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Effects of anesthetic agents on the adrenocortical system of female baboons.

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American Journal of Primatology 13:325-332 (1987)
BRIEF REPORT
Effects of Anesthetic Agents on the Adrenocortical System
of Female Baboons
MARGARET L. WALKER1, GERALD J. PEPE3, NELSON L. GARNET?, AND
EUGENE D. ALBRECHT'
'Departments of Obstetricd@necology and Physiology, University of Maryland School of
Medicine, Baltimore, 'Central Animal Facility, University of Maryland School of Medicine,
Baltimore, 3Department of Physiology, Eastern Virginia Medical School, Norfolk, Virginia
Invasive surgical procedures are often used to study the reproductive and
adrenocortical endocrine systems in primates. Anesthetic agents must,
therefore, be used that have the least confounding effects on these systems.
The present study was designed to characterize various adrenocortical endocrine responses of female baboons (Papw anubis), each treated for 120
minutes with an infusion of ketamine HC1 (6 mg/min) in 5% dextrose in
water (0.40 mumin), a combination of ketamine and acetylpromazine (0.6
mg acetylpromazine and 6 mg ketamine HCllmin) in 5%dextrose in water,
or inhalation of vaporized halothane (1.0% halothane, NzO 25%, 1litedmin;
0 2 75%, 3 liters/min). Blood samples were collected throughout the treatment period, and serum was assayed for prolactin (PRL), dehydroepiandrosterone (DHA), dehydroepiandrosterone sulfate (DHAS), and cortisol (F).No
significant elevations in DHA, F, or PRL concentrations were found following infusion of ketamine alone. Only serum DHAS concentrations were
significantly altered after long-term exposure to ketamine. Acetylpromazine
increased PRL concentrations tenfold to levels significantly greater than
those in ketamine- and halothane-treated animals but had no effect on
serum DHA, DHAS, or F. Treatment with halothane had no effect on serum
PRL, DHA, or DHAS but did suppress F (>40%) concentrations over time.
These data indicate that ketamine is best suited for the collection of biological samples when deep analgesia is not required but that halothane is
preferable in the latter situation.
Key words: cortisol, prolactin, dehydroepiandrosterone, ketamine, acetylpromazine,
halothane
INTRODUCTION
The baboon is an excellent nonhuman primate model for the study of reproductive endocrinology because the patterns of hormone production and metabolism are
relatively similar to those in humans [Goldzieher & Axelrod, 1969;Albrecht & Pepe,
1985;Pepe & Albrecht, 1985al. Invasive experimental procedures are often employed
Received February 2, 1987; revision accepted April 14, 1987.
Address reprint requests to Dr. Eugene D. Albrecht, Department of Obstetrics and Gynecology,University
of Maryland School of Medicine, 655 West Baltimore Street, Baltimore, MD 21201.
0 1987 Alan R. Liss, Inc.
326 I Walker et a1
in our studies with baboons to determine the factors essential to the regulation of
the hypothalamic-hypophyseal-adrenocorticalaxis, including the formation of cortisol @'), dehydroepiandrosterone (DHA), and DHA sulfate (DHAS). Because the endocrine system is sensitive to the differential effects of various anesthetics [Ferin et
al, 1976; Quadri et al, 1978; Puri et al, 19811, it is essential that anesthetic agents
are used that produce the fewest possible confounding effects.
Previous studies [Albrecht et al, 19771 with baboons have demonstrated that
ketamine prevents the stress-induced rise in F typically associated with handling
and restraint. However, ketamine alone apparently does not provide the analgesia
required for major abdominal surgery [Hughes & Lang, 19831. To circumvent this
problem, a combination of ketamine and a central nervous system depressant such
as acetylpromazine is commonly employed to induce the analgesidanesthesia required for major surgery. Similarly, general anesthesia achieved by inhalation of
halothane is an effective means of attaining the necessary state of analgesia.
The present study was designed to determine the effects of ketamine, the
combination of ketamine and acetylpromazine, and halothane on the adrenocortical
hormones F, DHA, and DHAS in baboons. In addition, the influence of these anesthetics on prolactin (PRL), a hormone elevated by stress [Aidara et al, 19811 and
recently shown to regulate adrenal gland function [Pepe & Albrecht, 1985b], was
assessed. Elucidation of the effects of these anesthetics on endocrine parameters has
important implications for the study of baboon reproductive physiology.
MATERIALS AND METHODS
Subjects
Six adult female baboons (Papio anubis) weighing 10-12 kg were studied at
random times of the menstrual cycle. The animals were maintained in an environmentally controlled animal facility in which food (Purina Monkey Chow 5037,
Ralston-Purina, St. Louis, MO) and fruit were provided twice daily and water was
available ad lib. Each animal was used as its own control and was treated with the
anesthetic agents described below in a randomly assigned fashion to control for
carry-over effects. A minimum of seven days elapsed between anesthetic treatments.
Procedures
Between 1000 and 1100 hours, baboons were briefly restrained in their cages for
initial sedation with 100 mg ketamine HC1 Parke, Davis & Co., Detroit, MI), which
was administered intramuscularly. Blood (5.0 ml) was immediately withdrawn from
a peripheral saphenous vein to serve as the time 0 sample. The animals were
immediately transferred to a surgical suite and subjected to one of the following
regimes for 120 minutes.
Ketamine HCl. Six animals were placed on their left side and maintained
under anesthesia with ketamine HC1 (6 mg/min; 0.40 ml/min) in 5% dextrose in
water infused with an I-Med 922 infusion pump (I-Med Corp, San Diego, CA) via a
catheter (Bard 1917R, 24 inches) inserted into the saphenous vein.
Ketamine HC1-acetylpromazine. In six baboons, 1.0 ml of acetylpromazine (10
mg/ml; Aveco Co., Inc., Fort Dodge, IA) was added to 9.0 ml ketamine (100 mg/mU,
and this solution was constantly infused via the saphenous vein at a rate of 6 mg
ketamine-0.6 mg acetylpromazine/min (0.40 ml/min) in 5% dextrose in water.
Halothane. Following ketamine sedation, four of the six baboons were placed
in a supine position and intubated with an endotrachial tube through which a
mixture of vaporized halothane (Fluothane, Ayerst Laboratories, NY) and other
gases (NzO, 25%, 1 literlmin; and 02,75%, 3 literdmin) were passed. Between 0
and 30 minutes only, 1.5%halothane and 0 2 were administered. From 30 minutes
Endocrine Response to Anesthesia in Baboons I 327
until termination of treatment, the concentration of the halothane was reduced t o
1.0% in the presence of N20 carrier.
Blood samples (5.0 ml) were collected at 5, 10, 15, 30, 45, 60, 75, 90, 105, and
120 minutes during anesthesia. Samples were kept on ice and centrifuged, and
serum was stored at -20°C until assay.
Assays
Concentrations of F, DHA, and DHAS were measured by using radioimmunoassays described previously [Pepe et al, 1977; Townsley & Pepe, 19771. The cortisol
assay had a sensitivity of 31.0 pg/ml. Intra- and interassay coefficient of variation
(CV) were assessed by using serum pools and averaged 8.0 and 9.1% respectively.
The sensitivity of the DHA assay was 16.0 pg/ml, and the intra- and interassay CV
were 8.2 and 9.8% respectively. The DHAS assay could detect 41 pg/ml with intraand interassay CV of 6.1 and 10.2% respectively.
PRL concentrations were determined by using a double antibody RIA kit developed by Diagnostic Products Corp (Los Angeles, CAI. The antiserum was generated
against porcine PRL. The ability of this antibody to detect baboon PRL was validated
by measuring serum PRL concentrations in a female baboon infused for 60 minutes
with thyroid-releasing hormone (TRH; 3 pgkg body weight). Within ten minutes of
TRH infusion, PRL concentrations rose from a baseline of 8.5 ng/ml to 108 ng/ml,
representing a tenfold increase. The PRL assay was sensitive to 5 ng/ml and the
assay is optimized for linearity throughout the 5-200 ng/ml range.
Analysis of Data
The effects of anesthetic agent on hormone concentrations were evaluated for
each hormone by using analysis of variance with repeated measures (sampling
times) and a nested variable (anesthetic). This statistical evaluation permitted determination of the main effect of anesthetic, the pattern of hormone change over time,
and the interaction between time and anesthetic. Effects were considered significant
at P < .05. Post hoc comparisons were made by using Newman-Keuls (N-K) procedure [Keppel, 19821. Data are expressed as mean SE.
RESULTS
Serum PRL Concentrations
Basal (ie, time 0) serum PRL concentrations in animals subsequently treated
with ketamine, ketamine-acetylpromaine, and halothane were 8.4 0.8,7.4 +_ 3.6,
and 7.2 + 3.6 ng/ml respectively. The administration of ketamine-acetylpromazine
elevated serum PRL t o concentrations significantly above those achieved with halothane (N-K, P < .Ol) and ketamine (N-K, P < .01; Fig. 1)alone. Treatment with
ketamine-acetylpromaine increased PRL, from a baseline of 7.4 +_ 3.6 ng/ml to a
maximum at 120 minutes of 79.5 2 18.0ng/ml, which represented a tenfold increase
in serum PRL concentrations.
Serum F Concentrations
Basal serum F concentrations (pg/lOO ml) were similar in ketamine (17.9 _+ 1.5)-,
ketamine-acetylpromaine (18.4 1.4)-,and halothane (21.9 & 2.0)-treated animals
respectively (Fig. 2). Treatment with halothane depressed F concentrations slightly, to
13.1 +_ 2.1 pg/100 ml at 120 minutes (N-K, P < .05).However, serum F was not significantly affected by ketamine or ketamine-acetylpromaine.
Serum DHA and DHAS Concentrations
Basal serum DHA concentrations were 10.6 +_ 2.3, 10.3 & 2.3, and 10.2 k 4.0
ng/ml in animals treated with ketamine, ketamine-acetylpromaine, and halothane
328 I Walker et a1
15
0
30
45
60
75
90
105
120
MINUTES
Fig. 1. Concentrations (mean 5 SE) of peripheral serum prolactin (PRL) in female baboons receiving
ketamine (0---O), ketamine-acetylpromazine (@-@),
or halothane (A-A)
anesthesia over a 120minute period.
1
0
.
1
1
15
1
1
1
1
1
1
x)
45
60
75
90
105
1
120
MINUTES
Fig. 2. Concentrations (mean k SE) of serum cortisol (F) in female baboons receiving ketamine
(0---O), ketamine-acetylpromaine (0-01, or halothane (A-A)
anesthesia over a 120-minute
period.
Endocrine Response to Anesthesia in Baboons I 329
E
)a: "
E 10-
-*
Y
--
I
n
0
15
30 45
60
75
90 105 120
MINUTES
Fig. 3. Concentrations(mean f SE) of serum dehydroepiandrosterone(DHA) in female baboons receiving
ketamine (0---O),
ketamine-acetylpromazine (0-O), or halothane (A-A) anesthesia over a 120minute period.
respectively. Serum concentrations of DHA were not significantly changed by the
anesthetic agents (Fig. 3).
Serum concentrations of DHAS are depicted in Figure 4. At time 0, baseline
DHAS concentrations in animals treated with ketamine, ketamine-acetylpromaine,
and halothane were 28.5 I 8.0, 30.2 k 6.8, and 28.5 k 11.4 pg/lOO ml respectively.
Serum concentrations of DHAS were significantly elevated (N-K, P < .01) after 75
minutes of treatment with ketamine alone and continued to rise throughout the
treatment period, reaching levels 100% above baseline. Treatment with ketamine
plus acetylpromazine appeared to increase DHAS concentrations throughout the
final 60 minutes of treatment; however, this increase was not significant. Halothane
treatment had no effect on serum DHAS concentrations.
DISCUSSION
In the present study, the effects of three commonly used anesthetics on the
serum concentrations of hormones typically measured in the study of the hypothalamic-hypophyseal-adrenocorticalaxis were determined in baboons. Serum concentrations of PRL, which characteristically are elevated by stress [Aidara et al, 19811,
were rapidly and strikingly elevated by acetylpromazine, a centrally acting dopamine receptor blocker. Because dopamine is a putative PRL-inhibitoryfactor [Meites
& Clemens, 19721, inhibition of the dopaminergic system results in increased PRL
release. A well-established relationship exists between hyperprolactinemia and altered reproductive function in which elevated PRL inhibits luteinizing hormone
release directly [Williams et al, 19791 or via diminished gonadotropin-releasing
hormone secretion [Nakano et al, 19751. Furthermore, Pepe & Albrecht [1985b]
recently have shown that PRL regulates adrenal androgen production in fetal/
neonatal baboons. Therefore, when studying endocrinological events such as menstrual cyclicity, gonadotropin secretion, regulation of the adrenal gland, or any
physiological response affected by PRL, acetylpromazine does not appear to be a
suitable anesthetic.
Ketamine alone, when administered iv continuously, had no effect on serum
PRL, F, or DHA concentrations in baboons. In contrast, others [Quadri et al, 1978;
Aidara et al, 1981;Puri et al, 19811have reported that ketamine increased PRL and
F in various cercopithecines. Methodological or species differences may account for
330 / Walker et al
0
15
30 45
60 75
90 105 120
MINUTES
Fig. 4. Concentrations (mean k SE)of serum DHA sulfate @HAS) in female baboons receiving ketamine
(0---O), ketamine-acetylpromazine (0-01, or halothane (A-A)
anesthesia over a 120-minute
period.
these apparently discrepant findings. For example, Quadri et a1 [19781 administered
a single injection of ketamine and found that PRL concentrations increased within
30 minutes but returned to baseline by 60 minutes; Puri et a1 [1981] reported that
only multiple injections of ketamine increased serum PRL concentrations. In the
present study, serum concentrations of DHAS only, the most abundant adrenal
androgen produced during baboon pregnancy [Townsley & Pepe, 1977; Walker et al,
19871 were elevated by long-term treatment with ketamine alone. This effect appears to be confined to the sulfated form of the adrenal androgen DHA and does not
appear t o be mediated by either PRL or ACTH, because neither PRL nor F concentrations were affected by ketamine treatment. Caution should be used, therefore,
when studies of the metabolism of DHAS requiring long-term sedation of the animal
are performed.
Halothane, an alternative to ketamine-acetylpromaine, decreased slightly the
serum concentrations of F, a steroid hormone that originates primarily from the
adrenal cortex; but halothane had no significant effect on DHA and DHAS. This
decline in peripheral serum adrenocortical steroid may have reflected a decrease in
production and/or an increase in the metabolic clearance rate of this steroid. Halothane produced no effect on serum concentrations of PRL in baboons, despite reports
to the contrary in rhesus monkeys [Quadri et al, 19781. Halothane administered to
immobilized animals increased PRL concentrations above those found during immobilization alone, yet PRL was also significantly elevated as a result of immobilization alone [Quadri et al, 19781. Therefore, it is possible that immobilization, and
not halothane, may have been the primary stimulant of PRL release in these
animals. Overall, the effects of halothane on the hormones examined in the present
study were relatively slight, particularly when compared to those observed with
Endocrine Response to Anesthesia in Baboons I 331
acetylpromazine.However, when coupled with reports that halothane may adversely
affect blood chemistry (eg, gases and acid-basebalance) and other regulatory mechanisms [Goosen et al, 19841, we suggest that caution should be used and results
interpreted carefully when using halothane in nonhuman primates.
In summary, ketamine alone had no influence on the serum concentrations of
PRL, F, or DHA and produced a rise in DHAS only after 75 minutes of administration. Ketamine therefore appears to be an excellent dissociative agent for the study
of hormones important in primate reproductive physiology. However, the ability of
ketamine to produce analgesia to visceral pain is suspect, and it is no longer
considered a suitable anesthetic agent for use in major surgery [Hughes & Lang,
19831. As surgical intervention is often required to study definitively the regulation
of the endocrine system, alternative anesthetic agents are required. The present
study indicates that halothane had relatively minor effects on hormones of the
adrenocortical system. It would appear that in baboons, ketamine is well suited for
the collection of biological samples when deep analgesia is not required, but that
halothane anesthesia is preferable in the latter situation.
CONCLUSIONS
1. In baboons, ketamine administration had no effect on serum PRL, DHA, or F
concentrations.
2. Long-term exposure (>60 minutes) to ketamine significantly elevated serum
concentrations of DHAS.
3. The dopamine antagonist acetylpromazine significantly elevated serum PRL
concentrations in baboons.
4. Halothane slightly decreased serum F concentrations.
5 . In baboons, ketamine was best suited for the collection of biological samples
when deep analgesia was not required, and halothane anesthesia was preferable
when deep anesthesia was necessary.
ACKNOWLEDGMENTS
The authors would like to thank Ms. Susan J. Larsen and Mr. Richard J. Roh
for their expert technical assistance. This work was supported by NIH grants R01
HD-13294 and F32 HD 06903.
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