Cocaine as a cause of congenital malformations of vascular origin: experimental evidence in the rat

Cocaine hydrochloride was administered to pregnant Sprague-Dawley rats as a single intraperitoneal dose or as two doses 1-4 hours apart. A single dose administered on day 16 of gestation was teratogenic in a dose-dependent manner, with 40 mg/kg being a no-effect dose and 50 mg/kg the lowest teratogenic dose; 80 mg/kg was lethal to the dam. Forty-eight hours after exposure to a teratogenic dose on day 16 of pregnancy, the fetuses showed severe hemorrhage and edema in the their extremities, particularly the footplates, tail, genital tubercle, and upper lip/nose. When the fetuses were examined on day 21 of gestation, the main externally visible malformations were reduction deformities of the limbs and tail. When two doses of cocaine were administered 1-4 hours apart, the incidence of affected fetuses increased as the time interval between the two doses decreased. Two doses of cocaine administered 2 hours apart were not teratogenic on day 9, 10, 11, 12, 13, or 14 of gestation but did induce reduction deformities on days 15, 16, 17, 18, or 19. The same dose administered 1 hour apart was teratogenic on days 14-19. In general, cocaine administration on gestational days 14, 15, or 16 induced more severe and more widespread hemorrhage and edema than administration on days 17, 18, or 19. In the latter cases, damage was restricted to the distal parts of the hindlimb digits and the tail. The results show that in the rat cocaine is only teratogenic during the late organogenic or postorganogenic period.(ABSTRACT TRUNCATED AT 250 WORDS)     

Cerebrovascular effects of prostanoids: in-vitro studies in feline middle cerebral and basilar artery

The effect of prostaglandin (PG) E2, F2 alpha, the thromboxane-A2 mimetic U46619 (9,11-dideoxy-9 alpha,11 alpha-methanoepoxy-prostaglandin F2 alpha) and the prostacyclin mimetic iloprost was investigated in cat middle cerebral and basilar arteries in vitro precontracted with 5-hydroxytryptamine (5-HT) (50nM) in the absence and presence of the cyclooxygenase inhibitor indomethacin or the thromboxane receptor blocker AH23848B [1 alpha (z),2 beta,5 alpha]-(+)-7-[5-[1,1'-(biphenyl)-4-yl] methoxy]-2-(4-morpholinyl)-3-oxocyclopentyl]-4-heptenoic acid). PGF2 alpha and U46619 both produced further concentration-related contractions of basilar and middle cerebral artery, U46619 being approximately 1,000 times more potent than PGF2 alpha. Iloprost produced concentration-related relaxations of precontracted basilar and middle cerebral artery, the mean maximum relaxations produced at a concentration of 1.3 microM being 57.3% and 80.6%, respectively of the contraction produced by 50nM 5-HT. PGE2, 100nM relaxed the basilar and middle cerebral artery, 46.7% and 38.5% respectively. However, at 1 microM, PGE2 caused contraction. Indomethacin, 2.8 microM had no effect on contractile or relaxant responses to any of the prostanoids. Oxyhaemoglobin inhibited the relaxation of both arterial preparations but had no effect on responses to PGE2 or iloprost. The thromboxane-receptor blocker AH23848B antagonised the contractile responses to U46619, PGF2 alpha and PGE2 and had no effect against relaxant responses to PGE2 or iloprost. It is concluded that both contraction- and relaxation-inducing prostanoid receptors are present in the in vitro preparation of feline basilar and middle cerebral artery. Under sustained tension conditions, endothelial factors do not appear to be involved in the responses to dilating prostanoids.     

Effect of alpha-adrenergic blocking agents on uterine activity in the ewe at the end of gestation

The electromyographic activity (EMG) of the uterus was recorded in vivo in 8 unanaesthetized ewes from the 140th day of gestation up to parturition. The effects on uterine activity of treatments with an alpha 1-receptor blocker (prazosin) and an alpha 2-receptor blocker (yohimbine) were studied. During the last days of gestation, EMG activity consisted of periodic active phases (1-2/h). During the last 16-17 hours, uterine activity increased sharply; this period was referred to a labour. Intravenous perfusion of prazosin (0.03 mg/kg/mn over 1 h) or intravenous injections (1 mg/kg) did not modify uterine activity either before or during labour. Intravenous perfusion of yohimbine (0.03 mg/kg/mn during 1h) inhibited uterine activity before and during labour. In all cases, lambing occurred between the 142nd and 145th day of gestation, which corresponds to the normal lambing period. These results suggest that, in the ewe, uterine alpha 2-receptors are important for normal uterine activity at the end of gestation and in. parturition.     

Cardiopathic effects of dichloroacetate in the fetal Long-Evans rat.

Dichloroacetic acid (DCA) is a by-product of the chlorine disinfection of water and may occur in treated water at levels exceeding 100 micrograms/L. Previous studies revealed teratogenic effects, particularly heart malformations, at high doses (900-2,400 mg/kg given on days 6-15 of pregnancy). In a series of three studies, groups of 7-10 Long-Evans rats were dosed with 1,900 mg/kg of DCA on days 6-8, 9-11, or 12-15; with 2,400 mg/kg on days 10, 11, 12, or 13; and with 3,500 mg/kg on days 9, 10, 11, 12, or 13, in an attempt to determine the most sensitive period and further characterize the heart defect. In a fourth study, six dams were treated with 1,900 mg/kg of DCA days 6-15 of pregnancy, and 56 fetuses were harvested for light microscopy of the heart. Eight control fetuses from four litters were also examined. No heart malformations were seen in the groups treated with 1,900 mg/kg DCA days 6-8 but were present in the group treated on days 9-11 and 12-15, with the higher incidence occurring on days 12-15. Single doses of 2,400 mg/kg DCA given on days 10, 11, 12, or 13 resulted in a much lower incidence of cardiac malformations, which occurred only on days 10 and 12. The high dose of DCA (3,500 mg/kg) did not increase the incidence of heart defects but showed that dosing on day 9 as well as on days 10 and 12 would produce the defect. The defects seen were characterized as high interventricular septal defects (H-IVSD). Light microscopy showed that the defect was caudal to the semilunar valves, with the anterior right wall of the aorta communicating with the right ventricle. Another aspect of the defect is at the level of the semilunar valves, with the right cusp or sinus of Valsalva in communication with the right ventricle. The defects are discussed more fully and methods for further study suggested.     

Developmental toxicity evaluation of orthovanadate in the mouse

Sodium orthovanadate in deionized water was administered once daily by gavage on gestational days 6-15 to mice at doses of 0, 7.5, 15, 30, and 60 mg/kg. Dams were killed on day 18 of pregnancy, and fetuses were examined for external, visceral, and skeletal defects. Maternal toxicity was observed at the highest doses of sodium orthovanadate, as evidenced by a significant number of deaths (60 and 30 mg/kg/d) and reduced weight gain and food consumption (30 and 15 mg/kg/d). Embryolethality and teratogenicity were not observed at maternally toxic doses and below, but fetal toxicity was evidenced by a significant delay in the ossification process of some skeletal districts at 30 mg/kg/d. The no-observed-adverse-effect level (NOAEL) for maternal toxicity was 7.5 mg/kg/d, and 15 mg/kg/d represented a NOAEL for developmental toxicity in mice under the conditions of this study.     

Thromboxane receptors can modulate gastric acid secretion in the rat

The effects of PGE2 and the thromboxane A2 mimetic, U-46619, have been investigated on gastric secretion in the rat isolated gastric mucosa. Both compounds produced concentration-related inhibitions of histamine-induced secretion whereas only U-46619 inhibited methacholine-stimulated and basal secretion, and neither compound had any effect on the secretory response to dbcAMP. Indomethacin had no effect on the antisecretory activity of PGE2 but markedly reduced the potency of U-46619 suggesting that endogenous prostaglandins play a role in the U-46619 responses. However, direct inhibitory effects of U-46619 were seen at high concentrations. The thromboxane receptor antagonist AH23848, at concentrations selective for thromboxane receptors, had no effect on responses to PGE2 but markedly inhibited the effects of U-46619. We conclude that the antisecretory profile of U-46619 differs from that of PGE2. U-46619 has both direct and indirect antisecretory effects and these are mediated via thromboxane receptors in the rat gastric mucosa.     

Cortisone cure of the eyelid closure defect in lidgap-Stein fetal mice: a dose-response and time-response study as a test of the hypomorph hypothesis for the lidgap alleles

The lidgap-Stein mutation is one of a series of alleles that cause the birth defect open eyes in mice. Previously it was known that cortisone administered during pregnancy prevents the defect in some lidgap-Stein fetuses. In this study, the hypothesis that lidgap-Stein is a hypomorph of effect intermediate between that of its alleles lidgap-Miller (least abnormal) and lidgap-Gates (most abnormal) was tested in a dose-response, time-response, and scanning electron microscopic study. Cortisone produced a response at doses of 20-80 mg/kg, with maximum cure of 30% in right eyes, 24% in left eyes, and 13% bilaterally. There was significantly more response in right than in left eyes. The response was slight at doses of less than 20 mg/kg and dropped to zero at the highest dose of 120 mg/kg. Treatment on days 13 or 14 gave the maximum response, with little or no response to treatment on days 10, 11, 12, 15, or 16. Severity of defect, measured as the size of gap in open eyes on day 18, was not reduced as the frequency of open eyes was reduced; most unclosed eyes remained wide open. The much lower level of maximum bilateral response to cortisone in lidgap-Stein (13%) than in lidgap-Miller (94%) is entirely compatible with the hypomorph hypothesis.     

Inositol 1,4,5-trisphosphate (IP3) induced rapid formation of thromboxane B2 in saponin-permeabilised human platelets: mechanism of IP3 action

The mechanism of IP3-induced activation of saponin-permeabilised platelets has been examined. Saponin permeabilization resulted in the leakage of low-Mr substances into and from the cells without loss of cytoplasmic proteins. Addition of IP3 rapidly induced a dose-related formation of thromboxane B2 and release into the medium, leading to the responses of shape change, aggregation and [14C]5HT release. These responses were inhibited by the thromboxane A2 receptor antagonist AH23848. The IP3-induced release of 45Ca from intracellular stores was not affected by indomethacin. Synthesis of thromboxane was inhibited if Ca2+ elevation was prevented by using Ca-EGTA buffers during permeabilization. These studies indicate that IP3-induced activation was due to Ca2+ mobilisation leading to phospholipase activation and thromboxane synthesis.     

AH23848 accelerates inducible nitric oxide synthase degradation through attenuation of cAMP signaling in glomerular mesangial cells.

Excessive release of nitric oxide (NO) by mesangial cells contributes to the pathogenesis of glomerulonephritis. Prostaglandin E(2) (PGE(2)) produced at inflammatory sites regulates the release of NO through its downstream signaling. In glomerular mesangial cells (MES-13 cells), PGE(2) modulated NO production mainly through EP4 receptor in a cAMP-dependent manner. Lipopolysaccharide and interferon-gamma (LPS+IFNgamma)-induced NO production, inducible nitric oxide synthase (iNOS) gene and protein expression were greatly inhibited by AH23848, an EP4 antagonist. Further investigation indicated that AH23848 attenuated endogenous cAMP accumulation in MES-13 cells and modulated NO production through declination of iNOS gene expression and acceleration of iNOS protein degradation. AH23848 downregulated the iNOS protein in MES-13 cells through protein kinase A (PKA) since KT5720, a PKA-specific inhibitor, reduced iNOS protein stability. A short exposure of activated MES-13 cells to okadaic acid augmented iNOS activity. AH23848 and KT5720 attenuated serine/threonine phosphorylation of iNOS protein in LPS + IFNgamma-stimulated MES-13 cells. The results of this study led us to speculate that cAMP might regulate iNOS-stimulated NO synthesis through posttranslational mechanisms. Attenuation of cAMP signaling and the phosphorylation status of the iNOS protein may account for the effect of AH23848 in accelerating iNOS protein degradation in MES-13 cells.     

Opposing hemodynamic effects of substance P on pulmonary vasculature in rabbits

Substance P is a peptide implicated in the control of a variety of physiological processes. Although substance P-containing neurons impinge on the pulmonary vasculature, the effects of substance P on the pulmonary circulation have not been systematically investigated. Rabbits were anesthetized with methohexital sodium and paralyzed with pancuronium bromide. Injection of substance P (0.002-0.10 microgram/kg) in the vena cava produced dose-dependent pulmonary vasoconstriction and systemic vasodilation. Pulmonary arterial pressure reached a peak within 15-20 s and declined toward base line over 10 min. Aortic pressure fell rapidly, reaching minimum at 5-10 s. At higher doses cardiac output fell transiently, resulting in a 65% fall in pulmonary vascular conductance. If repeat substance P dosages were administered 15 min apart, there was no tachyphylaxis. Pulmonary vasoconstriction was inhibited by the cyclooxygenase blocker meclofenamate (10 mg/kg) and the thromboxane synthase inhibitor Dazmegrel (UK-38,485) (2 mg/kg). In contrast, vasoconstriction was enhanced by atropine (2 mg/kg). In Dazmegrel-treated animals in whom pulmonary vasoconstriction was established by epinephrine infusion, low doses of substance P produced vasodilation. Our findings indicate that substance P produces pulmonary vasoconstriction via prostaglandin (particularly thromboxane) generation and pulmonary vasodilation via activation of cholinergic pathways.     

Congenital defects in the offspring of male mice treated with ethylnitrosourea

Daily doses of ENU (25-100 mg/kg) were injected intraperitoneally into ICR strain male mice for 5 days. The males were mated to untreated virgin females of the same strain on days 1-16 and 64-80 after the last dose. Copulations during these periods involve, respectively, treated postmeiotic cells and spermatogonial stem cells. The uterine contents were examined on day 18 of pregnancy for evidence of dominant lethal effects. The fetuses were examined for external and skeletal abnormalities. ENU treatment of either postmeiotic cells or spermatogonial stem cells caused dose-dependent significant increases in the incidence of abnormal fetuses over the control level. The induction rate per live fetus per unit dose in mg/kg by treating spermatogonial stem cells was estimated to be 1.0 X 10(-4), which is 3-fold lower than the rate previously estimated for the same endpoint at the same germ cell stage with MNU. Cleft palate was the most frequent external abnormality in the ENU-treated and the control series. Malformed vertebrae was the most frequent skeletal abnormality in the treated series. Rib fusion was the only skeletal malformation seen in the control series. Dominant lethals were clearly induced when germ cells were treated as postmeiotic cells.     

Effects of aminoglutethimide phosphate administration on the incidence of pregnancy and number of live fetuses in superovulated rats

This study examined the effects of aminoglutethimide phosphate (AGP) administration on the incidence of pregnancy and number of live fetuses in superovulated rats. Immature (31- to 33-day-old) young (8- to 9-week-old) and (14- to 16-week-old) rats were treated with 20 or 40 IU of PMSG and the same dosses of hCG at 52 to 54 hour-intervals (GTH rats). One half of the GTH rats were injected with 10 mg AGP twice a day for the first 3 days of pregnancy (AGP rats). Pregnancy rates of all AGP rats (95 to 100%) were much higher than the GTH rats (12.7). Young and adult AGP rats treated with 20 IU of GTH had 14.7 and 15.3 live fetuses per dam, respectively. These numbers of live fetuses increased to 17.8 and 18.4 after treatment with 40 IU of GTH. These values were significantly greater than those of the control rats (12.3 to 12.8; P<0.01), but the fetal weights were lower. The results show that AGP administration improves pregnancy rates and the number of live fetuses in young and adult superovulated rats.     

Protective actions of a new thromboxane synthetase inhibitor in arachidonate induced sudden death

A new thromboxane synthetase inhibitor, OKY-046, at doses of 0.5 and 1.0 mg/kg prevented mortality induced by sodium arachidonate in 100% of the rabbits studied. Sodium arachidonate at a dose of 1.25 mg/kg uniformly decreased mean arterial blood pressure to values approximately 0 mm Hg, stopped respiration and produced sudden death within 3-5 minutes in all rabbits studied. OKY-046 prevented all these sequelae of the sodium arachidonate. Untreated rabbits challenged with sodium arachidonate develop large increases in circulating thromboxane B2 (TxB2) and 6-keto PGF1 alpha of about 12- to 18-fold. In contrast, OKY-046 prevented the increase in TxB2 concentrations and the pulmonary thrombosis, but did not block the rise in 6-keto PGF1 alpha following arachidonate injection. These results suggest that the protective mechanism of OKY-046 in arachidonate induced sudden death is via selective inhibition of thromboxane synthesis.     

Exposure of the pregnant rat to warfarin and vitamin K1: an animal model of intraventricular hemorrhage in the fetus

Pregnant Sprague-Dawley rats were given daily oral doses of sodium warfarin (100 mg/kg) and concurrent intramuscular injections of vitamin K1 (10 mg/kg). This dosing regimen did not have any apparent deleterious effect on the dams and did not affect the fetuses when administered from day 1 to day 12 of pregnancy. However, similar treatment from day 9 to 20 caused hemorrhage in the fetuses examined on day 21 of gestation. There were no hemorrhages in the control fetuses from dams receiving vitamin K1 only. The lowest effective dose of warfarin, in conjunction with daily doses of vitamin K1, was 3 mg/kg. This dose caused hemorrhage in 28% of fetuses; the incidence of affected fetuses was not further increased by doses of warfarin up to 100 mg/kg. Hemorrhages affected the fetal brain, face, eyes, and ear and occasionally the limbs. Brain hemorrhages were frequently intraventricular and caused various degrees of hydrocephaly. Bony defects were not a feature of prenatal exposure to warfarin. These results show that prenatal exposure of the rat to warfarin and vitamin K duplicates the hemorrhagic abnormalities and pathology associated with prenatal exposure to warfarin in the human. It did not induce bony or facial defects probably because the vitamin K-dependent components of bone development occur postnatally in the rat. This model should allow detailed determination of the role of vitamin K-dependent proteins in development.     

Endogenous EP4 prostaglandin receptors coupled positively to adenylyl cyclase in Chinese hamster ovary cells: pharmacological characterization

The purpose of these studies was to investigate the pharmacology of E-series and selected prostaglandins of other classes on adenylyl cyclase activity in Chinese hamster ovary (CHO) cells expressing an endogenous prostanoid receptor and to compare these responses with those from immortalized human non-pigmented ciliary epithelial (NPE) cells containing the EP2 receptor. 11-deoxy-PGE2 was the most potent of the 16 prostanoid agonists tested for stimulating cAMP formation with a potency (EC50) value of 26 +/- 6 nM in the CHO cells. The endogenous ligand, PGE2, exhibited potencies of 40 +/- 7 nM (n = 24) in the CHO cells and 67 +/- 9 nM (n = 46) in the NPE cells. The EP2 receptor agonist, butaprost, produced an EC50 value of 212 +/- 58 nM (n = 4) in the NPE cells while being inactive (EC50 > 10,000 nM, n = 6) in the CHO cells. The EP4 receptor selective antagonists, AH22921 and AH23848B, at a concentration of 30 microM, caused a 2.2 +/- 0.5 (n = 4) and 8.2 +/- 2.7 (n = 4) fold rightward shift in the PGE2 concentration-response curves in the CHO cells, yielding apparent pKb values of 4.6 +/- 0.6 and 5.3 +/- 0.2 (n = 4), respectively. AH22921 and AH23848B were non-competitive antagonists at the CHO cell EP4 receptor, but did not shift the PGE2 concentration-response curves in the NPE cells containing the EP2 receptor. These studies have characterized the functional prostaglandin receptors in CHO cells pharmacologically and shown them to be consistent with the EP4 subtype.     

Inhibition of platelet thromboxane A2 synthase activity by sodium 5-(3'-pyridinylmethyl)benzofuran-2-carboxylate

This report outlines the activity of a new thromboxane synthase inhibitor sodium, 5-(3-pyridinylmethyl)-2-benzofurancarboxylate, (U-63557A). U-63557A is a potent inhibitor of the thromboxane synthase in human platelets in vitro, as well as in rhesus monkey platelets ex vivo. A single oral dose of 3.0 mg/kg U-63557A inhibits the platelet thromboxane synthase in rhesus monkeys approximately 80% for at least 12 hrs. U-63557A has been administered to monkeys twice a day, (10 mg/kg) for 14 days, without evidence of drug tachyphylaxis or rebound. U-63557A does not inhibit thrombin-stimulated PGI2 biosynthesis in human endothelial cells, the 5-lipoxygenase in human neutrophils, or the cyclo-oxygenase in a variety of test systems. In anesthetized dogs, U-63557A injected i.v. at 0.1 to 5 mg/kg prevented the blockage of stenosed coronary arteries caused platelet aggregation. Similar effects were obtained by oral administration of 1-5 mg/kg. The thromboxane synthase inhibitor was more efficacious than cyclooxygenase inhibitors and equal to PGI2 in efficacy. Under appropriate conditions the protective effects of U-63557A could be reversed by i.v. cyclooxygenase inhibitors suggesting that its efficacy depended in part on endogenous PGI2 formation. Due to its specificity, oral activity, and extended duration of action, U-63557A is a promising compound for the evaluation of the role of thromboxane synthase in a variety of pathophysiological states.     

Prevention by tiopronin (2-mercaptopropionyl glycine) of methylmercuric chloride-induced teratogenic and fetotoxic effects in mice

Previous investigations (Fuyuta et al., '76, '79) have shown that a single oral administration of 25 mg/kg methylmercuric chloride (MMC) to pregnant ICR mice on day 10 of pregnancy induced cleft palate in a remarkably high incidence in fetuses. Based on these findings, the present study dealing with the prevention of cleft palate by Tiopronin, (2-mercaptopropionyl glycine, Tp), was initiated. Twenty females in the positive control group were given 25 mg/kg MMC orally on day 10 of pregnancy and then given physiological saline intraperitoneally. Twenty females in the negative control group were given distilled water orally and then given saline intraperitoneally. Cleft palate was found in 98.1% of fetuses in the positive control group and none of them in the negative control group. Twenty females were pretreated with a single oral dose of 25 mg/kg MMC on day 10 of pregnancy and were posttreated with Tp intraperitoneally, immediately and at every 24, 48 and 72 hours after the MMC treatment. The doses of Tp were 320, 160 and 80 mg/kg/day. The incidences of cleft palate in fetuses were reduced to 1.49, 31.3 and 47.8% in the Tp-treated groups with the doses of 320, 160 and 80 mg/kg/day, respectively. Tiopronin could effectively prevent the expected incidence of cleft palate. Other types of abnormalities as well as fetotoxicity represented by reduced fetal body weight were also effectively prevented with the Tp-treatment.     

Effects of the Ay gene on susceptibility to hydrocortisone fetotoxicity and teratogenicity in mice

Effects of the Ay gene, a coat color gene, on susceptibility to hydrocortisone fetotoxicity and teratogenicity were investigated by using the congenic strain of C57BL/6-Ay (Ay/a) which had been maintained by repeated back-crosses of the Ay gene to the C57BL/6 (a/a) background. Matings were conducted as follows (female x male): group I, a/a; group II, a/a x Ay/a; and group III, Ay/a x a/a. Pregnant females were subcutaneously given daily doses of 0, 12.5, 25, or 50 mg/kg of hydrocortisone on days 10-13 of pregnancy. On day 18 of pregnancy, fetuses were sexed, weighed, and examined for external abnormalities. In group I, the mean fetal weight was significantly decreased at a dose of 25 mg/kg or more. The incidences of cleft palate were 3.2 and 22.7% at 25 and 50 mg/kg, respectively. In group II, in which half of the fetuses were expected to carry the Ay gene, the mean fetal weight was decreased significantly at 12.5 mg/kg or more. The incidence of cleft palate in group II at 50 mg/kg was 44.2%, which was significantly higher than that in group I. In group III, in which maternal mice as well as half of their fetuses carried the Ay gene, a decrease in the mean fetal weight was greater than in group II. In addition, the mean percentage of fetal resorptions was significantly increased at 50 mg/kg. The incidence of cleft palate in group III was significantly increased at 25 mg/kg (10.5%) when compared with those in groups I and II. These results indicate that the Ay gene may be associated with susceptibility to hydrocortisone fetotoxicity and teratogenicity in mice.     

Postnatal closure of membranous ventricular septal defects in Sprague-Dawley rat pups after maternal exposure with trimethadione

BACKGROUND: Congenital membranous ventricular septal defects (VSD) have been shown to close during postnatal development in rats [Solomon et al., Teratology 55:185-194, 1997]. Although they may differ in size, spontaneous and treatment-related VSD are histologically similar; however, the postnatal fate of treatment-induced VSD is not known. The objective of this study was to determine if treatment-induced VSD persist throughout postnatal development. METHODS: Groups of 40 female rats were given oral doses of trimethadione (TMD) at 400 mg/kg/day (200 b.i.d.) or 600 mg/kg/day (300 b.i.d.) on Gestation Days (GD) 9 and 10. Twenty dams in each group were designated for Cesarean section and 20 were allowed to deliver and rear their offspring to Postnatal Day (PND) 21. The integrity of the ventricular septum was evaluated in fetuses (GD 21) and pups (PND 21). RESULTS: The incidence of membranous VSD was 0.6, 7.6, and 49.8% per litter in the Control, 400, and 600 mg/kg groups, respectively, on GD 21. Both the incidence and severity of VSD increased with dose. The VSD at 400 mg/kg were small in size and initially detected by the presence of blood flowing through the defect from the closed right ventricle. In the 600 mg/kg dose group, the VSD, although still membranous, were larger and more readily detected without the need to examine the blood flow. At 600 mg/kg, not only were the VSD larger than those in the Control or the 400 mg/kg group, 10.1% per litter of the affected fetuses had other vessel anomalies associated with the VSD, which were incompatible with pup survival. On PND 21, VSD was noted in 0.3, 0, and 6.4% per litter evaluated in the Control, 400, and 600 mg/kg groups, respectively. This demonstrates that the small, isolated treatment-related VSD can resolve postnatally; however, the closure of the larger or more severe VSD may be prolonged or may not occur at all. Although TMD exposure reduced group mean fetal weights at both dose levels, there was no difference between the mean weight of fetuses with VSD and those fetuses without VSD in the same group. CONCLUSION: Treatment-induced VSD close postnatally, and appears to be a delay in cardiac development not associated with fetal weight. The timing of closure and survivability during closure is dependent on the severity of the VSD. Further characterization of the two sizes of VSD may provide diagnostic clarity; however, the current data support the smaller VSD as a variation with no significant impact on viability and growth, and the more severe VSD to be a malformation.     

Histological effects of 6-mercaptopurine on the fetal rat central nervous system: a light-microscopic study.

Wistar rats were administered single doses of 16 or 50 mg/kg 6-mercaptopurine (6-MP) on day 12 of pregnancy. Necrosis in the fetal forebrain and spinal cord was studied by light microscope 6, 12, 14, 48, 72, and 81 h and 8 days afterward. The extent of necrosis was dose dependent. The first necroses were seen after 24 h, regardless of location (brain, spinal cord) or dose; but the extent was greatest after 48 h. All necrotic cells had a typical appearance; they were ballooned and often fragmented, their nuclei were darkly colored and frequently pyknotic, and they were often karyorhexic. Necroses appeared almost exclusively at sites of beginning cellular differentiation, i.e., in the intermediate zone. In the spinal cord the ventricular zone was also necrotic and the alar plate (dorsal horn) always affected. Phagocytizing cells (macrophages) appeared in the spinal cord after 48 h and in the brain after 72 h. After 81 h all the necrotic material had been phagocytized, at which time there was a massive congestion of the extra- and intracerebral vessels. Hemorrhages appeared in defined localizations. Eight days after exposure to 16 mg/kg 6-MP fetuses no longer showed any visible deviations. Fetuses exposed to 50 mg/kg showed deviations in the cytoarchitecture of the neopallium: an extremely broadened ventricular zone, few cells in the intermediate zone, and extensive rarefaction cells in the cortical plate with no clear layer structure. In the spinal cord, cleft formations were especially noticeable in the dorsal-horn region. All fetuses showed a hydrocephalus externus after 50 mg/kg. The mechanism leading to necrosis is discussed.