Comparison of Cardiopulmonary Parameters in Awake and Anesthetized Chickens THOMAS E .
NIGHTINGALE
USDA Poultry Research Laboratory, R.D. 2, Box 600, Georgetown, Delaware 19947 (Received for publication May 5, 1976)
POULTRY SCIENCE 56: 147-153, 1977
inherent in handling, restraint, and intravenous injection of birds, Gandal (1956) reported the advantage of intramuscular administration of a solution containing chloral hydrate, magnesium sulfate, and pentobarbital. This mixture has become accepted for many types of investigations because of its ease of administration, safety, and relatively long duration of a surgical plane of anesthesia. However, its influence on specific cardiopulmonary parameters is unknown because most studies report only postanesthesia values as control data. This study was therefore designed to examine the influence of a commonly used avian anesthetic mixture on resting hemodynamic and respiratory parameters. In an initial series of experiments, the stability and repeatability of respiratory parameters in awake birds over a period of hours on 3 days were determined. In a second series of experiments, cardiopulmonary values in awake and anesthetized birds were compared.
INTRODUCTION
D
ETERMINATION of steady-state conditions in physiological investigations of birds has often been a problem in trials with both awake and anesthetized states. Cardiovascular parameters have been reported to change substantially over 2-3 hours in awake, restrained chickens that were not subjected to any other experimental procedures (Whittow et al., 1965). Results were similar in trials with conscious turkeys that were placed in a holding device for determinations of indirect blood pressure (Krista et al., 1963). On the other hand, alteration of many physiological parameters by anesthetic agents has been generally known for years. Although awake chickens have been tested in numerous cardiovascular and respiratory studies, anesthetized birds must be tested in many other studies due to humane or methodological considerations. Intravenous barbiturate anesthesia in birds is accompanied by a small margin of safety and short duration (Lee, 1953), as well as notable alterations in blood gas tensions (Besch et al., 1971), blood pressure (Ray and Fedde, 1969), and respiratory data (Fedde et al., 1963). To avoid some of the problems
MATERIALS AND METHODS Repeated Trials. Six adult male White Leghorn chickens weighing between 2.0 and 2.3 kg. were tested on three occasions, 7-14 days apart. Heart rate (HR), respiratory volumes 147
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ABSTRACT Steady state cardiovascular and respiratory parameters in adult male chickens while they were awake and after anesthetization with a mixture of chloral hydrate, magnesium sulfate, and pentobarbital were compared. Blood pressure (BP), heart rate (HR), cardiac output (CO), stroke volume (SV), peripheral resistance (TPR), tidal volume (V T ), respiratory rate (RR), minute ventilation (V), end-expiratory carbon dioxide partial pressure (P A C0 2 ), and arterial blood gases and pH were measured simultaneously on birds spontaneously breathing air. Anesthetization resulted in increased HR and RR and decreased BP, CO, TPR, V T , P A C0 2 , and blood gas tensions. The data indicate a depression of cardiovascular function but no change in total ventilation although the relative contributions of V T and RR were changed. Anesthetization increased variability in SV although the other parameters were maintained in a steady-state condition over a 2-h period.
148
T. E .
NIGHTINGALE
Minimum P A 0 2 was monitored on a breath-to-breath basis with a polarographic analyzer (Model OM-11, Beckman Instruments), and maximum P A C 0 2 was measured with an infrared analyzer (LB-1 with linearizer, Beckman Instruments), that sampled at the rate of 100 ml./min. from the distal side of the pneumotachograph. Air flow, V T , ECG, P A C 0 2 and P A 0 2 were continuously recorded on a Beckman Type RM pen recorder. Acute Trials. Eight adult male White Leghorn chickens weighing 2.1 to 2.6 kg. were tested. Under local anesthesia, the trachea was cannulated in a low cervical position, and cannulae were placed in both femoral arteries, in a femoral vein, and into the right atrium by way of the right cutaneous ulnar vein. Positions of all cannulae were verified at the end of the experiments. Rectal temperature was monitored and maintained at 40 ± 1°C. with a heating pad and an infrared lamp. Arterial BP was measured from a femoral
artery with a Statham P23Gb transducer. Mean pressure was determined by electrical damping. HR was determined from the pressure trace. Cardiac output (CO) was measured by dye-dilution with indocyanine green (Hynson, Wescott & Dunning) and a Waters 0-500 densitometer with an XC-350 cuvette. The dye was injected as a bolus into the right atrium. Blood was continuously drawn through the cuvette from a femoral artery at a rate of 7.4 ml./min. by a roller pump (Holter Model 911, Extracorporeal Medical Specialties, Inc.) and returned to the femoral vein. The recorded curves were manually digitized and CO calculated using the computer program of Hall and Tyler (1971). Stroke volumes (SV) (ml./kg./beat) were calculated by dividing CO by HR. Total peripheral resistance (TPR) in arbitrary units, was calculated as the quotient of mean arterial BP (mm.Hg) and CO (inl./min.). This value was multiplied by 100 to obtain a convenient unit. Because central venous pressure is usually near zero, it was disregarded in the calculations. Volume of the tracheal cannula and pneumotachograph in these acute trials was 2.3 ml., which is significantly less than the normal upper airway volume of chicken (Fedde, 1970). V T , RR, V, P A 0 2 and P A C 0 2 were determined as described above. Arterial blood oxygen ( P a 0 2 ) and carbon dioxide (P a C0 2 ) partial pressures and pH (pHa) were measured on an Instrumentation Laboratory Model 113 blood gas system at 40.0° C. Blood removed for analysis was returned to the bird immediately after determinations were completed. Heparin (initial dose of 200 units) was infused slowly intravenously; additional 25 unit doses were administered periodically to prevent clotting in the cannulae. In all trials, the birds were lightly restrained in lateral recumbency, their eyes were covered, and the pneumotachograph was attached to a large T-piece through which air was drawn at 2 L./min. by a vacuum pump.
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and rates, and end-expired oxygen partial pressure ( P A 0 2 ) and carbon dioxide partial pressure (P A C0 2 ) were continuously determined while the birds breathed air. The trachea was intubated with a thin-walled endotracheal tube of sufficient length that a pneumotachograph (Fleisch, Model 00, Instrumentation Associates, New York City) could be attached at the mouth. The oral portion of the endotracheal tube and pneumotachograph resulted in an additional 2.0 ml. of respiratory dead space. Tidal volume (V T , BTPS) was determined by electrical integration of the flow signal (Statham PM15 differential pressure transducer and Beckman 9873B coupler), and respiratory rate (RR) was counted from the V T trace. Minute ventilation (V, BTPS) was derived as the product of V T and RR. HR was determined from the ECG recorded by subcutaneous needle electrodes.
149
CARDIOPULMONARY CHANGES IN CHICKENS
Statistics. An unpaired t test was used to compare group means. Variability within a parameter was calculated and expressed as a coefficient of variation ((SEM -f- X) 100 = % variation). Changes in each value over the 2 h period were analyzed by simple linear regression (Sokal and Rohlf, 1969). The relationship of these variables with time was
expressed and compared by the slope of the regression line, b. RESULTS Repeated Trials. HR and respiratory values for six birds studied on different days are summarized in Table 1. Each bird had a discrete range for each parameter. Values for the group show coefficients of variation of 4% or less for HR, P A 0 2 , and P A C 0 2 and more variable coefficients of 8% for V T and 7% for RR, respectively. Variability of V, which was derived as the product of V T and RR, was 12%. No bird had a cardiopulmonary pattern that might be associated with excitement or stress; e.g. tachycardia, hyperpnea, and hypocapnia. Figure 1 presents the data for a typical bird during the first hour of study on three occasions to show the variability of the parameters. Acute Trials. Average cardiopulmonary data at 10-min. intervals over a 2-h. awake (control) period and for the first 2 h. after anesthetization are compared in Fig. 2. While birds were awake, HR, CO, V T , RR, and V increased gradually, whereas TPR decreased. After anesthetization, BP fell substantially within 10 min., then continued to decline at a slower rate over the next 60 to 70 min., and finally reached essentially stable
TABLE 1.—Heart rate and respiratory values obtained by repeated measurements over a 2 h. period on different days in each of 6 chickens Bird No. 1 2 3 4 5 6
n 35 34 20 28 11 12
HR (beats/min.) 295 ± 3 294 ± 13 285 ± 15 368 ± 4 330 ± 8 288 ± 14
RR VT (ml.) (breaths/min. ) 34.1 ± 3.5 18.6 ± 0.5 20.1 ± 1.3 52.1 ± 6 . 8 14.9 ± 2.4 42.3 ± 4.2 12.3 ± 0.8 42.2 ± 4.3 17.2 ± 0.9 53.2 ± 2.3 16.4 ± 0.6 32.6 ± 3.0
V (ml./min.) 632 ± 56 1046 ± 147 640 ± 164 520 ± 84 913 ± 68 536 ± 59
PAO2 (mm.Hg.) 106 ± 4.8 101 ± 5.6 107 ± 2.5 101 ± 2.0 95 ± 1.4 100 ± 5.2
p A co 2 (mm.Hg) 38.1 ± 2.6 43.7 ± 0.4 38.5 ± 0.4 38.5 ± 1.1 37.8 ± 0.7 39.3 ± 2.7
Group 140 42.8 ± 3.5 16.6 ± 1.1 715 ± 88 102 ± 1.7 39.4 ± 0.9 309 ± 12 values Values are means ± SEM. n = number of observations. HR = heart rate; V T = tidal volume; RR = respiratory rate; V = minute ventilation; P A 0 2 = end-expired P 0 2 ; P A C 0 2 = end-expired PCO,.
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Preliminary trials were conducted for a 4to 8-h. period; data sampled at intervals were used to examine the stress of handling, the laboratory environment, and the influence of the apparatus as a source of variability, as well as to determine whether the chicken would accept the procedure without struggling. After analysis of data from repeated trials indicated no untoward effects of the procedure and after no consistent changes were noted over periods of up to 8 h., more detailed trials on awake birds examined the initial 2 h. interval. After the acute trials were completed, the birds were anesthetized with a mixture of chloral hydrate (4.25 gms./lOO ml.), magnesium sulfate (2.126 gms./lOO ml.), and pentobarbital (0.972 gms./100 ml.), at 2.5 ml. /kg., given intramuscularly at three sites in both the left and the right pectoral muscles. Because a surgical plane of anesthesia is attained within 15-30 min., trials were conducted during this initial 30 min. and for 2 h. thereafter.
150 3 5 0 T
T. E. NIGHTINGALE
HR
4 0 0 T HR (»«*»/n>ln)
(6.et,/m/„,
2 0 0 T ^ P '""Ho) TT
fci - •'
*
CO (i»//i»rn/*j)
300 100
SV (ml/ntot)
1.0 0 60
(unfit)
[^UM
6 0 T V T '""'
_.
Z0X
*
« » (nr*9fn«/min)
"OrRRiiii^'. 20
40
60
TIME (Min)
FIG. 1. Sequential changes in heart rate and respiratory parameters in one conscious chicken determined on 3 occasions. Values shown from top to bottom are: heart rate (HR) in beats/min., tidal volume (VT) in ml., respiratory rate (RR) in breaths/min., minute ventilation (V), in ml./min., and end-expiratory C0 2 partial pressure (P A C0 2 ) in mm. Hg. levels that were approximately 50%of control values. Other parameters found to be significantly different as a result of anesthesia were V T , P A C 0 2 , P a 0 2 , and P a C 0 2 at various times over the 2-h. period. For evaluation of the significance of slow changes in individual parameters, each data set was analyzed by linear regression with time as the independent variable. The slopes of the calculated regression lines, (£>), along with mean values (± SEM), are presented in Table 2. A t test of the regression lines, assuming (3 = 0, revealed significant changes in HR, CO, TPR, V T , RR, V, P A C 0 2 , and P a C 0 2 over the 2-h. awake period. After anesthesia, immediate changes such as BP decreases and RR increases would be misleading when steady-state values are determined (Fig. 2). Accordingly, mean values in Table 2 were calculated for a 2-h. period
20 1500 T V (ml/min) (ml/min) j V 1000
40-?°°«
(mm Ho)
P c t , (mm Hi) 4 00 T « ^ Z
760 r 7.50
pH
a '""""'
[^^4^ 0
30
60
90 120
T I M E (min)
FIG. 2. Mean cardiopulmonary values over a 2 h. period while awake (0O) and after anesthetization with chloral hydrate-magnesium sulfatepentobarbital (A A). Vertical lines are SEM. Open symbols indicate values significantly different from value at time zero (P < 0.05). * indicates anesthetized values significantly different from awake value (P < 0.05). Values shown from top to bottom are: heart rate (HR); mean blood pressure (BP); cardiac output (CO); stroke volume (SV); total peripheral resistance (TPR); tidal volume (V T ); respiratory rate (RR); minute ventilation (V); endexpired PC0 2 (P A C0 2 ); arterial P0 2 (P a 0 2 ); arterial PC0 2 (P a C0 2 ); and arterial pH (pHa).
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0
TPR
Awake (control) Coeff. of X ± SEM Measurement variation, % b n X ± SE HR, beats/min. 295 ± 8 3 (82) 330 ± 12 1.02f Mean BP, mm.Hg. 162 ± 3 2 0.136 (83) 80 ± 3* CO, ml./kg./min. 222 ± 13 6 0.966t (32) 173 ± 13 SV, ml./kg./beat 0.66 ± 0.03 5 0.001 (32) 0.56 ± 0.0 TPR, units 38.8 ± 3.2 8 (32) 25.3 ± 2.0 -0.176t V T , ml. 41.9 ± 0.9 2 0.091t (69) 34.0 ± 1.4 RR, breaths/min. 23.8 ± 0.8 3 (68) 33.7 ± 1.8 0.066t V, ml./min. 990 ± 37 4 (69) 1086 ± 57 4.0t P A C 0 2 , mm. Hg 34.9 ± 0.6 2 -0.005 (64) 27.8 ± 0.7 P a C 0 2 , mm. Hg 29.2 ± 0.8 3 0.056t (56) 24.1 ± 0.8 P 0 2 , mm. Hg 70.0 ± 1.6 2 0.023 (50) 62.9 ± 1.7 p H a , units 7.536 ± 0.007 -0.00005 (58) 7.545 ± 0.0 — HR = heart rate. Mean BP = meap blood pressure. CO = cardiac output. SV = stroke volu tidal volume. RR = respiratory rate. V = minute ventilation. P A C 0 2 = end-expired P C 0 2 . P a C 0 = arterial pH. Number of measurements shown in parenthesis, t indicates regression line slopes (b) significan comparisons between awake_and anesthetized values indicated by * P < 0.05, ** P < 0.01, *** P < 0 Anesthetized values for X ± SEM calculated for 30 to 150 min. post-anesthesia. Anesthetized administration (0 to 120 min.).
TABLE 2.—Cardiopulmonary values for 2-h period before and after a
om http://ps.oxfordjournals.org/ at North Dakota State University on May 16, 2015
152
T. E .
NIGHTINGALE
DISCUSSION The initial series of repeated trials indicated that the experimental protocol and apparatus presented no handicap to respiratory function because there were no significant changes over periods of up to 8 hr., and the birds did not struggle nor appear to be in distress. These repeated trials provided a basis for determining alterations due to the more extensive preparation and handling during the acute awake trials; and because HR in the two series were not different, all the awake
birds were considered to be in a basal, nonexcited state (Tummons and Sturkie, 1969). Data from acute awake trials agree closely with comparable values reported for hens (Piiper et al., 1970), and for adult males (Butler and Taylor, 1974; King and Payne, 1964; Sturkie and Vogel, 1959), as to both means and standard errors. Cardiovascular changes resulting from anesthesia were a slight tachycardia and reductions in CO, BP, and TPR. Similar changes have been noted in barbiturate-anesthetized mammals and attributed to a vagolytic response (Barlow and Knott, 1964; Olmsted and Page, 1966), or, more likely, to a direct myocardial depressing action of the barbiturates (Cox, 1972; Nash et al, 1956; Price, 1960). Previous reports dealing with either pentobarbital- (Ray and Fedde, 1969; Richards and Sykes, 1967), or Equithesin-anesthetized chickens (Nightingale, 1976; Tschorn and Fedde, 1974), have indicated hypotension although only one of these papers (Nightingale, 1976) reported data concerning CO (169 ± 16 ml. /kg. /min.) and TPR (41.7 ± 5.2 units) under anesthesia. V was not altered by anesthetization although the relationships of V T and RR were significantly changed. Although V T was reduced, it was not nearly as depressed as previously observed in pentobarbital-anesthetized chickens (Nightingale, 1971), nor was there any indication of hypercapnia and respiratory acidosis, which is a common feature of pentobarbital anesthesia (Besch et al., 1971; Nightingale and Fedde, 1972). The lower P a 0 2 was suggestive of a reduced effective ventilation although this change was not as profound as that under pentobarbital anesthesia (Besch et al., 1971). Although anesthesia-related changes in resting cardiopulmonary values with chloral hydrate-magnesium sulfate-pentobarbital were significant, comparisons of changing cardiopulmonary values over 2 hour trials while birds were awake as well as those
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beginning 30 min. after the anesthetic was administered because by 30 min., the animals were in a relatively steady state. Cardiovascular changes due to anesthetization included tachycardia and decreases in BP, CO, and TPR. The change in SV was not significant. As a result of the decreased V T and increased RR, V was unchanged. P A C 0 2 decreased, along with both P a 0 2 and P a C 0 2 , whereas arterial pK (pHa) increased. Regression analysis of data obtained during anesthesia revealed that BP, TPR, P A C 0 2 , and P a C 0 2 all decreased significantly (P < 0.05) with time when analyzed from the time the anesthetic was administered. However, if the first 30 min. were assumed to be an "unsteady state" and regression analyses were performed on data taken from 30 to 120 min. after anesthesia, only BP changed significantly, and at a lower rate than that calculated from 0- to 120-min. data (b was reduced from -0.522 to -0.261 mm.Hg/ min.). In awake trials, the coefficient of variation was 4% or less for all parameters except CO and two values that are derived from CO (SV and TPR). The variability for these three parameters was 5 to 8%. After anesthetization, variability was unchanged for two values (TPR & P a C O , ) , increased in variability by 1 to 2% for eight values, and increased substantially for only SV (Table 2).
CARDIOPULMONARY CHANGES IN CHICKENS
beginning 30 min. after anesthesia are suggestive of normal variation and not of an unsteady state resulting from anesthesia.
ACKNOWLEDGMENTS T h e author gratefully achnowledges the technical assistance of Mr. J o h n Richardi and Mrs. Jaci Wheatley, and the secretarial assistance of M r s . Patricia Palmer.
Barlow, G., and D. H. Knott, 1964. Hemodynamic alterations after 30 minutes of pentobarbital sodium anesthesia in dogs. Am. J. Physiol. 207: 764-766. Besch, E. L., R. R. Burton and A. H. Smith, 1971. Influence of chronic hypoxia on blood gas tensions and pH in domestic fowl. Am. J. Physiol. 220: 1379-1382. Butler, P. J., and E. W. Taylor, 1974. Responses of the respiratory and cardiovascular systems of chickens and pigeons to changes in P a 0 2 and P a C0 2 . Respiration Physiol. 21: 351-363. Cox,R. H., 1972. Influence of pentobarbital anesthesia on cardiovascular function in trained dogs. Am. J. Physiol. 223: 651-659. Fedde, M. R., 1970. Peripheral control of avian respiration. Federation Proc. 29: 1664-1673. Fedde, M. R., R. E. Burger and R. L. Kitchell, 1963. The effect of anesthesia and age on respiration following bilateral, cervical vagotomy in the fowl. Poultry Sci. 42: 1212-1223. Gandal, C. P., 1956. Satisfactory general anesthesia in birds. J. Am. Vet. Med. Assoc. 128: 332-334. Hall, R.C., and M.J. Tyler, 1971. A computer program to estimate cardiac output directly from dye-dilution curves. J. Appl. Physiol. 32: 145-147. King, A. S., and D. C. Payne, 1964. Normal breathing and the effects of posture in Gallus domesticus. J. Physiol., London, 174: 340-347. Krista, L. M., R. E. Burger and P. E. Waibel, 1963. Blood pressure and heart rate in the turkey as measured by the indirect method and their modifica-
tions by pharmacological agents. Poultry Sci. 42: 646-652. Lee, C. C , 1953. Experimental studies on the actions of several anesthetics in domestic fowls. Poultry Sci. 32: 624-627. Nash, C. B., F. Davis and R. A. Woodbury, 1956. Cardiovascular effects of anesthetic doses of pentobarbital sodium. Am. J. Physiol. 185: 107-112. Nightingale, T. E., 1971. Respiratory control in Gallus domesticus. Ph.D. Thesis. Kansas State University, Manhattan. Nightingale, T. E., 1976. Acute isovolemic anemia in anesthetized chickens. Am. J. Physiol. In press. Nightingale, T. E., and M. R. Fedde, 1972. Determination of normal buffer line for chicken blood. Respiration Physiol. 14: 353-365. Olmsted, F., and I. H. Page, 1966. Hemodynamic changes in dogs caused by sodium pentobarbital anesthesia. Am. J. Physiol. 210: 817-820. Piiper, J., F. Drees and P. Scheid, 1970. Gas exchange in the domestic fowl during spontaneous breathing and artificial ventilation. Respiration Physiol. 9: 234-245. Price, H. L., 1960. General anesthesia and circulatory homeostasis. Physiol. Rev. 40: 187-218. Ray, P. J., and M. R. Fedde, 1969. Responses to alterations in respiratory P0 2 and PC0 2 in the chicken. Respiration Physiol. 6: 135-143. Richards, S. A., and A. H. Sykes, 1967. The effects of hypoxia, hypercapnia and asphyxia in the domestic fowl (Gallus domesticus). Comp. Biochem. Physiol. 21:691-701. Sokal, R. R., and F. J. Rohlf, 1969. Biometry. Freeman, San Francisco. Sturkie, P. D., and J. A. Vogel, 1959. Cardiac output, central blood volume and peripheral resistance in chickens. Am. J. Physiol. 197: 1165-1166. Tschorn, R. R., and M. R. Fedde, 1974. Cardiopulmonary responses to carbon monoxide breathing in the chicken. Respiration Physiol. 20: 303-311. Tummons, J. L., and P. D. Sturkie, 1969. Nervous control of heart rate during excitement in the adult White Leghorn cock. Am. J. Physiol. 216: 14371440. Whittow, G. C , P. D. Sturkie and G. Stein, Jr., 1965. Cardiovascular changes in restrained chickens. Poultry Sci. 44: 1452-1459.
MAY 18-20, 1977. CELLULAR FUNCTION AND MOLECULAR STRUCTURE: A SYMPOSIUM ON BIOPHYSICAL APPROACHES TO BIOLOGICAL PROBLEMS, UNIVERSITY OF MISSOURI-COLUMBIA, COLUMBIA, MISSOURI JULY 18-23, 1977. XXVIITH INTERNATIONAL CONGRESS OF PHYSIOLOGICAL SCIENCES, PARIS
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REFERENCES
153
NIGHTINGALE
USDA Poultry Research Laboratory, R.D. 2, Box 600, Georgetown, Delaware 19947 (Received for publication May 5, 1976)
POULTRY SCIENCE 56: 147-153, 1977
inherent in handling, restraint, and intravenous injection of birds, Gandal (1956) reported the advantage of intramuscular administration of a solution containing chloral hydrate, magnesium sulfate, and pentobarbital. This mixture has become accepted for many types of investigations because of its ease of administration, safety, and relatively long duration of a surgical plane of anesthesia. However, its influence on specific cardiopulmonary parameters is unknown because most studies report only postanesthesia values as control data. This study was therefore designed to examine the influence of a commonly used avian anesthetic mixture on resting hemodynamic and respiratory parameters. In an initial series of experiments, the stability and repeatability of respiratory parameters in awake birds over a period of hours on 3 days were determined. In a second series of experiments, cardiopulmonary values in awake and anesthetized birds were compared.
INTRODUCTION
D
ETERMINATION of steady-state conditions in physiological investigations of birds has often been a problem in trials with both awake and anesthetized states. Cardiovascular parameters have been reported to change substantially over 2-3 hours in awake, restrained chickens that were not subjected to any other experimental procedures (Whittow et al., 1965). Results were similar in trials with conscious turkeys that were placed in a holding device for determinations of indirect blood pressure (Krista et al., 1963). On the other hand, alteration of many physiological parameters by anesthetic agents has been generally known for years. Although awake chickens have been tested in numerous cardiovascular and respiratory studies, anesthetized birds must be tested in many other studies due to humane or methodological considerations. Intravenous barbiturate anesthesia in birds is accompanied by a small margin of safety and short duration (Lee, 1953), as well as notable alterations in blood gas tensions (Besch et al., 1971), blood pressure (Ray and Fedde, 1969), and respiratory data (Fedde et al., 1963). To avoid some of the problems
MATERIALS AND METHODS Repeated Trials. Six adult male White Leghorn chickens weighing between 2.0 and 2.3 kg. were tested on three occasions, 7-14 days apart. Heart rate (HR), respiratory volumes 147
Downloaded from http://ps.oxfordjournals.org/ at North Dakota State University on May 16, 2015
ABSTRACT Steady state cardiovascular and respiratory parameters in adult male chickens while they were awake and after anesthetization with a mixture of chloral hydrate, magnesium sulfate, and pentobarbital were compared. Blood pressure (BP), heart rate (HR), cardiac output (CO), stroke volume (SV), peripheral resistance (TPR), tidal volume (V T ), respiratory rate (RR), minute ventilation (V), end-expiratory carbon dioxide partial pressure (P A C0 2 ), and arterial blood gases and pH were measured simultaneously on birds spontaneously breathing air. Anesthetization resulted in increased HR and RR and decreased BP, CO, TPR, V T , P A C0 2 , and blood gas tensions. The data indicate a depression of cardiovascular function but no change in total ventilation although the relative contributions of V T and RR were changed. Anesthetization increased variability in SV although the other parameters were maintained in a steady-state condition over a 2-h period.
148
T. E .
NIGHTINGALE
Minimum P A 0 2 was monitored on a breath-to-breath basis with a polarographic analyzer (Model OM-11, Beckman Instruments), and maximum P A C 0 2 was measured with an infrared analyzer (LB-1 with linearizer, Beckman Instruments), that sampled at the rate of 100 ml./min. from the distal side of the pneumotachograph. Air flow, V T , ECG, P A C 0 2 and P A 0 2 were continuously recorded on a Beckman Type RM pen recorder. Acute Trials. Eight adult male White Leghorn chickens weighing 2.1 to 2.6 kg. were tested. Under local anesthesia, the trachea was cannulated in a low cervical position, and cannulae were placed in both femoral arteries, in a femoral vein, and into the right atrium by way of the right cutaneous ulnar vein. Positions of all cannulae were verified at the end of the experiments. Rectal temperature was monitored and maintained at 40 ± 1°C. with a heating pad and an infrared lamp. Arterial BP was measured from a femoral
artery with a Statham P23Gb transducer. Mean pressure was determined by electrical damping. HR was determined from the pressure trace. Cardiac output (CO) was measured by dye-dilution with indocyanine green (Hynson, Wescott & Dunning) and a Waters 0-500 densitometer with an XC-350 cuvette. The dye was injected as a bolus into the right atrium. Blood was continuously drawn through the cuvette from a femoral artery at a rate of 7.4 ml./min. by a roller pump (Holter Model 911, Extracorporeal Medical Specialties, Inc.) and returned to the femoral vein. The recorded curves were manually digitized and CO calculated using the computer program of Hall and Tyler (1971). Stroke volumes (SV) (ml./kg./beat) were calculated by dividing CO by HR. Total peripheral resistance (TPR) in arbitrary units, was calculated as the quotient of mean arterial BP (mm.Hg) and CO (inl./min.). This value was multiplied by 100 to obtain a convenient unit. Because central venous pressure is usually near zero, it was disregarded in the calculations. Volume of the tracheal cannula and pneumotachograph in these acute trials was 2.3 ml., which is significantly less than the normal upper airway volume of chicken (Fedde, 1970). V T , RR, V, P A 0 2 and P A C 0 2 were determined as described above. Arterial blood oxygen ( P a 0 2 ) and carbon dioxide (P a C0 2 ) partial pressures and pH (pHa) were measured on an Instrumentation Laboratory Model 113 blood gas system at 40.0° C. Blood removed for analysis was returned to the bird immediately after determinations were completed. Heparin (initial dose of 200 units) was infused slowly intravenously; additional 25 unit doses were administered periodically to prevent clotting in the cannulae. In all trials, the birds were lightly restrained in lateral recumbency, their eyes were covered, and the pneumotachograph was attached to a large T-piece through which air was drawn at 2 L./min. by a vacuum pump.
Downloaded from http://ps.oxfordjournals.org/ at North Dakota State University on May 16, 2015
and rates, and end-expired oxygen partial pressure ( P A 0 2 ) and carbon dioxide partial pressure (P A C0 2 ) were continuously determined while the birds breathed air. The trachea was intubated with a thin-walled endotracheal tube of sufficient length that a pneumotachograph (Fleisch, Model 00, Instrumentation Associates, New York City) could be attached at the mouth. The oral portion of the endotracheal tube and pneumotachograph resulted in an additional 2.0 ml. of respiratory dead space. Tidal volume (V T , BTPS) was determined by electrical integration of the flow signal (Statham PM15 differential pressure transducer and Beckman 9873B coupler), and respiratory rate (RR) was counted from the V T trace. Minute ventilation (V, BTPS) was derived as the product of V T and RR. HR was determined from the ECG recorded by subcutaneous needle electrodes.
149
CARDIOPULMONARY CHANGES IN CHICKENS
Statistics. An unpaired t test was used to compare group means. Variability within a parameter was calculated and expressed as a coefficient of variation ((SEM -f- X) 100 = % variation). Changes in each value over the 2 h period were analyzed by simple linear regression (Sokal and Rohlf, 1969). The relationship of these variables with time was
expressed and compared by the slope of the regression line, b. RESULTS Repeated Trials. HR and respiratory values for six birds studied on different days are summarized in Table 1. Each bird had a discrete range for each parameter. Values for the group show coefficients of variation of 4% or less for HR, P A 0 2 , and P A C 0 2 and more variable coefficients of 8% for V T and 7% for RR, respectively. Variability of V, which was derived as the product of V T and RR, was 12%. No bird had a cardiopulmonary pattern that might be associated with excitement or stress; e.g. tachycardia, hyperpnea, and hypocapnia. Figure 1 presents the data for a typical bird during the first hour of study on three occasions to show the variability of the parameters. Acute Trials. Average cardiopulmonary data at 10-min. intervals over a 2-h. awake (control) period and for the first 2 h. after anesthetization are compared in Fig. 2. While birds were awake, HR, CO, V T , RR, and V increased gradually, whereas TPR decreased. After anesthetization, BP fell substantially within 10 min., then continued to decline at a slower rate over the next 60 to 70 min., and finally reached essentially stable
TABLE 1.—Heart rate and respiratory values obtained by repeated measurements over a 2 h. period on different days in each of 6 chickens Bird No. 1 2 3 4 5 6
n 35 34 20 28 11 12
HR (beats/min.) 295 ± 3 294 ± 13 285 ± 15 368 ± 4 330 ± 8 288 ± 14
RR VT (ml.) (breaths/min. ) 34.1 ± 3.5 18.6 ± 0.5 20.1 ± 1.3 52.1 ± 6 . 8 14.9 ± 2.4 42.3 ± 4.2 12.3 ± 0.8 42.2 ± 4.3 17.2 ± 0.9 53.2 ± 2.3 16.4 ± 0.6 32.6 ± 3.0
V (ml./min.) 632 ± 56 1046 ± 147 640 ± 164 520 ± 84 913 ± 68 536 ± 59
PAO2 (mm.Hg.) 106 ± 4.8 101 ± 5.6 107 ± 2.5 101 ± 2.0 95 ± 1.4 100 ± 5.2
p A co 2 (mm.Hg) 38.1 ± 2.6 43.7 ± 0.4 38.5 ± 0.4 38.5 ± 1.1 37.8 ± 0.7 39.3 ± 2.7
Group 140 42.8 ± 3.5 16.6 ± 1.1 715 ± 88 102 ± 1.7 39.4 ± 0.9 309 ± 12 values Values are means ± SEM. n = number of observations. HR = heart rate; V T = tidal volume; RR = respiratory rate; V = minute ventilation; P A 0 2 = end-expired P 0 2 ; P A C 0 2 = end-expired PCO,.
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Preliminary trials were conducted for a 4to 8-h. period; data sampled at intervals were used to examine the stress of handling, the laboratory environment, and the influence of the apparatus as a source of variability, as well as to determine whether the chicken would accept the procedure without struggling. After analysis of data from repeated trials indicated no untoward effects of the procedure and after no consistent changes were noted over periods of up to 8 h., more detailed trials on awake birds examined the initial 2 h. interval. After the acute trials were completed, the birds were anesthetized with a mixture of chloral hydrate (4.25 gms./lOO ml.), magnesium sulfate (2.126 gms./lOO ml.), and pentobarbital (0.972 gms./100 ml.), at 2.5 ml. /kg., given intramuscularly at three sites in both the left and the right pectoral muscles. Because a surgical plane of anesthesia is attained within 15-30 min., trials were conducted during this initial 30 min. and for 2 h. thereafter.
150 3 5 0 T
T. E. NIGHTINGALE
HR
4 0 0 T HR (»«*»/n>ln)
(6.et,/m/„,
2 0 0 T ^ P '""Ho) TT
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*
CO (i»//i»rn/*j)
300 100
SV (ml/ntot)
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(unfit)
[^UM
6 0 T V T '""'
_.
Z0X
*
« » (nr*9fn«/min)
"OrRRiiii^'. 20
40
60
TIME (Min)
FIG. 1. Sequential changes in heart rate and respiratory parameters in one conscious chicken determined on 3 occasions. Values shown from top to bottom are: heart rate (HR) in beats/min., tidal volume (VT) in ml., respiratory rate (RR) in breaths/min., minute ventilation (V), in ml./min., and end-expiratory C0 2 partial pressure (P A C0 2 ) in mm. Hg. levels that were approximately 50%of control values. Other parameters found to be significantly different as a result of anesthesia were V T , P A C 0 2 , P a 0 2 , and P a C 0 2 at various times over the 2-h. period. For evaluation of the significance of slow changes in individual parameters, each data set was analyzed by linear regression with time as the independent variable. The slopes of the calculated regression lines, (£>), along with mean values (± SEM), are presented in Table 2. A t test of the regression lines, assuming (3 = 0, revealed significant changes in HR, CO, TPR, V T , RR, V, P A C 0 2 , and P a C 0 2 over the 2-h. awake period. After anesthesia, immediate changes such as BP decreases and RR increases would be misleading when steady-state values are determined (Fig. 2). Accordingly, mean values in Table 2 were calculated for a 2-h. period
20 1500 T V (ml/min) (ml/min) j V 1000
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(mm Ho)
P c t , (mm Hi) 4 00 T « ^ Z
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30
60
90 120
T I M E (min)
FIG. 2. Mean cardiopulmonary values over a 2 h. period while awake (0O) and after anesthetization with chloral hydrate-magnesium sulfatepentobarbital (A A). Vertical lines are SEM. Open symbols indicate values significantly different from value at time zero (P < 0.05). * indicates anesthetized values significantly different from awake value (P < 0.05). Values shown from top to bottom are: heart rate (HR); mean blood pressure (BP); cardiac output (CO); stroke volume (SV); total peripheral resistance (TPR); tidal volume (V T ); respiratory rate (RR); minute ventilation (V); endexpired PC0 2 (P A C0 2 ); arterial P0 2 (P a 0 2 ); arterial PC0 2 (P a C0 2 ); and arterial pH (pHa).
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0
TPR
Awake (control) Coeff. of X ± SEM Measurement variation, % b n X ± SE HR, beats/min. 295 ± 8 3 (82) 330 ± 12 1.02f Mean BP, mm.Hg. 162 ± 3 2 0.136 (83) 80 ± 3* CO, ml./kg./min. 222 ± 13 6 0.966t (32) 173 ± 13 SV, ml./kg./beat 0.66 ± 0.03 5 0.001 (32) 0.56 ± 0.0 TPR, units 38.8 ± 3.2 8 (32) 25.3 ± 2.0 -0.176t V T , ml. 41.9 ± 0.9 2 0.091t (69) 34.0 ± 1.4 RR, breaths/min. 23.8 ± 0.8 3 (68) 33.7 ± 1.8 0.066t V, ml./min. 990 ± 37 4 (69) 1086 ± 57 4.0t P A C 0 2 , mm. Hg 34.9 ± 0.6 2 -0.005 (64) 27.8 ± 0.7 P a C 0 2 , mm. Hg 29.2 ± 0.8 3 0.056t (56) 24.1 ± 0.8 P 0 2 , mm. Hg 70.0 ± 1.6 2 0.023 (50) 62.9 ± 1.7 p H a , units 7.536 ± 0.007 -0.00005 (58) 7.545 ± 0.0 — HR = heart rate. Mean BP = meap blood pressure. CO = cardiac output. SV = stroke volu tidal volume. RR = respiratory rate. V = minute ventilation. P A C 0 2 = end-expired P C 0 2 . P a C 0 = arterial pH. Number of measurements shown in parenthesis, t indicates regression line slopes (b) significan comparisons between awake_and anesthetized values indicated by * P < 0.05, ** P < 0.01, *** P < 0 Anesthetized values for X ± SEM calculated for 30 to 150 min. post-anesthesia. Anesthetized administration (0 to 120 min.).
TABLE 2.—Cardiopulmonary values for 2-h period before and after a
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152
T. E .
NIGHTINGALE
DISCUSSION The initial series of repeated trials indicated that the experimental protocol and apparatus presented no handicap to respiratory function because there were no significant changes over periods of up to 8 hr., and the birds did not struggle nor appear to be in distress. These repeated trials provided a basis for determining alterations due to the more extensive preparation and handling during the acute awake trials; and because HR in the two series were not different, all the awake
birds were considered to be in a basal, nonexcited state (Tummons and Sturkie, 1969). Data from acute awake trials agree closely with comparable values reported for hens (Piiper et al., 1970), and for adult males (Butler and Taylor, 1974; King and Payne, 1964; Sturkie and Vogel, 1959), as to both means and standard errors. Cardiovascular changes resulting from anesthesia were a slight tachycardia and reductions in CO, BP, and TPR. Similar changes have been noted in barbiturate-anesthetized mammals and attributed to a vagolytic response (Barlow and Knott, 1964; Olmsted and Page, 1966), or, more likely, to a direct myocardial depressing action of the barbiturates (Cox, 1972; Nash et al, 1956; Price, 1960). Previous reports dealing with either pentobarbital- (Ray and Fedde, 1969; Richards and Sykes, 1967), or Equithesin-anesthetized chickens (Nightingale, 1976; Tschorn and Fedde, 1974), have indicated hypotension although only one of these papers (Nightingale, 1976) reported data concerning CO (169 ± 16 ml. /kg. /min.) and TPR (41.7 ± 5.2 units) under anesthesia. V was not altered by anesthetization although the relationships of V T and RR were significantly changed. Although V T was reduced, it was not nearly as depressed as previously observed in pentobarbital-anesthetized chickens (Nightingale, 1971), nor was there any indication of hypercapnia and respiratory acidosis, which is a common feature of pentobarbital anesthesia (Besch et al., 1971; Nightingale and Fedde, 1972). The lower P a 0 2 was suggestive of a reduced effective ventilation although this change was not as profound as that under pentobarbital anesthesia (Besch et al., 1971). Although anesthesia-related changes in resting cardiopulmonary values with chloral hydrate-magnesium sulfate-pentobarbital were significant, comparisons of changing cardiopulmonary values over 2 hour trials while birds were awake as well as those
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beginning 30 min. after the anesthetic was administered because by 30 min., the animals were in a relatively steady state. Cardiovascular changes due to anesthetization included tachycardia and decreases in BP, CO, and TPR. The change in SV was not significant. As a result of the decreased V T and increased RR, V was unchanged. P A C 0 2 decreased, along with both P a 0 2 and P a C 0 2 , whereas arterial pK (pHa) increased. Regression analysis of data obtained during anesthesia revealed that BP, TPR, P A C 0 2 , and P a C 0 2 all decreased significantly (P < 0.05) with time when analyzed from the time the anesthetic was administered. However, if the first 30 min. were assumed to be an "unsteady state" and regression analyses were performed on data taken from 30 to 120 min. after anesthesia, only BP changed significantly, and at a lower rate than that calculated from 0- to 120-min. data (b was reduced from -0.522 to -0.261 mm.Hg/ min.). In awake trials, the coefficient of variation was 4% or less for all parameters except CO and two values that are derived from CO (SV and TPR). The variability for these three parameters was 5 to 8%. After anesthetization, variability was unchanged for two values (TPR & P a C O , ) , increased in variability by 1 to 2% for eight values, and increased substantially for only SV (Table 2).
CARDIOPULMONARY CHANGES IN CHICKENS
beginning 30 min. after anesthesia are suggestive of normal variation and not of an unsteady state resulting from anesthesia.
ACKNOWLEDGMENTS T h e author gratefully achnowledges the technical assistance of Mr. J o h n Richardi and Mrs. Jaci Wheatley, and the secretarial assistance of M r s . Patricia Palmer.
Barlow, G., and D. H. Knott, 1964. Hemodynamic alterations after 30 minutes of pentobarbital sodium anesthesia in dogs. Am. J. Physiol. 207: 764-766. Besch, E. L., R. R. Burton and A. H. Smith, 1971. Influence of chronic hypoxia on blood gas tensions and pH in domestic fowl. Am. J. Physiol. 220: 1379-1382. Butler, P. J., and E. W. Taylor, 1974. Responses of the respiratory and cardiovascular systems of chickens and pigeons to changes in P a 0 2 and P a C0 2 . Respiration Physiol. 21: 351-363. Cox,R. H., 1972. Influence of pentobarbital anesthesia on cardiovascular function in trained dogs. Am. J. Physiol. 223: 651-659. Fedde, M. R., 1970. Peripheral control of avian respiration. Federation Proc. 29: 1664-1673. Fedde, M. R., R. E. Burger and R. L. Kitchell, 1963. The effect of anesthesia and age on respiration following bilateral, cervical vagotomy in the fowl. Poultry Sci. 42: 1212-1223. Gandal, C. P., 1956. Satisfactory general anesthesia in birds. J. Am. Vet. Med. Assoc. 128: 332-334. Hall, R.C., and M.J. Tyler, 1971. A computer program to estimate cardiac output directly from dye-dilution curves. J. Appl. Physiol. 32: 145-147. King, A. S., and D. C. Payne, 1964. Normal breathing and the effects of posture in Gallus domesticus. J. Physiol., London, 174: 340-347. Krista, L. M., R. E. Burger and P. E. Waibel, 1963. Blood pressure and heart rate in the turkey as measured by the indirect method and their modifica-
tions by pharmacological agents. Poultry Sci. 42: 646-652. Lee, C. C , 1953. Experimental studies on the actions of several anesthetics in domestic fowls. Poultry Sci. 32: 624-627. Nash, C. B., F. Davis and R. A. Woodbury, 1956. Cardiovascular effects of anesthetic doses of pentobarbital sodium. Am. J. Physiol. 185: 107-112. Nightingale, T. E., 1971. Respiratory control in Gallus domesticus. Ph.D. Thesis. Kansas State University, Manhattan. Nightingale, T. E., 1976. Acute isovolemic anemia in anesthetized chickens. Am. J. Physiol. In press. Nightingale, T. E., and M. R. Fedde, 1972. Determination of normal buffer line for chicken blood. Respiration Physiol. 14: 353-365. Olmsted, F., and I. H. Page, 1966. Hemodynamic changes in dogs caused by sodium pentobarbital anesthesia. Am. J. Physiol. 210: 817-820. Piiper, J., F. Drees and P. Scheid, 1970. Gas exchange in the domestic fowl during spontaneous breathing and artificial ventilation. Respiration Physiol. 9: 234-245. Price, H. L., 1960. General anesthesia and circulatory homeostasis. Physiol. Rev. 40: 187-218. Ray, P. J., and M. R. Fedde, 1969. Responses to alterations in respiratory P0 2 and PC0 2 in the chicken. Respiration Physiol. 6: 135-143. Richards, S. A., and A. H. Sykes, 1967. The effects of hypoxia, hypercapnia and asphyxia in the domestic fowl (Gallus domesticus). Comp. Biochem. Physiol. 21:691-701. Sokal, R. R., and F. J. Rohlf, 1969. Biometry. Freeman, San Francisco. Sturkie, P. D., and J. A. Vogel, 1959. Cardiac output, central blood volume and peripheral resistance in chickens. Am. J. Physiol. 197: 1165-1166. Tschorn, R. R., and M. R. Fedde, 1974. Cardiopulmonary responses to carbon monoxide breathing in the chicken. Respiration Physiol. 20: 303-311. Tummons, J. L., and P. D. Sturkie, 1969. Nervous control of heart rate during excitement in the adult White Leghorn cock. Am. J. Physiol. 216: 14371440. Whittow, G. C , P. D. Sturkie and G. Stein, Jr., 1965. Cardiovascular changes in restrained chickens. Poultry Sci. 44: 1452-1459.
MAY 18-20, 1977. CELLULAR FUNCTION AND MOLECULAR STRUCTURE: A SYMPOSIUM ON BIOPHYSICAL APPROACHES TO BIOLOGICAL PROBLEMS, UNIVERSITY OF MISSOURI-COLUMBIA, COLUMBIA, MISSOURI JULY 18-23, 1977. XXVIITH INTERNATIONAL CONGRESS OF PHYSIOLOGICAL SCIENCES, PARIS
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REFERENCES
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