Emeritus Faculty, Acad Council, Anesthesiology, Perioperative and Pain Medicine
Phenylethanolamine N-methyltransferase (PNMT), the final enzyme in the pathway for epinephrine biosynthesis, serves as a marker for tissues and cells producing epinephrine. The present study examines the developmental expression of PNMT in the rat embryo. A transient burst in PNMT mRNA expression begins on embryonic day 9.5 (E9.5), peaks between E10.0 and E11.0, and declines to barely detectable levels by E13.0. Regional localization of PNMT mRNA and enzyme activity demonstrates that PNMT is concentrated in the heart. PNMT has not previously been reported to be expressed at these early stages of development, and its presence in the developing heart suggests that this embryonic tissue may produce epinephrine. Because this catecholamine is known to increase cardiac output and promote the growth of cardiomyocytes, local production of epinephrine by the heart could play an important role in the development of cardiac structure and function.
View details for Web of Science ID A1996VF05900007
View details for PubMedID 8877775
We recently reported that alpha1-adrenoceptor agonists, administered at the beginning of neurulation (Stage 11a) in rat embryos grown in culture, interfere with normal development of the left/right body axis leading to situs inversus. Despite these pharmacological findings, expression of alpha1-adrenoceptor genes at such an early stage of development has not been demonstrated. In the present study, we examined the expression of mRNAs for cloned alpha1-adrenoceptor subtypes in rat embryos at Stage 11a. Timed-pregnant Sprague-Dawley rats were killed in the morning of gestational day 9 (vaginal plug day = day 0), and the implantation sites were removed. The implantation sites were separated into embryo, ectoplacental cone and decidua, only those at Stage 11a were selected, and these were immediately frozen on dry ice, and subsequently their total RNA was isolated. RNase protection assays were then performed for cloned alpha1a-, alpha 1b- and alpha1d-adrenocepter subtypes using 20-30 mug of total RNA for each hybridization. In all tissues, strong and weak signals were detected for alpha1b- and alpha1a-adrenoceptor subtype mRNAs, respectively. In contrast, a signal for alpha1d mRNA was not detected in any tissues. These results, together with previous pharmacological findings, suggest the existence of alpha1a- and alpha1b-adrenoceptor subtypes in rat embryos at Stage 11a.
View details for Web of Science ID A1995TC68900005
View details for PubMedID 20650135
In mouse and rat embryos, the embryonic disc develops within a cup-shaped "egg cylinder" and consists of an inner layer of ectoderm and an outer layer of endoderm. Because of this configuration, the embryo first develops in a dorsally flexed position and then undergoes "axial rotation" to a ventrally flexed position. In the present study, we first analyzed the morphological process of axial rotation in rat embryos using novel reference axes set in the egg cylinder that remained invariant during the process. Our new perspective allowed us to demonstrate that the process consists of three movements which start at different stages of development: twisting of the upper body at stage 12/s7-8, twisting of the middle body at stage 13/s11-12, and twisting of the lower body (so called "tail") at stage 14/s15-16. Axial rotation is an interesting developmental event not only because it is such a dynamic process but also because it is one of the earliest morphological signs of body asymmetry. This asymmetry is strongly biased in that the tail almost always finishes up on the right side of the embryo for reasons that are still unknown. In the second part of the study, we performed microsurgical experiments to extend our previous finding that removal of the allantois results in random determination of tail sidedness. We demonstrated that an allantois transplanted from another embryo can prevent this abnormal sidedness in an embryos whose allantois had been removed and that transecting the allantois did not lead to abnormal tail sidedness. A possible explanation is that the allantois produces a chemical factor that controls tail sidedness.
View details for Web of Science ID A1995QZ36200007
View details for PubMedID 7660327
Staurosporine, an alkaloid isolated from Streptomyces species, is commonly used as a protein kinase C (PKC) inhibitor in animal investigations. In the present study, we used this compound to determine whether alpha 1-adrenergic stimulation-induced situs inversus in rats is mediated by PKC. Embryos were explanted at 8 A.M. on day 9 of gestation. Those with a neural groove but with no visible neural folds (Stage 11a) were selected and were cultured in medium containing various concentrations of staurosporine with or without 50 microM of phenylephrine, an alpha 1-adrenergic agonist. At 10 A.M. on day 11 of gestation, embryos were examined for situs inversus and other abnormalities. Staurosporine, tested at 0.0001, 0.001, 0.01, 0.1, 0.375, and 0.5 microM (lethal concentration), did not block phenylephrine-induced situs inversus at any concentration. However, staurosporine alone produced situs inversus at concentrations above 0.1 microM. At 0.5 and 1.0 microM, staurosporine also caused cyst-like lesions projecting dorsally from the mesencephalon that we named "mesencephalic vesicles" and the formation of secondary somites. To confirm and further examine these unique effects of staurosporine both grossly and histologically, we conducted additional experiments using staurosporine from another source. Embryos were explanted between 6 A.M. and 9 P.M. on day 9 of gestation and were placed in one of the following groups according to their stage of development: 10b, 11a, 11b, 11c, 12/s1-2, 12/s3-4, and 12/s5-6. Embryos were then cultured with various concentrations of staurosporine. Those cultured from Stage 11a exhibited similar lesions to those seen in the initial experiment but at somewhat higher concentrations of staurosporine. Embryos cultured from Stage 10b showed a similar pattern of lesions as seen at Stage 11a, except that higher concentrations of staurosporine were required to cause mesencephalic vesicles and secondary somites formation. Embryos cultured from Stage 11b showed similar effects to those cultured from younger stages except that maximum incidences of situs inversus were much lower. Those cultured from Stage 11c showed similar dose-response to those cultured from Stage 11b except that the incidence of secondary somites formation was much higher. In addition, in approximately 40% (n = 25) of embryos treated with greater than 1.0 microM of staurosporine, the growing end of the allantois did not reach the chorion and remained unattached in the exocoelomic cavity.(ABSTRACT TRUNCATED AT 400 WORDS)
View details for Web of Science ID A1994QF41200001
View details for PubMedID 7716734
Nitrous oxide (N2O)-induced teratogenicity in rats is commonly believed to be due to decreased tetrahydrofolate, which results in decreased DNA synthesis. The role of decreased methionine has been largely ignored as have the sympathomimetic effects of N2O.A rat whole-embryo culture system was used to determine whether N2O-induced teratogenicity can be prevented with supplemental methionine or folinic acid and whether N2O-induced situs inversus is mediated by alpha 1-adrenergic stimulation. Embryos were explanted on day 9 of gestation, and those at stage 10b (late primitive streak stage) were cultured with or without N2O and the various chemicals, methionine (25 micrograms.ml-1), folinic acid (5 micrograms.ml-1), phenylephrine (range 0.5-50 microM) and prazosin (10 microM). Embryos in the N2O groups were exposed to a concentration of 75% for the first 24 h of culture. After 50 h of culture, embryos were examined for abnormalities including situs inversus.Treatment with N2O alone resulted in increased incidences of malformations and growth retardation. Methionine, but not folinic acid or prazosin, almost completely prevented N2O-induced malformations and growth retardation. N2O itself did not cause situs inversus but increased the incidence of phenylephrine-induced situs inversus. This additive effect was blocked by prazosin.Our results indicate that decreased methionine rather than decreased tetrahydrofolate plays the major role in N2O-induced teratogenicity in rats. They also indicate that N2O stimulates the alpha 1-adrenergic pathway in the embryo and thereby increases the incidence of phenylephrine-induced situs inversus.
View details for Web of Science ID A1994NW23500025
View details for PubMedID 8042787
In previous studies, we have demonstrated that stimulation of alpha 1 but not alpha 2 or beta adrenergic receptors in rat embryos grown in culture interferes with normal development of the left/right body axis leading to situs inversus. In the present study, we aimed to determine the alpha 1 adrenergic receptor subtype and signal transduction pathway involved in this phenomenon. Rat embryos at Stage 11a by a modified Theiler's staging system were cultured for 50 hr in medium containing various compounds which are known to activate or inhibit different sites of the signal transduction pathways associated with alpha 1 adrenergic receptors. They were then examined to determine the sidedness of asymmetric body structures. WB4101, a selective antagonist of alpha 1A adrenergic receptor subtype, but not chlorethylclonidine, a selective antagonist of alpha 1B adrenergic receptor subtype, inhibited phenylephrine (an alpha 1 adrenergic agonist)-induced situs inversus. Neither the protein kinase C (PKC) activators phorbol 12-myristate 13-acetate and SC-9 nor the PKC inhibitor calphostin C caused situs inversus. Furthermore, calphostin C did not block phenylephrine-induced situs inversus. A23187, a Ca2+ ionophore, induced situs inversus; nifedipine, a L-type Ca2+ channel blocker, partially blocked phenylephrine-induced situs inversus. The calmodulin antagonists trifluoperazine, W-7, and W-13 blocked phenylephrine-induced situs inversus, although they did not cause situs inversus by themselves. KN-62, a Ca2+/calmodulin-dependent protein kinase II (CaM kinase II) inhibitor, dose-dependently blocked phenylephrine-induced situs inversus. However, higher concentrations of this compound produced no block in the presence of phenylephrine and in its absence produced a 50% incidence of situs inversus. These results indicate that alpha 1 adrenergic stimulation-induced situs inversus is mediated by the alpha 1A adrenergic receptor subtype and that activation of CaM kinase II but not PKC may be involved.
View details for Web of Science ID A1994ND48700020
View details for PubMedID 8150214
Sidedness of left/right asymmetric body structures is strongly biased in most animals by mechanisms that are not well understood. In rat embryos, axial rotation starts at the 9-10 somite stage and is almost completed at the 17-18 somite stage. As a result, the ventrally flexed tail (caudal part of the body) and chorioallantoic placenta on the yolk sac take up their position normally on the right side of the embryo. Because the tail and chorion become connected via the allantois around the time when axial rotation takes place, we hypothesized that the allantois and possibly its connection to the chorion is important in determining sidedness of the tail. In the present study, we tested this hypothesis by surgically removing either the allantois or chorion before axial rotation started. Embryos were explanted at 8 AM on Day 9 of gestation (presomite stage), and either the allantois or chorion was removed using microforceps. Embryos were then cultured in rotating bottles, and sidedness of the tail, chorioallantoic placenta, and bulboventricular loop (heart) was determined after 50 hours (approximately 25-26 somite stage). Removal of the allantois (n = 55) resulted in absence of the umbilical cord and a 49.1% incidence of inverted tail; a chorioallantoic placenta-like structure developed on the yolk sac in the normal position.(ABSTRACT TRUNCATED AT 250 WORDS)
View details for Web of Science ID A1993LD38800009
View details for PubMedID 8367831
Because there is no standard developmental staging system for the early postimplantation period of rodent embryos, investigators must now choose between a variety of systems that differ significantly. We have reviewed many of these staging systems and have summarized the ambiguities within them and the inconsistencies among them. In order to compare systems, we first obtained a consensus of the order of developmental events from the literature, and then attempted to fit existing systems into this order taking into account inconsistencies in terminology and blurred borderlines between stages. We were able to do this for most systems but not all because some were too divergent. We found that inconsistencies in definition of some terms, such as "primitive streak stage" and those used to describe the early neurulation process (neural plate, neural groove, neural folds, and head fold) cause much confusion. In order to develop an unambiguous system which can be used by all investigators, we propose to modify Theiler's system, which is one of the most commonly used systems but is not defined precisely during the early postimplantation period. We suggest making subdivisions of the original stages as follows: 1) stage 8 into 8a and 8b, by the degree of extension of the proamniotic cavity into the extraembryonic region; 2) stage 10 into 10a and 10b, by the completion of amnion formation; 3) stage 11 into 11a, 11b, and 11c, by the appearance of neural folds and foregut pocket. After Stage 12, the number of somite pairs can be used to precisely stage embryos.(ABSTRACT TRUNCATED AT 250 WORDS)
View details for Web of Science ID A1992JF75400010
View details for PubMedID 1440421
Evidence of developmental toxicity of clinically used nondepolarizing muscle relaxants was sought in rat embryos grown in culture. Embryos were explanted at 8 AM on day 9 of gestation (presomite stage, plug day = day 0), and were cultured in rotating bottles with medium containing various concentrations of d-tubocurarine, pancuronium, atracurium, and vecuronium. At 10 AM on day 11 of gestation (forelimb bud stage), culture was terminated and embryos were examined for general morphology. Treatment with tested agents resulted in dose-dependent developmental toxicity; namely, growth retardation seen as decreased crown-rump length, decreased number of somite pairs, and morphologic abnormalities. However, the concentrations that caused toxicity were at least 30-fold greater than serum concentrations clinically achieved in the mother. We conclude that these muscle relaxants have a low potential for causing developmental toxicity during organogenesis.
View details for Web of Science ID A1992HY13700021
View details for PubMedID 1350890
Rat embryos at a single gestational time in the presomite period were studied for their variation in development and their fate after culture. They were explanted at 8 A.M. on day 9 of gestation from timed-pregnant Sprague-Dawley rats which were obtained by mating between 8 and 10 A.M. (plug day = day 0). In the first experiment, a total of 203 embryos from 20 litters were examined for their variation in development. Several dimensions of embryo/egg cylinder were measured and development of various embryonic/extraembryonic structures were assessed using a scoring system that we developed for the present study. Embryos were then divided into different stages of development based on their scores using the staging system that we developed previously. A large variation in developmental stage was demonstrated; the youngest embryo was at the early primitive streak stage with no signs of amniotic folds and the oldest one was at the late neural plate stage with a foregut pocket but without visible somites. No strong correlation was demonstrated between developmental stage and size of embryo/egg cylinder, nor between developmental stage and development of the proamniotic tube, ectoplacental cavity, or allantois. In the second experiment, embryos were explanted at the same time and those at different stages were cultured separately in rotating bottles and their outcomes were compared after 49 hours. The difference in mean somites number of embryos cultured from the mid primitive streak and late neural plate stages was 6.1. This difference corresponds to approximately 10 hours based on the known linear increase of somites number on day 11 of approximately 0.6 somites per hour. These results indicate a large variation in development of presomite period embryos supposedly of the same gestational age and suggest the importance of careful staging at the time of explantation if precision is needed for whole embryo culture experiments.
View details for Web of Science ID A1992HU66400010
View details for PubMedID 1412059
We recently reported that rat embryos cultured from the presomite stage in a medium containing the alpha-1 adrenergic agonist, phenylephrine, have a high incidence of situs inversus. In the present study, we have determined more precisely the critical period of development when situs inversus is induced. Rat embryos were harvested at 8 AM on Day 9 of gestation (plug day = Day 0), and divided into different stages of development, namely, early, mid, and late primitive streak stages and early, mid, and late neural plate stages. They then were cultured in rotating bottles to which phenylephrine, 0.5 mM, was added for various durations. After 49 hr of culture, embryos were examined for general morphology including sidedness of the bulboventricular loop, tail, and chorioallantoic placenta. Phenylephrine increased the incidence of situs inversus above control when administered throughout culture from either the early neural plate stage or before, and when administered for 4 hr or more from the early neural plate stage. This increase was significant even at the mid and late primitive streak stages when the control incidence was high. Our results suggest that sidedness of asymmetric body structures is determined during the early neural plate stage. This period is well before the 6-8-somite stage when morphological signs of body asymmetry first appear.
View details for Web of Science ID A1991GE31400010
View details for PubMedID 1962290
A rat whole embryo culture system was used to study the adverse reproductive effects of nitrous oxide. Embryos were removed on day 9 of gestation and exposed in culture to several concentrations of nitrous oxide for 24 h. After an additional 25 h of culture, embryos were examined for morphological abnormalities and were assayed later for protein content. Nitrous oxide resulted in growth retardation and an increased incidence of morphological abnormalities and altered body laterality. The conditions required to produce abnormalities were similar to those required in vivo. We conclude that nitrous oxide has direct toxic effects on the developing embryo and that day 9 embryo culture is a useful technique for studying mechanisms of nitrous oxide-induced teratogenicity.
View details for Web of Science ID A1991FH24100014
View details for PubMedID 2025479
In previous studies, we have shown that the reproductive toxicity of N2O in rats is prevented by the co-administration of either halothane or isoflurane, whereas treatment with folinic acid, which should reverse the effects of N2O on DNA production, does not prevent toxicity. These results cast doubt on the commonly held theory that inactivation of methionine synthase is the sole cause of N2O-induced reproductive toxicity, and suggest the need for other hypotheses. One such possibility is that N2O causes adverse reproductive toxicity secondary to its sympathomimetic effects. As a first step to test this theory, we studied the effects of phenoxybenzamine (PX), an alpha-1 adrenergic antagonist, on N2O-induced reproductive toxicity using a well-established in vivo rat model. On day 8 of gestation (plug day = day 0), 130 timed-pregnant Sprague-Dawley rats were injected s.c. with either 0.5 ml of either 0.9% saline (control and N2O alone groups) or PX (0.5, 5, or 50 micrograms/kg) in 0.9% saline, the latter the maximum tolerated PX dose. They were then exposed to either air (control) or 60% N2O for 24 hours (all other groups). On day 20 of gestation, cesarean sections were performed and the fetuses were removed and examined for either visceral of skeletal abnormalities. Compared with control, treatment with N2O alone resulted in increased incidences of fetal resorptions, major and minor visceral abnormalities, and minor skeletal abnormalities.(ABSTRACT TRUNCATED AT 250 WORDS)
View details for Web of Science ID A1991EV80500006
View details for PubMedID 2014480
Rat embryos were explanted either on day 8, 3 PM (primitive streak stage) or on day 9, 8 AM (presomite stage) and divided randomly into two groups (plug day = day 0). In one group, Reichert's membrane was removed starting from the side opposite to the ectoplacental cone which was left intact (standard method). In the second group, Reichert's membrane was removed starting from the same side as the ectoplacental cone after the latter structure was excised and the proamniotic cavity (day 8) or the ectoplacental cavity (day 9) destroyed (new method). Embryos were then cultured and examined at 10 AM on day 11. All embryos developed normally in size and general morphology. However, there was a high incidence of inverted heart, allantoic placenta and tail in all groups, except for day 9, new method group; this was the only group in which the ectoplacental cavity did not expand abnormally during culture. The new method, which is quick and easy to perform even when Reichert's membrane does not protrude well, lessens the chances of damaging embryonic ectoderm during the removal of Reichert's membrane. Furthermore, the absence of ectoplacental cone decreases embryo aggregation during culture. We hypothesize that the abnormally expanded ectoplacental cavity generates abnormal physical forces within the egg cylinder that disrupt the control of body asymmetry.
View details for Web of Science ID A1991ET41700010
View details for PubMedID 2006475
Day 11 rat embryos produced by overnight and morning short-period breeding regimens were compared for differences in interlitter variability and stage of development. Female Sprague-Dawley rats were mated either overnight (5 PM-9 AM, n = 22) or for 3 hr in the morning (8-10 AM, n = 20), and the presence of a vaginal plug was determined (day 0). At 11 AM on day 11 of gestation, rats were killed and embryos were examined for crown-rump length, somite number, and general morphology; protein content was determined later. There were no differences in mean number of implantations, live embryos, and abnormalities between the two regimens. There were significant differences in mean crown-rump length, somite number, and protein content but not in the magnitude of their variances. The difference in mean somite number was 1.2 (23.7 vs. 24.9), which corresponds to a time difference in development of approximately 2 hr. The range of mean somite number in morning short-period breeding group was 3.2 (22.2-25.4), corresponding to approximately 5.3 hr. These results are inconsistent with the popularly held hypothesis that the timing of fertilization is the major cause of interlitter variability.
View details for Web of Science ID A1990EG97900009
View details for PubMedID 2278028
Seventy timed-pregnant Sprague-Dawley rats were exposed to either air (control) or 75% nitrous oxide (N2O) for 24 hours on day 8 of gestation. Four rats from each group were killed on days 11-16, 18, and 20, and laparotomy was performed. The viability of the embryos/fetuses was determined, as was the side of tail flexion on days 11 and 12, the direction from which the umbilical artery emerged from the body on days 13 and 14, the side of the body facing the placenta on days 15 and 16, and the side to which the aortic arch curved on days 18 and 20. Mean mortality rate in the control group was 8.9 +/- 6.1% (+/- S.D.), and there were no control embryos/fetuses with altered laterality except the 9% that faced left on day 16. In contrast, N2O treatment on day 8 of gestation resulted in significantly increased mortality (40.8 +/- 3.3%) beginning on day 14 of gestation and increased incidence of altered laterality overall (31.3%) and at all stages of development. The mechanisms underlying these events remain to be defined, as do the implications of our findings for pregnant surgical patients and occupationally exposed workers.
View details for Web of Science ID A1990CM38300001
View details for PubMedID 2321158
The susceptible period of nitrous oxide (N2O) teratogenicity was studied in 170 Sprague-Dawley rats. Seven groups of 20 timed-pregnant rats were exposed to 60% N2O for 24 hours on each of days 6-12 of gestation; a control group of 30 timed-pregnant rats was exposed to air on day 9. On day 20 of gestation, dams were killed and reproductive indices were determined; their fetuses were subsequently examined for external, skeletal, and visceral abnormalities. There were no differences among the groups in the number of implantations and live fetuses, mean fetal weight, and sex ratio. The incidence of fetal wastage was higher than control in N2O-treated groups exposed on days 8 and 11 of gestation. Skeletal malformations of the ribs and vertebrae were increased following exposure on day 9 of gestation. However, the specific minor anomaly, cervical rib, was increased only following exposure on day 8 of gestation. The incidences of right-sided aortic arch and left-sided umbilical artery, abnormalities indicative of altered laterality, were increased following exposure on day 8 of gestation. Nitrous oxide administration during organogenesis causes several reproductive defects by mechanisms which remain to be determined.
View details for Web of Science ID A1989CB97200004
View details for PubMedID 2623632
A rat model was used to determine whether isoflurane exacerbates liver dysfunction and whether its metabolism is changed in the presence of cirrhosis. Male Wistar rats were gavaged weekly with carbon tetrachloride until cirrhosis was well advanced. They and control rats without pretreatment with carbon tetrachloride and without cirrhosis were then exposed to 1.45% (1 MAC) isoflurane for 3 hours. Blood and urine samples were taken before, immediately, as well as 4, 24, 48, and 72 hours after anesthesia to measure liver function and isoflurane defluorination. After the last samples had been obtained, the rats were sacrificed and the liver removed for histologic examination and in vitro metabolic studies. Serum levels of SGOT and SGPT and inorganic fluoride production in rats with cirrhosis were similar to those in control rats without cirrhosis. Concentrations of cytochromes b5 and p-450 and specific activities of microsomal defluorinase and several cytosolic enzymes were significantly lower in cirrhotic than in noncirrhotic liver, but their total amounts in whole liver were the same. The results imply that cirrhosis does not increase the risk of acute hepatotoxicity of isoflurane. They also demonstrate that metabolism of isoflurane and perhaps other volatile anesthetics may be unaffected in rats with cirrhosis, even though liver architecture is severely disrupted.
View details for Web of Science ID A1989T385700005
View details for PubMedID 2919755
A carcinogen bioassay of isoflurane was performed in groups of Swiss/Webster mice exposed to either air (n = 181), 0.1% isoflurane (n = 167), or 0.4% isoflurane (n = 165), for 4 h per day, 5 days per week. After 78 weeks of exposure, mice were left untreated for 3 weeks and were then killed. Mice killed at this time when they were 86 weeks of age, and those killed or dying at other times during the study were subjected to complete gross and microscopic examination. Throughout most of the study, mean body weights of mice exposed to 0.1% isoflurane and 0.4% isoflurane were less by 1-5% and 5-8%, respectively, than that of mice exposed to air alone. Otherwise, no gross toxic treatment effects were noted. The first neoplastic lesion was detected 23 weeks after starting treatment and, by the end of the study, 190 tumors had been detected in 179 mice. However, there were no statistical differences among the groups in the number of mice with a particular tumor at a specific site, the ratio of benign to malignant tumors, or the time to tumor appearance. It was concluded that isoflurane is unlikely to have carcinogenic potential and is a remarkably non-toxic anesthetic in mice.
View details for Web of Science ID A1988Q772100018
View details for PubMedID 3189921
The teratogenic effects of nitrous oxide (N2O) on postimplantation rat embryos were studied using a whole embryo culture system to separate the direct effects of N2O from those that are maternally mediated. A total of 100, 10-day-old rat embryos were cultured in either a control atmosphere (75% N2, 20% O2, and 5% CO2), or a N2O atmosphere (75% N2O, 20% O2, and 5% CO2). After 22 h of culture embryos were examined microscopically, and protein and DNA contents were determined. DNA content was significantly lower in the embryos exposed to N2O compared with the controls. Additionally, three malformed embryos and four embryos with left-sided tails were observed in the N2O group, whereas no abnormalities were observed in the control group. There were no differences in crown-rump length, somite numbers, limb bud index, and protein content between the two groups of embryos. The positive findings in this study indicate that whole embryo culture is useful for studying the mechanisms of N2O teratogenicity.
View details for Web of Science ID A1988P966800019
View details for PubMedID 3415019
The teratogenic effects of nitrous oxide (N2O) administered with halothane or folinic acid (FA) were studied in two separate experiments using a total of 206 timed-pregnant Sprague-Dawley rats. In each experiment, rats were exposed to either 1) air (n = 30-40); 2) N2O (50-75% for 24 h on day 8 of pregnancy, n = 20-30); 3) test agent (i.e., 0.27% halothane for 24 h on day 8 of pregnancy; or 5 mg/kg/day of FA on day 5-13 of pregnancy, subcutaneously by osmotic pump, n = 20-30); or 4) N2O + test agent (n = 20-30). Cesarean sections were performed on day 20, and fetuses were examined for visceral and skeletal abnormalities. There were no differences in pregnancy rate, number of implantations and live fetuses per rat, and fetal weight among any of the groups. Treatment with N2O resulted in significantly higher incidences of resorptions and of major visceral and minor skeletal abnormalities. Halothane administered with N2O protected against these effects; folinic acid did not. Using an additional 65 nonpregnant rats, hepatic methionine synthase activity was measured after treatment with 50% N2O, 50% N2O plus 0.27% halothane, or 50% N2O plus 5 mg/kg/day of folinic acid. Methionine synthase activity was equally depressed in all groups. These findings do not support the commonly held theory that inactivation of methionine synthase is the sole cause of N2O teratogenicity; rather, they suggest a multifactorial etiology, which may include changes in uterine blood flow.
View details for Web of Science ID A1988P542800003
View details for PubMedID 3262935
The reproductive and teratogenic effects of sufentanil and alfentanil were studied in a total of 168 Sprague-Dawley rats in two separate experiments. Either sufentanil (10, 50, or 100 micrograms.kg-1.day-1) or alfentanil (8 mg.kg-1.day-1) were administered continuously from day 5 through day 20 of pregnancy using subcutaneously implanted osmotic minipumps. Cesarean sections were performed on day 20 of pregnancy, reproductive indexes were determined, and the 1484 fetuses were examined for external, visceral, and skeletal abnormalities. No significant adverse reproductive or teratogenic effects were observed with either narcotic.
View details for Web of Science ID A1988L894400010
View details for PubMedID 2963565
The reproductive and teratogenic effects of nitrous oxide (N2O), isoflurane, and their combination were studied in 130 timed-pregnant rats. Rats were exposed to either air, 0.35% isoflurane (1/4 MAC), 50% N2O (a known teratogenic concentration), or 50% N2O plus 0.35% isoflurane for 24 h on day 8 of pregnancy. On day 20 of pregnancy, cesarean sections were performed; a total of 1268 offspring were delivered and immediately examined for external abnormalities. They were subsequently examined microscopically either for visceral or skeletal abnormalities. N2O caused significantly higher incidences of early and late resorptions, and major visceral malformations. The addition of isoflurane to N2O prevented the majority of these adverse effects. These results cast doubt on the methionine synthase inhibition theory of N2O teratogenicity.
View details for Web of Science ID A1987K989500014
View details for PubMedID 3688539
A rat model was used to determine whether the metabolism of halothane is changed in the presence of cirrhosis and whether exacerbation of liver dysfunction is correlated with such a change. Cirrhosis was produced by gavaging enzyme-induced male Wistar rats with carbon tetrachloride in corn oil once weekly for 12 weeks. Control rats received corn oil only. After a 3-week period without treatment, blood and urine were collected from each rat for determination of background levels of inorganic fluoride, bromide, and trifluoroacetic acid (halothane metabolites) and for assessment of liver function. Rats were then anesthetized with 1.05% halothane in 50% oxygen for 3 h. Following anesthesia, serial blood and urine samples were taken to monitor halothane metabolism and liver function. No differences were observed between cirrhotic and non-cirrhotic rats in serum levels and urinary excretion of halothane metabolites. However, serum levels of SGOT and SGPT were significantly increased about 1.5-fold in the noncirrhotic group and about 2.5-fold in the cirrhotic group after anesthesia. The increased levels observed in the cirrhotic group were significantly greater than in the noncirrhotic group. The results imply that the exacerbation of liver dysfunction after halothane anesthesia is most likely related to an indirect effect, such as change in liver blood flow, rather than to toxic metabolites.
View details for Web of Science ID A1987K611600008
View details for PubMedID 3674465
The reproductive and teratogenic effects of 35% and 50% nitrous oxide, fentanyl 500 micrograms kg-1/day and their combinations were studied using a total of 419 timed-pregnant Sprague-Dawley rats. On day 7 of pregnancy, osmotic minipumps which delivered either fentanyl 500 micrograms kg-1/day or normal saline for 2 weeks were implanted subcutaneously in the back of the rats. Animals then were exposed to air, 35% nitrous oxide or 50% nitrous oxide for 24 h on day 9 of pregnancy. On day 21 of pregnancy, Caesarean sections were performed and all 2693 offspring were preserved and subsequently examined microscopically for external, visceral and skeletal abnormalities. There were no reproductive or teratogenic effects observed with 35% nitrous oxide, but adverse effects were observed with 50% nitrous oxide. The addition of fentanyl to the nitrous oxide increased the mortality rate among the rats, but did not significantly add to the adverse reproductive or teratogenic effects of nitrous oxide.
View details for Web of Science ID A1987K270400018
View details for PubMedID 3676057
The effect of in vitro exposure to nitrous oxide on rat fetal and maternal methionine synthase activity was investigated. Enzyme solutions were prepared from livers of fetuses and mothers on day 19 of gestation and exposed to air, 50% oxygen or 50% nitrous oxide in oxygen for periods up to 24 h. Normal activity of methionine synthase in the fetus was about 65% of that in the mother. Activity decreased by about 25% over 24 h when the enzyme was incubated at 37 degrees C in the presence of either air or 50% oxygen. Nitrous oxide produced a time-dependent decrease in activity which generally was similar for both fetal and maternal enzyme. After 24 h exposure to nitrous oxide, activity has decreased to 14 and 17% of fetal and maternal control values, respectively.
View details for Web of Science ID A1987J438100016
View details for PubMedID 3651273
The mutagenic potential of halothane, enflurane and isoflurane in combination with nitrous oxide was investigated using the sex-linked recessive lethal assay in the fruit fly Drosophila melanogaster. Male wild-type flies were exposed for 1 h to 1 or 2% volatile anaesthetic with various concentrations of nitrous oxide up to 75%. They were then mated with untreated females of the Basc marker strain. A brooding pattern was used to assess mutagenicity at different germ cell stages. The rate of lethal mutations was assessed in the F2 generation. Halothane produced a dose-dependent increase in the rate of lethal mutations, but the mutagenicity was independent of the presence of nitrous oxide and of the stage of germ cell formation. Neither enflurane nor isoflurane was mutagenic in the presence of nitrous oxide.
View details for Web of Science ID A1987H756400018
View details for PubMedID 3111509
A carcinogen bioassay of nitrous oxide (N2O) was performed in groups of male and female Swiss-Webster mice exposed to either air (n = 179), 10% N2O (n = 152), or 40% N2O (n = 151) for 4 h per day, 5 days per week. After 78 weeks of exposure, there was a 5-week period without treatment following which surviving mice were killed. Mice killed at this time or dying in extremis at other times were subjected to complete autopsy unless advanced autolysis or cannibalism precluded examination. Mean body weights for male and female mice in the 10% N2O group were the same as those in the air control group throughout the study, whereas they were 5% less in the 40% N2O group. Mean organ weights for N2O-treated mice were not statistically different from those of control mice. Gross and microscopic examination of tissues revealed a variety of neoplastic and nonneoplastic lesions; however, their presence was unrelated to treatment.
View details for Web of Science ID A1986C527700012
View details for PubMedID 3717637
A total of 305 timed-pregnant Sprague-Dawley rats were exposed for 6 h a day on each of three consecutive days in one of three periods, i.e., pregnancy days 14-16, 11-13, or 8-10, either to 0.55 times the minimum alveolar concentration (MAC) of nitrous oxide (75%) or to 0.75 MAC of halothane (0.8%), isoflurane (1.05%) or enflurane (1.65%); an additional 232 positive-control (retinoic acid) and air control rats were studied. Reproductive indices were determined, and the 5178 offspring delivered at cesarean section were examined for external, internal, and skeletal abnormalities. There were no major or minor teratologic effects in anesthetic treated groups, although several developmental variants were observed in halothane- and enflurane-treated groups. Nitrous oxide exposure on days 14-16 resulted in a three-fold increase in fetal resorptions. The results suggest that the volatile anesthetics are not teratogenic and confirm that nitrous oxide may be associated with increased reproductive loss.
View details for Web of Science ID A1986A233400007
View details for PubMedID 3954129
Swiss Webster mice were treated to determine if subchronic intermittent exposure to the inhalation anesthetic isoflurane causes organ toxicity or enhances its own metabolism or that of other anesthetics. One-hundred twenty, four-week-old male and female mice were exposed to compressed air or to 0.02%, 0.1% or 0.5% of isoflurane for four hours per day, five days per week for nine weeks. Body weights among the groups were the same prior to exposure. Overall, there were no significant differences in body weights among exposure groups (ANOVA with day as a repeated measure: females - F = 2.12, P = 0.1085; males - F = 1.80, P = 0.1583). There was, however, a significant interaction of group and days; differences were isolated to the start of exposure (weeks 1 through 3 for females; week 2 for males). At all times, differences remained within 10% of the control body weights. Organ weights (liver, spleen, kidney, testis and uterus), hematocrits, and SGOT levels were similar among exposure groups. Histologic evaluation of organs revealed no anesthetic-related organ toxicity. The concentration of hepatic cytochromes, b5 and P-450, per mg of microsomal protein were similar among exposure groups and between sexes. The rates of hepatic microsomal metabolism (defluorination) of three volatile halogenated ether anesthetics (methoxyflurane, enflurane, and isoflurane) were not different among groups following nine weeks of exposure. Isoflurane exposures of 0.5% or less for four hours per day for five days per week would appear to be the maximum tolerated concentration for any chronic study. Since there was no evidence of organ toxicity or of enhanced or inhibited hepatic microsomal enzyme activity, isoflurane seems to be relatively non-toxic inhalation anesthetic under the conditions of this study.
View details for Web of Science ID A1986C921700013
View details for PubMedID 3712244
Swiss Webster mice were treated to determine whether the inhalational anesthetic, nitrous oxide (N2O), causes organ toxicity and enhances anesthetic defluorination. Two-hundred and sixteen young adult male and female mice were exposed to room air or to 5,000 (.5%), 50,000 (5%) or 500,000 (50%) N2O for four hours per day, five days per week for periods up to fourteen weeks. Body weight was measured twice weekly throughout the experiment. Liver, kidney, spleen and testis were weighed and examined histologically along with brain, stomach, seminal vesicle, and ovary for evidence of drug induced damage. Blood smears were examined microscopically and complete blood count, differential white cell count, and reticulocyte and platelet counts were performed. In addition, liver microsomal cytochrome P-450 content and the rates of defluorination of enflurane and methoxyflurane were determined. The maximum tolerated concentration of N2O was approximately 5000,000 ppm. Even at this high dose, there was no evidence of organ damage. Following N2O exposure, neither the hepatic microsomal cytochrome P-450 content nor the rates of anesthetic defluorination were increased; the rate of in vitro inorganic fluoride production was greater for methoxyflurane than for enflurane. Since there was no evidence of specific organ toxicity or of enzyme induction or inhibition, it was concluded that N2O is a comparatively nontoxic inhalational anesthetic under the conditions of this study.
View details for Web of Science ID A1985AXJ3500012
View details for PubMedID 4078695
Halothane, enflurane, isoflurane, and fentanyl were examined for their potential to exacerbate liver dysfunction in rats with preexisting cirrhosis. Male Wistar rats given sodium phenobarbital for 2 weeks are assigned randomly to two groups. One group (cirrhotic) was exposed by inhalation to carbon tetrachloride (CCl4) in air at weekly intervals for 12 weeks to induce cirrhosis. The other group (noncirrhotic) was handled similarly but received air only. Five weeks after the last exposure to CCl4, cirrhotic and noncirrhotic rats were given three hours of 1 MAC halothane, enflurane, or isoflurane in 50% oxygen, or 350 micrograms fentanyl per kg of body weight and 50% oxygen, or 50% oxygen only. Blood gas tensions and blood glucose levels were measured before, during, and at the end of exposure. Forty-eight hours after exposure, serum chemistries were measured in each rat for comparison with preexposure values. Rats were then killed by CO2 overdose, and liver, kidney, and testis were prepared for microscopic examination. Enflurane, isoflurane, and halothane, but not fentanyl, produced mild respiratory acidosis and no change in serum glucose levels. All anesthetics resulted in a mild but similar degree of acute liver dysfunction as indicated by small increases in SGOT or SGPT in both cirrhotic and noncirrhotic rats. Liver histology revealed mild to moderate portal cirrhosis with fibrosis and well-developed micronodules in rats exposed to CCl4, but no superimposed acute hepatocellular damage was noted. It is concluded that all the anesthetics used in this study were associated with the same minimal degree of postanesthetic hepatic dysfunction and that the dysfunction was similar in both cirrhotic and noncirrhotic rats.
View details for Web of Science ID A1985AVM8700009
View details for PubMedID 4061900
The potential effects of enflurane, isoflurane, halothane, fluroxene and nitrous oxide on mortality and fertility in Drosophila melanogaster were examined. The LD50 value 1 day after exposure for 1 h was the same for males and females and was about 8% for enflurane, isoflurane and halothane, about 20% for fluroxene, and greater than 100% for nitrous oxide. Male and female fertility, however, were not affected by any of the anaesthetics, even at concentrations well above those used clinically.
View details for Web of Science ID A1985AQM3300013
View details for PubMedID 3927955
The mutagenic effects of several inhaled anesthetic agents were investigated using the sex-linked recessive lethal assay in the fruit fly, Drosophila melanogaster. Male wild-type flies were exposed for 1 hr to either halothane, enflurane, isoflurane, or fluroxene at vapor concentrations of 1 or 2% or to nitrous oxide at concentrations of 40 or 80%. Control flies were exposed to air alone. Following treatment, male flies were mated with untreated virgin females of the Basc strain and the rate of sex-linked recessive lethals was determined in the F2 generation. Halothane and fluroxene produced a dose-dependent and statistically significant increase in the rate of sex-linked recessive lethals, whereas enflurane, isoflurane, and nitrous oxide were not mutagenic at the concentrations tested.
View details for Web of Science ID A1985ADK7100016
View details for PubMedID 2858171
The toxic and teratogenic effects of inhaled anesthetics were assessed in an in vivo assay using the fruit fly Drosophila melanogaster. Eggs were exposed during development (metamorphosis) to enflurane, isoflurane or halothane at a vapor concentration of 0.1 or 0.2% (v/v), to fluroxene at 0.025 or 0.05% (v/v), or to nitrous oxide at 20 or 40% (v/v). Flies produced in each group were counted and were examined for morphological abnormalities within one day of hatching. All the anesthetics except nitrous oxide produced a dose-dependent increase in the duration of metamorphosis and a decrease in the number of flies. Despite these effects on development, no morphological abnormalities were observed in any fly.
View details for Web of Science ID A1985ALJ3300008
View details for PubMedID 3925599
The authors have refined a model of cirrhosis in the rat and used it to determine whether the administration of halothane anesthesia adversely affects preexisting liver disease. Male Wistar rats were placed on phenobarbital water and were assigned randomly to two groups. Group 1 rats were exposed by inhalation to carbon tetrachloride (CC14) at weekly intervals for 12 exposures while Group 2 rats received only air. All treatment including phenobarbital then was withdrawn for 4 weeks. Rats then were bled for SGOT and SGPT determinations and 24 h later were exposed to 1.8% halothane in oxygen for 3 h (HAL); the remaining rats from each group were exposed to 100% oxygen for 3 h (O2). Twenty-four hours later, rats were killed and blood was obtained for SGOT and SGPT by cardiac puncture. Light microscopic histologic examination was performed blind on liver sections for cirrhosis and scored for superimposed acute focal necrosis. The weekly sublethal CCl4 exposure resulted in histologically demonstrable cirrhosis in all surviving Group 1 animals. The mean (+/- SD) SGOT (128 +/- 32 IU/1) and SGPT (86 +/- 24 IU/1) values for the Group 1 rats were significantly greater (P less than 0.01) than those for Group 2 rats (98 +/- 18 IU/1 and 57 +/- 12 IU/1, respectively). Cirrhotic animals showed neither deterioration in liver function nor acute liver cell necrosis after Hal compared with O2. However, Group 2 rats showed a modest but significant increase in SGOT (P less than 0.05) after HAL, while this change was not noted after O2. Thus, 1.8% halothane anesthesia in oxygen did not result in superimposition of acute liver cell injury in already cirrhotic rats.
View details for Web of Science ID A1985AAQ6500001
View details for PubMedID 3966663
The MAC of halothane, isoflurane, and enflurane was determined using the tail-clamp technique in pregnant female, nonpregnant female, and male Swiss Webster mice (n = 216) and Sprague-Dawley rats (n = 112). Mean MAC values (+/-SD) for halothane, isoflurane, and enflurane in mice were 0.95 +/- 0.07%, 1.34 +/- 0.10%, and 1.95 +/- 0.16%, respectively; values in rats were 1.03 +/- 0.04%, 1.46 +/- 0.06%, and 2.21 +/- 0.08%, respectively, all significantly higher than in mice. Neither the sex of the animals nor whether female animals were pregnant influenced the results.
View details for Web of Science ID A1985ADK7100021
View details for PubMedID 3977116
The developmental toxicity of trace (0.006%), subanesthetic (0.06%), and light anesthetic (0.6%) exposure to isoflurane was examined in Swiss/Webster mice. No adverse effects were demonstrated following exposure of dams to 0.006% (n = 26) and 0.06% (n = 27) isoflurane for 4 hr daily on days 6-15 of pregnancy. Exposure to 0.6% isoflurane (n = 23) for the same period resulted in significantly decreased fetal weight, decreased skeletal ossification, minor hydronephrosis, and increased renal pelvic cavitation. The incidence of cleft palate also was significantly increased, abnormalities occurring in 12.1% of fetuses and affecting 11 of 23 litters. This incidence was considerably higher than that of the combined treatment and colony control groups (0.75%) and those that we have found in previous experiments with this mouse strain following exposure to halothane (1.2%) or enflurane (1.9%).
View details for Web of Science ID A1985AVY0500002
View details for PubMedID 4082064
The activity of the enzyme methionine synthase in fetal and maternal liver was investigated before and after exposure to nitrous oxide. Timed-pregnant rats on day 19 of gestation were exposed to either 10% or 50% nitrous oxide for various times up to 240 min. After exposure, fetal and maternal livers were removed and methionine synthase activity assayed. Normal methionine synthase activity in the fetus was about 50% of that in the mother. Both fetal and maternal methionine synthase activity decreased progressively with increasing time of exposure to nitrous oxide and recovered only slowly after the agent was discontinued.
View details for Web of Science ID A1984SV26900013
View details for PubMedID 6721960
Methionine synthetase (MS) activity in the brain and liver is decreased following nitrous oxide (N2O) exposure. Since MS is important for DNA synthesis, this interaction would be expected to have the most serious consequences on actively replicating tissue. The authors therefore measured MS activity in rat testes following 1 h exposure to either 10% or 50% N2O. Animals exposed to 10% N2O had a 29% reduction in MS activity, and exposure to 50% N2O caused a 63% reduction in enzyme activity. Testicular MS activity returned to normal by 24-48 h in the 10% exposure group and by 72 h in the 50% exposure group. This biochemical effect on testicular enzyme activity could be the basis for the reported deleterious effects of N2O on spermatogenesis.
View details for Web of Science ID A1984TA88800011
View details for PubMedID 6742486
The effects of 24 hours of nitrous oxide exposure on reproductive indices and fetal development were examined in Sprague-Dawley rats. Four different experiments employing four concentrations of nitrous oxide--0.75%, 7.5%, 25% and 75%--established that the threshold of toxicity was greater than 25%. At 75% nitrous oxide there was a significant increase in early and late resorptions, and a consistent teratogenic effect (e.g., runts, ocular malformations, limb deformities). Neither the stress of shipping dams while pregnant nor the withholding of food during nitrous oxide exposure resulted in additional adverse effects. Exposure to 25% nitrous oxide was associated with increased deoxyuridine suppression values; however, adverse reproductive effects were not seen at this nitrous oxide concentration. The results of this and other studies which have examined the reproductive and teratogenic effects of nitrous oxide do not contraindicate its use in operating rooms nor, when necessary, as an anesthetic for pregnant surgical patients.
View details for Web of Science ID A1984TP21700012
View details for PubMedID 6495226
The dose-dependent effects of nitrous oxide on thymidine and methionine syntheses were investigated in pregnant rats. Female Sprague-Dawley rats were exposed on day 9 of gestation to 0.75%, 7.5%, or 75% nitrous oxide for 24 h. Immediately and 72 h after exposure, a deoxyuridine-suppression test was performed on maternal bone marrow and a methionine synthetase assay was performed on maternal liver to assess thymidine and methionine syntheses, respectively. Inhibition of thymidine synthesis was seen after exposure to 7.5% and 75%, but not after 0.75%, nitrous oxide. Recovery was complete 72 h after exposure. Methionine synthetase activity was abolished at all concentrations of nitrous oxide tested and did not return to control values 72 h after exposure. Fetal weight and gross appearance were not affected by exposure to nitrous oxide; however, the observed decrease in thymidine and methionine syntheses after nitrous oxide exposure may account for its teratogenic effects.
View details for Web of Science ID A1983RA17600006
View details for PubMedID 6869860
Male and female Swiss Webster (SW) mice, age 13 to 14 weeks, were exposed by inhalation for 4 hr per day, 5 days per week, for 14 weeks, to either room air, 0.5% nitrous oxide, 5.0% nitrous oxide, or 50% nitrous oxide. Murine germ cells were examined for evidence of injury after this exposure. A group of male mice were treated with methyl methanesulfonate (MMS) as a positive control for sperm abnormalities while a group of female mice were treated with 3-methylcholanthrene (3-MC) as a positive control for oocyte destruction. There were no significant differences among the four inhalation exposure groups in testes weight, percentage of abnormally shaped sperm, sperm count, or histologic appearance of the testes; the mean percentage (+/- SE) of abnormal sperm ranged from 8.9 +/- 2.4 (5.0% nitrous oxide) to 13.5 +/- 0.5 (50% nitrous oxide) with a concurrent control value of 10.4 +/- 2.3%. In the positive control experiment, 25.2 +/- 4.1% of sperm from mice treated with MMS were abnormal compared with 2.5 +/- 0.3% of sperm from mice treated with saline (p less than 0.001), indicating that sperm of SW mice are sensitive to chemical damage. There was no significant difference between the mean number of oocytes in mice treated with 50% nitrous oxide (33.3 +/- 14.4) and in control mice (29.8 +/- 8.0). In the positive control experiment, mice treated with 3-MC had significantly fewer (p less than 0.001) primordial oocytes, 67.2 +/- 19.5 compared with control mice, 222.4 +/- 21.9, indicating that this strain is sensitive to chemical damage of the ovary. Thus, murine germ cells showed no evidence of toxic effects due to prolonged exposure to nitrous oxide.
View details for Web of Science ID A1983QM41200008
View details for PubMedID 6845366
Enflurane has been shown to increase slightly the percentage of abnormal spermatozoa in mice. To examine in more detail possible adverse effects of enflurane on male reproductive organs, 125 Swiss/ICR mice were exposed to 0.3% enflurane for up to 18 months; a control group of 125 mice was exposed concurrently to air. Exposures were for 4 hours/day, 5 days/week. After 12 months of exposure, groups of 25 enflurane-treated and control mice were killed, and chromosomal analysis of spermatogonial cells was performed. After 18 months exposure, remaining mice were killed and epididymal sperm were collected for morphologic examination. In addition, the reproductive organs of all mice dying or killed throughout the study were examined histologically. Approximately 1% of spermatogonial cells from both enflurane-treated and control mice had aberrant chromosomes. The average percentage of morphologically abnormal sperm was 9.4 +/- 1.7 for enflurane-treated mice and 6.5 +/- 1.3 for control mice; the difference was not statistically significant (p = 0.18). Finally, there was no difference between the treated and control mice in histologic appearance of reproductive organs. It was concluded that long-term exposure to enflurane has no adverse effects on male reproductive organs in mice.
View details for Web of Science ID A1982MZ36700005
View details for PubMedID 7198408
The effects of exposure to nitrous oxide on reproductive indices, fetal development, and male fertility were examined in Swiss/ICR mice. In experiment I, female mice were exposed for 4 hours per day on days 6-15 of pregnancy, to 0.5% (5,000 ppm), 5.0% (50,000 ppm), or 50% (500,000 ppm) nitrous oxide. Control mice were untreated, exposed to compressed air, or treated with retinoic acid on day 8 of gestation. In experiment II, male mice were treated, as above, for 9 weeks and then mated nightly for 7 nights to untreated, virgin females. In experiment I, 1,761 fetuses from 154 dams were examined and found to be without evidence of adverse nitrous oxide treatment effects. In experiment II there were no differences among the groups in the ability of males to impregnate females or in litter size, fetal wastage, or fetal size. When we compare nitrous oxide with other inhalation anesthetics we have studied employing a similar protocol, we find the order of reproductive toxicity to be: halothane greater than enflurane greater than methoxyflurane greater than nitrous oxide. None of the agents were toxic, however, at the trace concentrations usually found in operating rooms.
View details for Web of Science ID A1982PF03400002
View details for PubMedID 7135253
Inorganic fluoride (F-) production and renal function were assessed in six groups of Fischer 344 rats administered either methoxyflurane (MOF) or deuterated methoxyflurane (d4-MOF). One untreated and one phenobarbital (PB)-treated group were exposed for two hours to either air, 0.5 per cent (V/v) MOF, or 0.5 per cent (v/v) d4-MOF. Serum and urinary F- and serum urea nitrogen and creatinine were measured. Urine volume and urinary F- excretion were only slightly greater among MOF than among d4-MOF exposed animals. Pretreatment with PB, however, greatly enhanced F- production in MOF-exposed animals leading to marked renal impairment but only slightly enhanced F- production in d4-MOF animals leading to mild renal impairment. Thus, only in PB-pretreated animals could a biologically significant difference in nephrotoxicity be demonstrated for MOF and d4-MOF.
View details for Web of Science ID A1982NE81600009
View details for PubMedID 7059030
A modification of the Ames bacterial assay system employing two histidine-dependent strains of Salmonella typhimurium, TA1535 and TA100, was used to test the mutagenicity of four experimental, inhalational anesthetic agents: sevoflurane, synthane, dioxychlorane, and dioxyflurane. None of the anesthetics was mutagenic. Increased activity was seen only with vinylidene chloride, the positive control.
View details for Web of Science ID A1982NY17500010
View details for PubMedID 7044187
A carcinogen bioassay of enflurane was performed in Swiss/ICR mice. Two groups of weanling mice, each of 125 males and 125 females, were exposed to eigher a maximum tolerated dose of enflurane, 3000 ppm (0.3 per cent v/v) or compressed air for 4 hs per day, 5 days per week. After 52 weeks exposure, 25 males and 25 females from each group were killed. After 78 weeks exposure, a 4-week period without treatment was allowed before the remaining mice were killed. Mice killed at the scheduled periods and those killed or dying at other times throughout the study, underwent extensive gross and histologic, examinations unless autolysis or cannibalism precluded examination. After 78 weeks exposure, male mice in the enflurane-treated group had a 36 per cent incidence of liver tumors compared with 24 per cent in the control group; however, the difference was no statistically significant. The incidence of lung tumors was about 20 per cent in all groups. Other neoplastic lesions occurred in small numbers and were unrelated to treatment. It was concluded that under the conditions of the present experiment, chronic administration of enflurane at its maximum tolerated dose did not lead to an increased incidence of neoplasia in Swiss/ICR mice.
View details for Web of Science ID A1982MY12800003
View details for PubMedID 7053680
The mutagenic and toxic effect of high helium and hydrostatic pressure were assessed in an in vitro microbial assay using two histidine-dependent strains of Salmonella typhimurium. TA98 and TA100. Bacteria in the presence or absence of mammalian enzyme system were subjected to either 50 or 100 atmospheres absolute (ATA) helium or 300 ATA hydrostatic pressure. The number of revertant colonies, colony forming units and cell growth rates were measured. There was no mutagenic effect of decrease in number of colony forming units at any of the pressures tested. However, a statistically significant decrease in growth rate of exponentially dividing cells was seen at 300 ATA hydrostatic pressure and 100 ATA helium, but not at 50 ATA helium.
View details for Web of Science ID A1981MM87500002
View details for PubMedID 7031991
The mutagenic and toxic potential of nitrous oxide were assessed in vitro by microbial assay using two histidine dependent strains of Salmonella typhimurium, TA98 and TA100. Bacteria on plates and in liquid suspension in the presence or absence of enzymes prepared from rat liver, were exposed in a pressure chamber to partial pressures of nitrous oxide ranging from 0.5 to 6 atmospheres. Nitrous oxide decreased viability of both strains of bacteria at 4 and 6 atmospheres but was not mutagenic at any pressure tested.
View details for Web of Science ID A1981LA87800013
View details for PubMedID 7013158
Many vinyl compounds, such as vinyl chloride and some inhalational anesthetics, are known to be mutagens. In the present study, 10 vinyl compounds or derived epoxides, widely used in industry, were assayed in the Salmonella typhimurium/mammalian microsome system. 3 strains of histidine-dependent S. typhimurium, TA1535, TA98 and TA100 were used. Of the 10 compounds, 4 were mutagens. They were 9-vinylanthracene, vinylcarbazole, 3-vinyl-7-oxabicyclo[4.1.0]heptane and 3-epoxyethyl-7-oxabicyclo[4.1.0]-heptane. The study confirmed the overall genotoxicity of vinyl compounds and epoxides and the need to carefully screen them for mutagenic/carcinogenic effects.
View details for Web of Science ID A1980JZ52500003
View details for PubMedID 7001215
Mutagenic activity of urines obtained from operating room personnel was assayed in the Ames Salmonella/mammalian microsome system using three strains of histidine-dependent S. typhimurium, TA1535, TA1538, and TA100. Two procedures were employed. In the first, 100- and 200-microliter aliquots of urine obtained from 28 subjects working in either scavenged or unscavenged operating rooms were tested. In the second, urine samples obtained from 13 physicians before and after starting an anesthesia residency, as well as 250-fold concentrates of these samples, were assayed. There was no statistically significant difference in urinary mutagenic activities between individuals working in scavenged and those working in unscavenged operating rooms. Furthermore, urines of anesthesiologists collected before and after beginning training had similar mutagenic activities. Only heavy smokers had mutagenic urine. It was concluded that the majority of operating room workers do not excrete mutagens in the urine.
View details for Web of Science ID A1980KG96700003
View details for PubMedID 7425332
Swiss/ICR mice were tested to determine whether the volatile ether anesthetic, enflurane, causes induction of anesthetic defluorination and organ toxicity. Mice were exposed in utero and postnatally to 0.01, 0.1 and 1.0 volumes percent enflurane vapor. Body weight was measured at frequent intervals throughout the experiment. Animals were sacrificed at 73 days of age and liver microsomal cytochrome P-450 content and the rate of defluorination of enflurane, isoflurane, methoxyflurane ans sevoflurane were determined. In addition, the liver, kidney and testis were weighed and examined histologically for drug induced damage. The maximum tolerated dose of enflurane delivered over a twelve week period was determined to be 0.5 volumes percent for four hours a day, five days a week. Even at this high dose there was no evidence in either sex of liver, kidney or testicular damage. Following enflurane exposure, neither the liver microsomal cytochrome P-450 content nor the rate of anesthetic defluorination was increased. The rate of in vitro inorganic fluoride production per unit time was greatest for methoxyflurane, and approximately equal for enflurane, isoflurane and sevoflurane. Since there was no evidence of enzyme induction or specific organ toxicity, it was concluded that enflurane is a comparatively nontoxic volatile anesthetic under conditions of this study.
View details for Web of Science ID A1980KR74100026
View details for PubMedID 7441115
Male and female Swiss/ICR mice were exposed to 0.3% enflurane in air, 4 h per day, 5 days per week for 52 weeks. A control group was exposed to air alone. After 52 weeks, all animals were sacrificed and peripheral blood and bone marrow samples were examined for alterations in haemopoiesis. In general, there was no difference between treated and control groups.
View details for Web of Science ID A1980JS80100002
View details for PubMedID 7387801
The mutagenicities of two volatile metabolites of halothane, 2-chloro-1,1,1-trifluoroethane (CF3CH2Cl) and 2-chloro-1,1-difluoro-ethylene (CF2CHCl), and a presumed halothane metabolite, 2-bromo-2-chloro-1,1-difluoroethylene (CF2CBrCl), were investigated in the bacterial assay system developed by Ames and co-workers. Both gas-phase and liquid culture tests were made. In addition, mutagenicity of CF2CBrCl was studied in a modified Ames test system using rapidly growing cells in enriched liquid medium. The purity of tested compounds was verified by combined gas chromatography and mass spectrometry. In the standard Ames test, CF3CH2Cl and CF2CBrCl were not mutagenic, and CF2CHCl was only weakly mutagenic. CF2CBrCl was detectable as weakly mutagenic, however, in the modified Ames test using rapidly growing cells. Although these results are reassuring, the effects of long-term exposure to halogenated anesthetics are still not fully known.
View details for Web of Science ID A1979HT20900011
View details for PubMedID 386857
The teratogenic potential of subanesthetic and anesthetic exposure to halothane was studied in Swiss/ICR mice. Two treatment regimens were employed: daily exposure of males and females for nine weeks prior to conception and on days 1 through 17 of pregnancy; and exposure of females only on days 6 through 15 of pregnancy. Mice were exposed to subanesthetic concentrations of halothane for 0.025, 0.1, 0.4, and 1.2 MAC hours/day; anesthetic exposure was 4.0 MAC hours/day. Fetal morphologic development was normal at the two lowest exposures. Exposures of 0.4 MAC hours/day and more were associated with decreased fetal ossification. At the 1.2 MAC hour/day exposure, renal pelvic masturation was retarded and the incidence of skeletal variants was increased. The incidences of major malformations and minor anomalies were not increased following exposure to subanesthetic concentrations of halothane. Anesthetic exposure to 4.0 MAC hours/day was lethal to both dams and embryos, and resulted in major developmental malformations in surviving fetuses. These effects were probably due to altered maternal physiologic status. It is concluded that exposure of mice to subanesthetic concentrations of halothane does not result in important morphologic abnormalities in their offspring.
View details for Web of Science ID A1979JB04500010
View details for PubMedID 517780
The mutagenic potential of trichloroethylene, divinyl ether, nitrous oxide and cyclopropane was assessed in vitro by microbial assay employing two histidine-dependent strains of Salmonella typhimurium, TA1535 and TA100. Anaesthetic agents in various concentrations were incubated with bacteria in the presence or absence of an enzyme system prepared from enzyme-induced rat liver. Nitrous oxide and cyclopropane were not mutagenic, whereas divinyl ether gave a strongly positive response. Results for trichloroethylene were equivocal. These and previous studies with the salmonella system, together with mutagenicity studies using different test systems, indicate that modern inhalation anaesthetic agents are unlikely to be mutagenic.
View details for Web of Science ID A1979GW23200005
View details for PubMedID 375954
A simplified in-vivo bioassay system was used to test the carcinogenic potential of halothane in Swiss/ICR mice. Halothane was tested only at its maximum tolerated dose, and histologic examination was performed only on tumor masses and other grossly abnormal tissues found at necropsy. Two groups, each of 15 timed pregnant mice, were exposed to either halothane, 500 ppm (0.05 per cent), or compressed air for two hours on days 10--19 of pregnancy. Five days after birth the offspring were similarly exposed, three times weekly, for 78 weeks. After a ten-week, no-treatment, observation period, all remaining mice were examined by necropsy. Mice dying or killed in extremis before final sacrifice at 88 weeks of age also underwent complete gross necropsy unless extensive cannibalism or autolysis precluded examination. The incidences of malignant tumors, hepatomas or modular hyperplasias, and benign tumors in halothane-treated mice were 7, 6, and 20 per cent, respectively; there were similar incidences of these lesions in control animals. It is concluded that under the conditions of this experiment, lifetime administration of halothane at its maximum tolerated dose is not associated with an increased incidence of neoplasia in Swiss/ICR mice.
View details for Web of Science ID A1979HB75200005
View details for PubMedID 222171
The commercially available volatile anesthetic fluroxene (2,2,2-trifluoroethyl vinyl ether) which contains the stabilizer N-phenyl-1-napthylamine, was tested for mutagenicity using four strains of S. typhimurium, TA1535, TA1537, TA98 and TA100, and one strain of E. coli, WP2. In addition, purified fluroxene; N-phenyl-1-napthylamine; trifluoroethanol, a major metabolite of fluoroxene; and urine from rats anesthetized with fluroxene were tested. Several procedures were utilized including exposure of bacteria to vapor in desiccators and in liquid suspension. Results indicate that fluroxene, but not its stabilizer, was mutagenic to strains TA1535, TA100 and WP2 only in liquid suspension and only in the presence of a rat-liver enzyme system. Trifluoroethanol and urine from fluroxene-treated rat were not mutagenic to any strain of bacteria. These findings indicate that fluroxene is a promutagen which requires preincubation before it is recognized. Further experiments were performed with enzymes prepared from mouse, hamster and human liver. Fluroxene was mutagenic only in the presence of enzymes prepared from Aroclor 1254 pretreated rodents. Since fluroxene was not mutagenic in the presence of enzymes prepared from three human livers, the significance of these findings to man are unclear.
View details for Web of Science ID A1978GA33400008
View details for PubMedID 34096
Reproductive studies were performed in Swiss/ICR mice chronically exposed to subanesthetic and anesthetic concentrations of halothane. Male and female mice were treated five or seven days a week for nine weeks prior to mating; exposure of females was continued daily throughout pregnancy. Halothane exposures were 0.025, 0.1, 0.4, 1.2, and 4.0 MAC hours per day. No adverse effect on reproduction was observed at the lowest two exposure levels studied. Exposures to 0.4 MAC hour per day or more were associated with decreased maternal weight gain, fetal fetal length and weight, and early postnatal weight gain. Pregnancy rate, implantation rate, and number of live fetuses per litter were significantly decreased at 1.2 MAC hours per day. The percentage of resorption or fetuses dead in utero was not increased, and postnatal survival of offspring was unaltered. Subsequent matings between untreated females and males exposed to halothane, 1.2 MAC hours per day for 17 weeks, resulted in normal reproductive performance; this suggests that the adverse reproductive changes observed when both males and females were exposed represented a primary effect on females. The least exposure at which effects were seen is approximately 40 times greater than the level of human occupational exposure is unscavenged operating rooms.
View details for Web of Science ID A1978ER29300002
View details for PubMedID 626421
An in vitro microbial assay system employing two histidine-dependent strains of Salmonella typhimurium, TA1535 and TA100, was used to test the mutagenicities of enflurane, methoxyflurane, isoflurane and fluroxene. Enflurane, isoflurane and fluroxene in concentrations ranging from 0.01 to 30 per cent and methoxyflurane in concentrations ranging from 0.01 to 7 per cent were incubated with bacteria in the presence or absence of homogenates of liver prepared from rats pretreated with the enzyme inducer, Aroclor 1254. Enflurane, methoxyflurane, isoflurane, and urines from patients anesthetized with these agents were not mutagenic. Fluroxene, however, was highly mutagenic, and therefore poses a possible hazard for operating room personnel and patients.
View details for Web of Science ID A1977DD60500010
View details for PubMedID 15474
The mutagenicity of halothane was tested in an in-vitro microbial assay system employing two histidine-dependent mutants of Salmonella typhimurium, TA98 and TA100, Halothane in concentrations ranging from 0.1 to 30 per cent was incubated with bacteria in the presence or absence of a metabolic activation system prepared from either rat liver treated with Aroclor 1254 or human liver. Trifluoroacetic acid, a major metabolite of halothane, and urine from patients anesthetized with halothane also were tested. Halothane, trifluoroacetic acid, and patients' urines were not mutagenic.
View details for Web of Science ID A1976CD56100007
View details for PubMedID 786079