardiograph (Burdick EK III), and a kymograph (Watson Bird). Initially 100% 002 was administered until the calves fell into lateral recumbency, following this 65% 002 and 35% 02 was used to maintain anesthesia and the following data were collected before and during 002 anesthesia: Total leukocyte and erythrocyte counts, differential blood counts, packed cell volumes (hematocrit), hydrogen ion concentrations, Seref Nezihi Ersoz arterial blood pressures, electrocardiograms, pneumograms, and body temperatures. During anesthesia response to cor- neal and cutaneous pain stimuli were periodically observed. The principal findings were: 1) on the basis of experimental data no consistent significant changes were noted in the hematocrit values, total erythrocyte, leuko- cyte, and differential counts, 2) a consistent decrease of blood pH varying between 0.3 and 1.1, 3) an increase in blood pressure ranging from 13 to 112 mm. Hg., 4) an increase of heart rate between 30 and 120 beats per minute above normal, 5) significant changes in the conduction sys- tem of the heart during 002 administration were tachycardia, possible heart injury, and on one calf the Wenchebach phe- nomena were observed. The pertinent observations were: 1) blood stains in the nasal discharge on three calves, 2) deep respiratory movements, 3) defecation at the beginning or during 002 administration, 4) intermittent bellowing, 5) involuntary movements of the limbs of two calves, 6) a partial to com- plete absence of response to corneal and cutaneous pain stimuli. The results of the findings and observations are discussed with a summary followed by references and a bib- liography. CARBON DIOXIDE ADMINISTRATION TO CALVES BY Seref Nezihi Ersoz A THESIS Submitted to Michigan State University in partial fulfillment of the requirements for the degree of MASTER OF SCIENCE Department of Surgery and Medicine 1962 ACKNOWLEDGMENT I would like to express my sincere appreciation and gratitude to the following persons whose contributions made this thesis possible. To Dr. Gabel H. Conner, Professor, Graduate Advisor, Department of Surgery and Medicine, for his guidance, advice, and valuable suggestions in research and preparation of the thesis, and his patience throughout the period of the study. To Dr. Ralph D. Earner, Professor, Department of Surgery and Medicine, for his guidance, advice, encourage- ment in research and preparation of the thesis, and help in supplying me with the most important references. To Dr. Charles Coussens whose preliminary investi- gations enhanced these studies. To Miss Joyce Remsberg, Medical Technician, Depart- ment of Surgery and Medicine, for her assistance in the hematological procedures. The author is indebted to the Animal Welfare Insti— tute, 230 Park Avenue, New York 13, New York, whose finan- cial support made these research studies possible. ii Dedicated to My Instructors Prof. Dr. S. Nejat Yalki Prof. Dr. Yasar Altan iii TABLE OF CONTENTS CHAPTER PAGE I. INTRODUCTION. . . . . . . . . . . . . . . . . . I II. REVIEW OF THE LITERATURE. . . . . . . . . . . . 3 III. MATERIALS AND METHODS . . . . . . . . . . . . . 11 Experimental Animals. . . . . . . . . . . . . ll Experimental Equipment. . . . . . . . . . . . 11 Methods . . . . . . . . . . . . . . . . . . . 20 IV. RESULTS AND DISCUSSION. . . . . . . . . . . . . 27 Physiological Data. . . . . . . . . . . . . . 27 HematOIOgical Findings. . . . . . . . . . . . 3O Electrocardiographic Findings . . . . . . . . 31 Observations on Anesthetic Effect of CO . . . 33 V. SUMMARY . . . . . . . . . . . . . . . . . . . . 63 REFERENCES. . . . . . . . . . . . . . . . . . . . . . . 65 BIBLIOGWHY. O O O O O O O O O O O O O O O O O O O O O 69 iv FIGURE 10. ll. 12. l3. l4. l5. 16. The CO2 Apparatus. . . . . . . . . . The 002 Gas Analyzer (Ranarex) . . . The Beginning of 002 Administration. Calf Under CO 2 LIST or FIGURES Anesthesia. Eleotrocardiograph (Burdick EK III). . Kymograph (Watson-Bird Variable Speed 70-142). Pneumogram, Pneumogram, Pneumogram, Pneumogram, Pneumogram, Electrocardiogram, Calf Calf Calf Calf Calf No. 1 No. 2 No. 3 No. 4 No. 5 Calf Electrocardiogram, Calf Electrocardiogram, Calf Electrocardiogram, Electrocardiogram, Calf Calf PAGE 13 15 l7 19 24 44 46 4e 50 52 54 56 58 6O 62 TABLE \0 CD \1 ON Ul .p \N I\) |—‘ O H O '11. 12. 13. 14. 15. Hematological Physiological Hematological Physiological HematolOgical Physiological Hematological Physiological Hematological Physiological Electrocardiographic Findings, Electrocardiographic Findings, Electrocardiographic Electrocardiographic Findings, Electrocardiographic LIST OF TABLES Findings, Calf Data, Calf No. Findings, Calf Data, Calf No. Findings, Calf Data, Calf No. Findings, Calf Data, Calf No. Findings, Calf Data, Calf NO. vi Findings, Findings, No. l. l. . . No. 2. 2. . . No. 3. 3. . . No. 4. 4. . . No. 5. 5. . . Calf Calf Calf Calf Calf \n-P-UJNH PAGE 35 35 36 36 37 37 38 3a 39 39 4o 40 41 41 42 CHAPTER I INTRODUCTION Carbon dioxide (002)--its production in the body, effects upon body systems, and elimination from the body-- has always been of concern to anesthesiologists, as well as physiologists. In human medicine, 002 has been used for a number of different conditions: child delivery (MacRae, 1951), treatment of neuroses and alcoholism (LaVerne, 1953). treat- ment of stutterers (Smith, 1953), and muscular rigidity i (Fay, 1953). The most extensive studies with 002 have been made by Meduna (1953 and 1958) who has written a text book entitled "Carbon Dioxide Therapy--A Neurophysiological Treat- ment of Nervous Disorders." Within recent years, an increas- ing interest has been taken in the anesthetic properties of CO2 in veterinary medicine (Swem, 1952; Murphy, 1953; Blomquist, 1957; Wernberg, 1957; Kitchell, et al., 1960), as well as its usage in meat packing plants to produce surg- ical anesthetic effects before exsanguination. The "Humane Slaughter Law" passed by the 85th U.S. Congress as a public law in 1958 required slaughter houses to dispatch with do- mestic animals in a humane manner. Carbon dioxide has been approved for a pre-surgical anesthetic for swine, sheep, 2 and calves (Title 9, Code of Federal Regulations, 1959).* This study was conducted as a result of interest aroused by the work of previous investigators and the ap- parent lack of scientific data regarding the effect of 002 on calves. * . Regulations governing the meat inspection of the United States Department of Agriculture, United States Gov- ernment Printing Office, Washington, 1960. CHAPTER II REVIEW OF THE LITERATURE In 1827, Hickman, cited by McQuacker (1929), first demonstrated that animals exposed to an atmosphere contain- ing a high concentration of 002 became anesthetized. He performed experimental surgery on a dog, puppy, mouse and rabbit using 002 as the anesthesia inducing agent. In 1878 Bert, cited by Loewy (1923), recommended the gas for anesthesia and reported that 002 concentrations of 25 to 30% in the inspired air would effect paralysis of the central nervous system with cessation of corneal and other reflexes. Zuntz and Loewi (1897) and Kropeit (1898) observed that in rabbits there occurred a short ex- citation stage, but no such stage has been observed in cold blooded animals (Winterstein, 1911). Albitzki (1912) stated in experiments with rabbits that if the concentration of CO2 in inspired air was lower than 25 to 30% complete anes- thesia was not attained. But following recovery from anes- thesia they showed disturbances of balance, as well as a failure in the ability to keep upright and to change their positions. Prausnitz (1928) showed that 25 to 30% 002 was re— quired in the inspired air to abolish corneal and light reflexes in experiments on dogs, cats, rabbits, guinea pigs, rats, and mice. The excitation stage was slight, and awak- ening occurred quickly even though the anesthesia lasted for one hour. The animals tolerated a concentration of 50% 00 resulting in deep anesthesia for 60 to 90 minutes. 2 Unfortunately, after this period of anesthesia the animals showed convulsions and died of pulmonary edema. Leaks and Waters (1928), in experiments on dogs, found that 30 to 40% C02 with oxygen (02) will anesthetize the dogs in about one minute usually without struggling. The pulse rate was usually reduced as was the respiratory rate, but the blood pressure rose rapidly. The depth of respiration was greatly increased, but breathing did not seem to be labored. Upon opening the abdominal cavity, the intestines seemed relaxed and the muscles were usually flaccid. Dogs have been maintained under fairly satisfac- tory anesthesia for two hours. Respiratory movements became more regular and the depth of anesthesia deeper the longer the animal received the same mixture. According to Leaks and Waters (1928), the lower percentages of CO2 did not i seem to anesthetize as well. Concentrations higher than 40% tended to depress respiration. Cardiac function and anoxemia became a factor with concentrations above 80%. With 100% 002, the blood pressure became lower and the heart 'and respiration stopped at about the same time. In another experiment one year later, these investigators demonstrated anesthesia with 30-40% 002 in a variety of animals obtain- ing comparable results. They also pointed out that it was theoretically significant that high concentrations of a metabolic waste product (002) would depress the central nervous system enough to produce anesthesia. These inves- tigators concluded that the anesthetic effect, in the case of a mixture of 30 to 40% CO2 with 02, was not due to as- phyxia. The theory being that the increased 002 tension in the blood prevented the escape of CO2 from the cells, and that the gradual accumulation of this metabolic product may progressively depress cellular function. In 1926, Losvenhart, Lorenz and Waters first intro- duced 002 inhalation therapy into psychiatry. They theorized that a disordered function of the brain may be corrected by physiological and biochemical means. The report of this study was published in 1929 but their technique has since been abandoned. Hsttwsr (1938) examined the relation of excitability of frog sciatic nerve to 002 tension of the external medium by systemic, graphic action potential threshold determina- tions and found a reduced irritability of isolated nerves after exposure to 002. Barbour and Seevers (1941a) conducted a series of experiments to determine the influence of cold on the nar- cotic action of 002 on rats, rabbits and dogs in a chamber at 5 C. They reported the minimal narcotic concentrations at this temperature as follows: rat, 11%; rabbit, 17%; and dog, 14%. The induction time varied from 3 to 24 hours. These concentrations of 002 did not induce narcosis at 25 C. The same authors (1941b) showed in albino rat eXperi- ments that the maximum tolerated concentration of CO2 for the first time was between 15 to 20%. At 20% an occasional animal survived. Atmospheres containing between 25 to 30% 002 were uniformly lethal. Survival time varied from 0.5 to 36 hours, depending on the concentration of 002. At 50% 002, no animal survived longer than five hours. Death seemed to be due primarily to pulmonary injury rather than narcosis. The edema and hemorrhage noted in the lungs were also seen on all visible mucous membranes. Continuing the experiments, Barbour and Seevers (1943a) also pointed out that during the exposure of albino rats to 11% 002 for 30 days, the plasma pH fell to a level as low as 7.09 in the first half hour and remained below 7.15 during the remainder of the period. A marked reticulocytosis occurred in the rat during prolonged exposure to 11% 002, but no correspond- ing increase in the number of leukocytes or erythrocytes was observed. In another experiment they (1943b) concluded that the time required to produce the state of narcosis varied with the concentration of the 002, the species, and previous condition of the animal. Gsllhorn and Yesinick (1941) produced convulsions in anesthetized cats by the injection of metrazol, picrotoxin, corymyrtin, camphor, strychnine and other drugs. In the non-denervatsd cats, 002 exerted a greater anti- convulsant effect whereas in those that were denervated, anoxia was produced. In experiments on dogs, Draper and Vhitehead (1944), and Parry, et a1. (1949) succeeded in arresting respiration for 45 minutes by administering a barbiturate and CO2 anesthesia and permitting the dog to rebreathe expired air. By the end of the experiments, up to 50% CO2 was found in the alveolar air. In other experi- ments the acidity of the venous blood was determined after inspiration of different concentrations of 002. Inspira- tion of 40% 002 was found to effect an average fall of blood pH from 7.42 to 6.79. The only alteration in the electro- cardiogram was an increase in the amplitude of the T wave, which returned to normal following restoration of the normal blood pH. Paulsen (1949) described an apparatus with an ar- rangement for continuous production of anesthetic gas mix- tures consisting of 002 and ether. In a series of experi- ments on rabbits Paulsen (1952) used 30 to 35% CO2 and ob- served that 002 produced a marked rise of the CO2 concen- tration in the blood of these animals, and at the same time a fall in the blood pH from about 7.5 to 6.5. The corneal reflex also ceased at this concentration. In human medicine, Meduna (1950, 1953) conducted a series of experiments using 30% 002 and 70% 02 on psychoneurotic individuals. This line of therapy was given to more than 40,000 patients. No complications were re- ported other than tongue biting (1 case) and 3 or 4 spon- taneous urination (3 or 4 cases). macRae (1951) found that inhalation of suitable mixtures of CO2 and 02 produced coma which was both safe and beneficial in delivery at child birth. He stated that as an anesthetic for delivery "002 coma" is more satisfactory both for the mother and the baby than other anesthetics. However, some other anesthetic is needed for surgical repair because the "002 coma" is brief. LaVerne (1953), using higher concentrations of 002 such as 70% 002 and 30% 02, demonstrated the safety of this mixture on 4,000 patients. It should be taken into consider- ation that the CO2 treatment has some contraindications, such as: previous history of coronary attack, organic heart disease, advanced arterio-sclerosis, hypertension and em- physema. Within recent years the veterinary profession has shown an increased interest in the anesthetic properties of 002. Swem (1952) and Murphy (1953) at Geo. A. Hormel & Co. employed a mixture of 002 to immobilize the slaughter hogs for ease in shackling and sticking. By using this method, both the carcass yield and by-products were improved. Johnson (1960) believes that the ability of 002 to produce anesthesia is by loss of function of the con- soiousness center of the brain because of the associated anesthetic prOperties of the gas and the markedly reduced supply of 02 to the brain. This is accomplished before vital heart action ceases. It is known that an isolated nerve in high CO2 tension milieu demands greater irritation to react, that is, to send out impulses. In addition, it is known that the speed of the impulse along the nerve is reduced. 002 also has the effect of making the transfer of impulses from one nerve cell to another more difficult because the connections between the nerve cells (synapses) are partly blocked by 002. Meduna (1953) is in agreement with this concept. 0n the other hand, Murphy (1953), work- ing with hogs, contended that the CO2 unites with the hemo— globin of the blood and thereby causes unconsciousness. The compound rapidly separates from the hemoglobin and is discharged by exhalation as the hog continues to breathe fresh air before and after sticking. As a result of the swine studies, Kotula, Drewmak and Davis (1956) used 33 to 36% CO2 in stunning chickens, and Blomquist (1957) and Wernberg (1957, 1962) further in- vestigated reactions of hogs to CO2 stunning during an ex- periment carried out by the Danish Meat Research Institute in which 70% CO2 was used. Pistraszek (1958) and Bartels (1962) also outlined the effect of 002 in stunning hogs. In the United States the Humane Slaughter Law was passed by the 85th Congress as a Public Law 85-765 and be- came effective August 27th, 1958. The purpose of this law 10 was to require the use of humane methods in the slaughter of livestock. The animals must be exposed to the CO2 gas in such a way as to produce surgical anesthesia before they are shackled, hoisted, thrown, cast or cut. Carbon dioxide is a relatively cheap and non-flammable gas, commercially available in large quantities and when used with properly designed units, offers no hazard to employees. These fac- tors havs stimulated research to determine its value as a surgical anesthetic for use on animals. CHAPTER III MATERIALS AND METHODS A--Experimental Animals All experiments in this study were conducted on five calves as described below: Tag No. nggg Agg _gx Weight 1 Holstein 3 months M 260 lbs. 2 Holstein 4 months M 335 lbs. 3 Holstein 4 months M 350 lbs. 4 Holstein 3 months F 272 lbs. 5 Jersey 3 months M 360 lbs. The above animals were housed in the Veterinary Clinic of the College of Veterinary Medicine, Michigan State University, and were fed good quality alfalfa hay. B--Experimental Equipment ‘The 002 apparatus used for induction of anesthesia consisted of: 1. Two 002 tanks each equipped with two gauges (cylinder contents and outflow regulator) and a flow-meter (Figure l). The tanks were connected with a COpper tubing in such a manner that the gas from one or both of them passed through an Automatic CO2 Heater (Figure 1) before going 11 Figure 1. 12 The 002 Apparatus A. B. C. D. E. C0 tanks 2 Automatic C02 Heater Cylinder Contents Gauge Outflow regulator Flow-meter l3 14 Figure 2. The 002 Gas Analyzer, Ranarex \ \.| “JUMP- \N 16 Figure 3. The Beginning of CO2 Administration l7 18 Figure 4. Calf under CO2 Anesthesia 19 20 into the inhalation mask.* This unit was mounted on a two- wheeled truck. 2. One 02 tank mounted on a truck and likewise equipped with two gauges (as above) and a flow-meter (Fig- ure 1). 3. An equine protective field mask, M4, which will be called an inhalation mask. 4. A 002 gas analyzer, Ranarex,** which gave a continuous analysis of the concentration of 002 being in- haled by the calf. The gas sampling tube was connected to the tube containing the 002-02 mixture 5 ft. from the mask. At this point, the gases had flowed 25 ft. from their source and should have been thoroughly mixed. C--Methods The initial stages of administering the anesthesia consisted of giving the calves 100% 002 flowing at a rate of 150 lts. per hour. Within 25 to 37 seconds after the calves went down, a sufficient flow of 02 was supplied to result in a C02 concentration of 65%. Induction of anes- thesia was also attempted with 65 and 80% CO2 in combination with either 02 or air. *- Chemical Supply Officer, New Cumberland General Depot, New Cumberland, Pa. *- *Manua1 R-466, Ranarex Instrument Division, The Permutit Company, 50 West 44th Street, New York 36, N.Y., 1957. 21 Electrocardiographic tracings were made on a Bur- dick EK III* using a paper speed of 25 mm./second. Only limb leads were used and the recordings were taken before and during CO2 administration. Blood pressures were taken directly from the caro- tid artery which had previously been permanently implanted beneath the skin by surgical procedures. For pressure de- terminations a 16 gauge 1% inch hypodermic needle was in- serted into the carotid artery with the point directed to- wards the heart. This needle was connected to a mercury manometer and the blood pressure was recorded in milli- meters of mercury. A series of blood pressure determina- tions were taken prior to any experimentation in order to establish normal ranges. Blood pressures were also taken during 002 administration. Frequency and depth of respiration were recorded immediately prior to and during 002 administration using a Watson-Bird Variable Speed Kymograph 70-142** (Figure 6). Kymograph paper speed used for this work was 25 mm/sec. The hydrogen ion concentration of arterial blood . U *** . was determined on a Beckman pH meter, model H—2 uSlng * Operating Manual EK III-858 for the Burdick EK III Dual—Speed Electrocardiograph, Aug., 1958. l"!- Tools for Biophysics, Catalog No. 5—60, Phipps and Birds, Inc., Silver Spring, Maryland. ***Beckman--Bulletin 190-3, Feb., 1948, National Technical Labs, South Pasadena, California. 22 a special blood electrode assembly (X 800-5) with a flow chamber. Total erythrocyte and leukocyte counts were made on the improved Neubauer Sharpline Hemocytometer.* The blood smears for differential counts were stained with Wright stain. The packed cell volumes were read from micro- capillary tubes. Rectal temperatures were taken before and during CO2 administration. The corneal reflexes were tested by touching the cornea with a moistened finger and cutaneous pain reflexes by pricking the skin between the toes with a needle or by pinching with a forceps. Each calf was subjected to CO2 administration three times with intervening intervals varying from 2 to 8 days. During 002 administration, a claw amputation was attempted on two calves and on one calf a 3 inch skin in- cision was made on the lateral surface of the foreleg (over the radius) and sutured. -I- . Sharpline Hemocytometer, Improved Neubauer Ruling, Chicago Apparatus 00., Chicago, Ill. 23 Figure 5. Electrocardiograph Burdick EK III 25 0-142 ” raph 7 " d kymog Variable apes —Bird a Watson 6. Figure CHAPTER IV RESULTS AND DISCUSSION The calves used in this study were considered to be normal and it is believed that their health status did not interfere with the investigational results herein re— ported. Physiological data Tables 2, 4, 6, 8, and 10 show the experimental physiological data on each of the five calves. Before 002 administration, the rectal temperatures varied between 101.0 and 103.0 F. The temperature of each calf taken 5 minutes before the end of CO2 administration did not reveal any appreciable elevation. However, the temperature of calf No. 2 was increased during the first (102.6 F. to 104.8 F.) and second experiments (102.4 F. to 103.8 F.). During 7 of the 15, there was an increase of temperature ranging from 0.1 F. to 0.7 F. In the remaining experiments, there was a decrease of temperature varying from 0.2 F. to 1.2 F. Because of the variations of temperatures observed and the small number of animals used, it is difficult to make a general statement regarding the effect of 002. Superficial observation of the pneumograms* (Figs. *Due to the similarity of the pneumograms only one is presented for each calf (recorded during first experiment). 27 28 7, 8, 9, 10, 11) might lead one to believe that the respira— tory rates during 002 anesthesia were increased. But actu- ally, the rates were often the same as before 002 adminis- tration. However, there was a marked increase in the depth of the respiratory movement. Best and Taylor (1961) state that an excess of CO2 in circulating blood depresses the respiratory center resulting in a decrease of respiratory rate. Before the 002 administration, the respiratory fre- quency varied between 20 and 60 per minute; during CO2 ad- ministration it varied between 36 and 60. The most signif— icant increase in respiratory rate was seen on calf No. 4 (Table 8, Fig. 10) where there was an increase from 20 to 44 during second experiment and from 28 to 40 during the third experiment. Also Calf No. 5 showed an increase from 24 to 48 during the second experiment. The most significant findings of the physiological data were those pertaining to the heart rate, blood pres- sure, and blood pH during 002 administration. The heart rate on each animal increased during 002 administration. This increase varied from 40 to 120 beats per minute. The most marked increase was on Calf No. 2 (Table 4 and Fig. 12) during the first experiment (from 80 to 200). The in- creased heart rate is mainly due to the effect of CO2 upon cardiac and vasomotor centers, and directly on the cardio- vascular musculature (Best and Taylor, 1961). The pH of venous blood decreased on each animal 29 during 002 administration. This was apparently the result of increased amounts of carbonic acid in the blood. This fall varied between 0.3 and 1.1. Pertaining to this decrease, it is noteworthy to state that the junctional tissues are particularly sensitive to high tensions of CO2 and auriculo- ventricular conduction becomes markedly depressed when the 002 excess is sufficient to cause a fall in pH of the fluids bathing the cardiac muscle fibers. At a pH of around 7.0 complete heart block occurs in human beings (Best and Taylor, 1961). The excessive fall in blood pH is probably a dis- advantage to the use of 002 in calves as a surgical anes- thetic. However, there did not appear to be any serious toxic effect during the experiments. The blood pressures also increased during experiments on each animal. According to Best and Taylor (1961) this increase is a result of high 002 tension in venous blood coming into the heart, thus enhancing the extensibility of the cardiac muscle fiber during diastole, and in consequence, exerts a more favorable effect upon the filling of the heart. The cardiac output is therefore increased and the hyper- tonicity of the vasoconstrictor center results in a high blood pressure. The highest increase was from 170 to 282 mm. Hg. on Calf No. 2 (Table 4) during the third experiment, and the smallest increase was on Calf No. 5 (Table 10) during the third experiment (from 157 to 170 mm. Hg.). In the 30 remaining experiments, the blood pressure increase ranged from 21 to 84 mm. Hg. above the recordings taken prior to CO2 administration. The corneal reflex was present on each animal with the exception of partial abolishment on Calf No. 1 during the first and second experiments. The cutaneous pain re- flexes were abolished on Calf No. 1 during first and second experiments. During the balance of the experiments, cutane- ous pain reflexes were either partially abolished (forelegs or hindlegs), or were present. HematolOgical findings The results of the hematological findings are pre- sented in Tables 1, 3, 5, 7, and 9. The hematocrit (Hot.) values varied between 27 and 36.5% in all calves before 002 administration. During CO2 administration in the first trials on each animal there was an increase in RBC counts varying from 290,000 to 3,840,000/mm3. As an example, in Calf No. 5 (Table 9), the Hot. value remained constant dur— ing the experiment but the RBC count increased from 8,190,000 to 10,380,000. This picture may indicate that a new supply of RBC's entered the blood stream but that they were appar- ently smaller in size. A similar observation was made on Calf No. 3 (Table 5) which also shows an increase of RBC's from 7,020,000 to 10,860,000 with only a slight increase in Hot. (27.5 to 29%). In the remainder of the experiments there was no consistent increase or decrease in RBC counts. 31 The WBC counts before CO2 administration varied between 5,700 and 13,200/mm3. During 002 administration, the counts ranged from 5.760 to 13.000/mm3. The foregoing WBC counts are generally considered to be the normal values. Inconsistent variations in WBC counts may be noted before and during 002 administration. Electrocardiographic findings The electrocardiograms* (Figs. 11, 12, l3, l4, and 15) were taken with the calves in the standing position be- fore starting the experiments and in a recumbent position during CO2 administration. All recordings were from Lead 1. In this study with calves in the standing position P waves were positive on Calf Nos. 1, 2, 3, and 4 (Figs. 11, 12, 13, and 14) and diphasic on Calf No. 5 (Fig. 15). As is known, the P wave is variable in dairy calves, at times diphasic in Lead I and sometimes inverted (Ardsel, Kreger, and Bogart, 1959), or positive (Platner, Kibler, and Brody, 1948). The PR interval varied between 0.132 and 0.182 seconds in all calves. These data are similar to the results obtained by Platner, Kibler, and Brody (1948) who reported intervals of 0.133 to 0.167 seconds. During CO2 administration with the calves in left lateral recumbency there was a significant change in the P *- . Due to the similarity of electrocardiographic re- cordings only one is presented for each calf (recorded dur- ing first experiment). 32 wave and the PR interval in all calves as follows: Calf Nos. 1 and 2 (Figs. 11 and 12, Tables 13 and 14). In Calf No. l the T wave was superimposed on the P wave whereas the P wave almost disappeared on Calf No. 2. The PR inter- val showed a variation between 0.14 and 0.16 seconds and widened on Calf No. 1. This picture together with the re- sults on Calf No. 2 may indicate a possible current injury of heart muscle. Both recordings present evidence of tachy— cardia. Calf No.43 (Fig. 13 and Table 15). The P wave for this calf is superimposed by the T wave, and there is an increase (up to 0.20 sec.) in the PR interval. This increase in the PR interval is sufficient to cause one heat to drop out. This picture reveals a special type of partial AV block, sometimes referred to in man as the Wenchebach phenomena. It occurs when the PR interval increases in length with each successive cardiac cycle until the impulse, originat- ing in the SA node, is completely blocked at the AV node and fails to initiate a QRS complex (Burch and Winsor, 1960). Calf Nos. 4 and 5 (Figs. 14 and 15, Tables 16 and 17). The P wave on Calf No. 4 was absent. On Calf No. 5 a pre-anes- thetic diphasic P wave changed to a monophasic wave with a positive deflection during 002 administration. Also, the PR interval increased on Calf No. 5 (0.20 to 0.22 sec.). 33 Before CO2 administration the QRS complex was di- phasic on Calf Nos. 1, 2, and 4 (Figs. 11, 12, and 14; Tables 13, 14, and 16), and positive on Calf Nos. 3 and 5 (Figs. 13 and 15; Tables 15 and 17). Arsel III, Krueger and Bogart (1959) reported similar findings. The QRS interval showed a variation between 0.04 and 0.072 second which is similar to the results reported by Platner, Kibler and Brody (1948). During CO2 administration, the QRS complex showed a negative deflection and amplitudes were increased on all calves. During the same time the QRS interval increased during each experiment ranging from 0.01 to 0.028 seconds with one exception--Calf No. 5--during the second experi- ment in which there was no change in the QRS interval. Prior to 002 anesthesia the T wave was positive on Calf No. 1 (Fig. 11), and negative on Calf Nos. 2, 4, and 5 (Figs. 12, 14, and 15), and there was no T wave on Calf No. 3 (Fig. 13). During the same time the QT interval showed a variation of 0.28 to 0.36 seconds which resembled the results obtained by Platner and his colleagues (1948). Observations on anesthetic effect of 002 As was pointed out in Chapter II, 100% 002 has been used to put calves down in lateral recumbency. Near the beginning of each C02 administration most calves coughed once or twice and became recumbent within 25 to 37 seconds. There was an absence of struggling in Calves No. 1, 2, 3. 34 and 4 with the following exceptions: Calves No. 2 and 4 during the second experiment struggled intermittently. Calf No. 5 struggled intermittently during each of the three experiments. During this series of 15 CO2 administrations the calves occasionally emitted a slight bellowing, passed feces, and showed some involuntary movements of limbs. At the end of some of the experiments there were blood stains mixed with nasal discharge in the inhalation mask. In an evaluation of the anesthetic effect of 65% CO2 and 35% 02 for surgical procedures, a 12 cm. skin in- cision was made on the forelimb over the radius (lateral surface) of Calf No. 4. The incision was made and sutur— ing was accomplished without the calf showing any evidence of pain. Two attempts were made to perform a claw amputation operation on two calves while under the influence of 65% 002 and 35% 02. There was not sufficient desensitization of the interdigital skin to permit this surgery. 35 Table l. Hematological findings, Calf No. 1 Differential Count* Experiment Hct. RB WBC Neut. Lymph. Mono. Eosin. 36 10 103 96 % a Before 27.0 6.52 6.55 25 67 3 5 let ' During** 28.0 6.81 7.00 4 89 4 3 Before 29.0 6.47 7.15 8 88 4 0 2nd During 34.5 8.56 8.30 6 91 l 2 Before 35.0 6.65 5.70 18 79 2 1 3rd During 36.5 6.65 5.76 21 74 2 3 Table 2. Physiological data, Calf No. 1 *** Experiment Duration Temp. Resp. Heart Blood Reflexes /min. F. rate rate pres.pH Cornea Skin g_/mint_/min. mm.Hg. Before 102.4 52 100 166 7.1 lst During** 17 102.5 38 160 200 6.6 PA A Before 102.4 36 100 166 6.9 2nd During 20 102.6 40 150 210 6.6 PA A Before 101.1 52 80 166 7.4 3rd During 45 101.8 36 120 250 6.4 P PA *Basophils were not observed on the blood smears. **Taken 5 minutes before the end of CO2 administration. ***Duration of 002 administration. Legend: A-—Abolished; PA—-Partially abolished; P-—Present. 36 Table 3. Hematological findings, Calf No. 2 Differential Count* Experiment Hot. RB WBC Neut. Lymph. IMono. Eosin. 9% 10 103 % % Before 29.0 5.78 8.35 25 72 2 1 let During** 30.0 7.11 12.55 36 60 4 0 Before 27.0 6.65 6.60 20 78 1 1 2nd During 27.0 6.35 10.45 18 82 0 0 Before 29.0 6.45 6.30 26 70 4 0 3rd During 31.0 7.10 6.73 28 68 4 0 Table 4. Physiological data, Calf No. 2 , *** Experiment Duration Temp. Resp. Heart Blood Reflexes /min. F. rate rate pres. pH Cornea Skin _g/min._/min. mm. Hg. Before 102.6 42 80 170 7.5 lst During 21 104.8 44 200 212 6.6 PA Before 102.4 60 80 170 6.9 2nd During 30 103.8 48 120 194 6.5 P Before 101.6 52 80 170 7.3 3rd During 27 101.2 56 110 282 5.6 PA *Bas0phils were not observed on the blood smears. administration. **Taken 5 minutes before the end of 00 ***Duration of 002 administration. Legend: 2 P-—Present; PA--Partially abolished. 37 Table 5. Hematological findings, Calf No. 3 Differential Count* Experiment Hot. RB WBC Neut. Lymph. Mono. Eosin. 0% 10 103 9% 56 3?» Before 27.5 7.02 13.20 42 53 l 4 1st During** 29.0 10.86 10.10 49 41 l 9 Before 28.0 6.16 10.60 19 77 3 1 2nd During 27.0 5.26 13.00 12 84 2 2 Before 32.0 7.06 11.25 44 55 l 0 3rd During 32.0 6.40 11.45 40 55 3 2 Table 6. Physiological data, Calf No. 3 . . , inm- Experiment Duration Temp. Resp. Heart Blood Reflexes /min. F. rate rate pres.pH Cornea Skin p/min._[min. mm. Hg. Before 102.4 44 60 178 7.2 lst During 30 102.2 44 90 223 6.6 PA Before 102.6 32 60 178 7.1 2nd During 22 103.0 52 100 212 6.5 PR Before 102.0 40 60 178 7.3 3rd . During 27 102.6 38 120 250 6.2 PR *Bas0phils were not observed on the blood smears. **Taken 5 minutes before the end of CO2 administration. ***Duration of 002 administration. Legend: P—-Pressnt; PA--Partially abolished; PR--Partially abolished rear legs. 38 Table 7. Hematological findings, Calf No. 4 Differential Count* Experiment Hot. RBC 'WBC Neut. Lymph. Mono. Eosin. 3’0 106 103 a 36 J5 3% Before 35.0 8.19 10.80 8 91 0 l IStDuring** 36.5 10.38 7.90 11 87 1 1 Before 28.0 7.70 9.10 24 76 0 0 2ndDuring 35.0 13.35 8.14 24 76 0 0 Before 32.0 8.08 11.45 16 82 2 0 3rdDuring 33.0 8.09 11.25 25 72 2 0 Table 8. Physiological data, Calf No. 4 Experiment Duration***Temp. Resp. Heart Blood Reflexes /min. F. rate rate pres. pH Cornea Skifi /min./min. mm. Hg. Before, 102.4 36 90 174 7.7 1st During 42 101.2 52 140 224 7.0 P PA Before 102.6 24 96 174 7.5 2nd During 25 102.4 48 148 210 6.7 P P Before 101.8 28 90 174 7.3 3rd During 18 101.2 32 140 220 7.0 P PF *Basophils were not observed on the blood smears. **Taken 5 minutes before the end of CO2 administration. ***Duration of 002 administration. Legend: P--Pressnt; PA--Partially abolished; PF--Partially abolished forelegs. 39 Table 9. Hematological findings, Calf No. 5 Differential Count* Experiment Hct. RBC WBC Neut. Lymph. Mono. Eosin. ~. 106 103 76 , 54 Before 36.5 8.19 10.80 8 91 0 1 1st During** 36.5 10.38 7.90 10 89 l 0 Before 30.5 9.49 10.20 13 84 2 1 2nd During 31.0 9.77 9.40 10 86 3 1 Before 30.0 8.25 10.50 18 78 2 2 3rd During 32.0 10.30 8.45 23 75 2 0 Table 10. Physiological data, Calf No. 5 *** Experiment Duration Temp. Resp. Heart Blood Reflexes /min. F. rate rate pres. pH Cornea Skin p/min.,/min. mm. Hg. Before 102.0 52 70 157 7.6 1st During 40 101.0 60 140 210 7.1 P P Before 101.8 20 100 157 7.6 2nd During 15 101.6 44 144 178 6.9 P P Before 101.6 28 70 157 7.4 3rd During 23 101.4 40 130 170 6.7 P P *Basophils were not observed on the blood smears. **Taken 5 minutes before the end of 00 ***Duration of CO2 administration. Legend: P——Present. 2 administration. 40 Table 11. Electrocardiographic findings, Calf No. 1 PR ens of Experiment interval interval interval sec. sec. sec. Before 0.14 0.072 0.28 lst During 0.16 0.080 0.22 Before 0.15 0.048 0.28 2nd During 0.16 0.080 0.23 Before 0.16 0.072 0.32 3rd During 0.16 0.080 0.24 Table 12. Electrocardiographic findings, Calf No. 2 PR QRS QT Experiment interval interval interval sec. sec. sec. Before 0.16 0.040 0.35 lst During No P wave 0.052 0.20 Before 0.14 0.044 0.32 2nd During No P wave 0.072 0.24 Before 0.16 0.040 0.30 3rd During No P wave 0.064 0.20 41 Table 13. Electrocardiographic findings, Calf No. 3 PR QRS QT Experiment interval interval interval sec. sec. sec. Before 0.180 0.040 No T wave 1st During 0.13-0.20 0.080 0.28 Before 0.182 0.040 No T wave 2nd During 0.13-0.20 0.070 0.28 Before 0.180 0.040 No T wave 3rd During No P wave 0.078 0.30 Table 14. Electrocardiographic findings, Calf No. 4 PR QRS QT Experiment interval interval interval sec. sec. sec. Before 0.160 0.060 0.36 lst During No P wave 0.070 0.28 Before 0.160 0.048 0.32 2nd During Not recorded* Before 0.168 0.048 0.35 3rd During No P wave 0.080 0.28 *Because of the movements of calf. 42 Table 15. Electrocardiographic findings, Calf No. 5 PR QRS QT Experiment interval interval interval sec. sec. sec. Before 0.140 0.080 0.328 lst During 0.220 0.128 0.360 Before 0.132 0.080 0.320 2nd During 0.200 0.080 0.360 Before 0.128 0.080 0.340 3rd During Not recorded* *Because of the movement of calf. Figure 7. 43 Pneumogram, Calf No. 1 A. Before CO2 administration B. During CO2 administration 0. Time interval 1 sec. Figure 8. Pneumogram, Calf No. 2 A. Before 002 administration B. During CO2 administration 0. Time interval 1 sec. 47 Figure 9. Pneumogram, Calf No. 3 A. Before CO2 administration B. During CO2 administration 0. Time interval 1 sec. lllLlllllnllllllJlnllllLJ lllll Figure 10. 49 Pneumogram, Calf No. 4 A. Before 002 administration B. During CO2 administration 0. Time interval 1 sec. 50 Figure 11. 51 Pneumogram, Calf No. 5 A. Before 00 administration 2 B. During CO2 administration 0. Time interval 1 sec. IIIIIIIIIIIIIIIIIIIIIIIIIIIIIII 53 Figure 12. Electrocardiogram, Calf No. l A. Before CO2 administration B. During 002 administration (Five large squares represent a time interval of 1 sec.) 54 55 Figure 13. Electrocardiogram, Calf No. 2 A. Before 002 administration B. During CO2 administration (Five large squares represent a time interval of 1 sec.) 56 57 Figure 14. Electrocardiogram, Calf No. 3 A. Before CO2 administration B. During CO2 administration (Five large squares represent a time interval of 1 sec.) 58 59 Figure 15. Electrocardiogram, Calf No. 4 A. Before CO2 administration B. During CO2 administration (Five large squares represent a time interval of 1 sec.) 6O 61 Figure 16. Electrocardiogram, Calf No. 5 A. Before CO2 administration B. During CO2 administration (Five large squares represent a time interval of 1 sec.) 62 CHAPTER V SUMMARY During the administration of 65% C02 and 35% O2 to calves, there was a consistent decrease of blood pH varying be- tween 0.3 and 1.1. During all 002 administrations, there was a definite increase in blood pressure ranging from 13 to 112 mm. Hg. The heart rats showed an increase between 30 and 120 beats per minute. The hematological studies conducted during 002 admin- istration revealed only minor fluctuations from the values determined before the administration of CO2. The most significant fluctuation was noted during the 'first 002 experiments on each calf where there was an increase in the RBC count varying from 290,000 to 3,800,000 cells per cubic millimeter. As evidenced by the electrocardiographic studies, there appeared to be a significant change in the conduction system of the heart during 002 administration. Tachy- cardia, possible heart injury, and on one calf (NO- 3; Fig. 13), the Wenchebach phenomena (partial AV block), were observed on the electrocardiograms. 63 10. 64 During CO2 anesthesia, the depth of respiratory move- ments increased but the rate did not appreciably change. The corneal reflex was present during 002 anesthesia. Cutaneous pain reflexes were sometimes absent, at other times present, especially in the interdigital areas. Rectal temperatures did not reveal any consistent de- crease or increase during 002 administration. The nasal discharges of 3 calves contained small amounts of blood noticed upon completion of CO2 administration. Calves would not fall into lateral recumbency when CO2 was administered in concentrations of 65 or 80%, in combination with either 02 or air. REFERENCES Albitzki, P.: Uber die Ruckwirkung Resp. "Nachwirkung" der 002 und Uber die Biologiske Bedentung der im Korper Gewohnlich Vorhandenen Kohlensaure. Arch. f. d. ges. 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