Time-response and dose-response to acetazolamide in the WB/ReJ and C57BL/6J mouse strains: genetic interaction in the ectrodactyly response

The WB/ReJ and C57BL/6J strains were compared in their time and dose responses to acetazolamide administered in a single subcutaneous injection regime. WB/ReJ has a genetically determined, high-frequency, transient fetal edema that has maximum expression on day 14 and is resolved by day 18. Acetazolamide, at 1,000 mg/kg, appears to induce edema in WB/ReJ with a time of response on days 9 and 10, and the induced edema follows the same time course of appearance and disappearance as the spontaneous trait. The dose-response analysis is not interpretable in the WB/ReJ and C57BL/6J strains and their reciprocal F1 fetuses because there was significant response only at the highest dose (2,000 mg/kg) used in this study. The time of ectrodactyly response is maximal on day 9 in both WB/ReJ and C57BL/6J strains. The dose-response analysis demonstrates that, for the usual measure of total fetuses with ectrodactyly (or penetrance), the Wb/ReJ and C57BL/6J strains and the WB/ReJ x C57BL/6J F1 (WB.B6F1) have the same slope of the dose-response curve and the strain difference in response can be interpreted as a difference in dosage tolerance. The tolerance of WB/ReJ is twofold greater than that of C57BL/6J. This overdominance of relative resistance to acetazolamide ectrodactyly supports the general finding of directional dominance of relative resistance among genetically different strain pairs. The median effective dose for penetrance of the ectrodactyly response of the reciprocal B6.WBF1 embryo is similar to the WB.B6F1, but the slope of the dose-response curve is significantly different, and a different teratogenic mechanism of response may be involved. Ectrodactyly was predominantly right sided in all genotypes, and, in bilaterally affected fetuses, the right forelimb was more severely affected. An unexpected difference between WB/ReJ and C57BL/6J was found when the laterality of ectrodactyly was analyzed further. There is a significant increase with dosage in bilaterally affected fetuses (a measure of expressivity) in C57BL/6J but not in WB/ReJ, even though the dose-response of total affected fetuses (penetrance) is similar in both strains. In C57BL/6J, the left and right forelimbs are correlated in their responses with the left, requiring approximately a threefold greater dose. The left and right forelimbs are symmetrical in response, and the difference can be interpreted in terms of a developmental (or teratogenic) gradient. In WB/ReJ, the right forelimb has the same dose response as C57BL/6J and requires a twofold greater dose than the right forelimb of C57BL/6J, but the left forelimb has a very flat slope and is not correlated with the response of the right.(ABSTRACT TRUNCATED AT 400 WORDS)     

Genetically determined transient edema found in the WB/ReJ mouse strain in a teratogenic survey with acetazolamide

A continuing survey of the genetic variability of different mouse strains to acetazolamide teratogenesis demonstrated the WB/ReJ strain expresses a high frequency of induced subcutaneous edema in day 15 fetuses. In treated WB/ReJ fetuses, the probability of expression of edema is independent of the expression of forelimb ectrodactyly and, with the dosage regime, there is no significant increase in acetazolamide-induced resorption. It was surprising to find a high frequency of spontaneous edema on day 15 in untreated WB/ReJ fetuses. The spontaneous edema is a transient trait with maximum expression (56%) on day 14, and it is resolved by day 17 without apparent consequence to the survival of previously affected fetuses. There is no sex dimorphism in the liability to express the transient edema. Preliminary genetic crosses to investigate the spontaneous edema were made between WB/ReJ and the C57BL/6J strain, which historically had not be observed to express spontaneous edema. A low frequency of spontaneous edema was observed on day 14 in both C57BL/6J and the reciprocal F1 fetuses. The trait is not additive because there is dominance deviation of the BC1 fetuses in the direction of the F1 fetuses. The data were fitted to a threshold model suggesting that the developmental liability to express the difference in transient edema is determined by more than one gene, but the data can be interpreted by a minimum of two loci with duplicate epistasis. The observed differences in frequencies of edema suggest the genetic model can be tested with relatively few test crosses.     

Differential teratogenesis of all-trans-retinoic acid administered on gestational day 9.5 to SWV and C57BL/6N mice: emphasis on limb dysmorphology

BACKGROUND: Mouse strain differences in teratologic response are well documented. However, because retinoids cause similar malformation syndromes across many species, the strain differences may be predicted to be minimal. The goals of this study were to characterize and explain the differences between the C57BL/6N and SWV mouse strains in terms of all-trans-retinoic acid (RA)-induced teratologic effects at the time of gestation that cause postaxial forelimb ectrodactyly. METHODS: Visceral and skeletal malformations were determined by Wilson's sectioning and double-staining techniques, respectively; developmental staging was performed according to the somite count; and retinoid concentrations were assessed by HPLC. RESULTS: C57BL/6N mice were more susceptible than SWV mice to induction of embryolethality, cardiovascular defects, and forelimb ectrodactyly, whereas the opposite was true for the induction of ear, thymus, and tail agenesis, and cleft palate, gastroschisis, and anal atresia. As determined by somite counts, 1 strain intercross was developmentally advanced compared to the parental strains and the reciprocal cross. Retinoid susceptibility was equivalent between the reciprocal crosses for some malformations and determined by the maternal genotype for others. Toxicokinetic experiments showed that whole-embryo peak retinoid concentrations did not differ between the strains, but the area under the curve (AUC) for all-trans-RA was 1.3 times higher in C57BL/6N than in SWV embryos. CONCLUSIONS: The malformation spectrum induced by RA was strain-specific, and the strain sensitivity for forelimb ectrodactyly was consistent with all previously tested teratogenic agents (i.e., C57BL/6N was more sensitive than SWV). The strain differences in teratologic effects were not explained by developmental timing differences or toxicokinetic differences at the whole-embryo level.     

Teratogenesis of acetazolamide in the CBA/J and SWV strains of mice. I. Teratology.

SWV mice were totally resistant to the teratogenic and embryolethal actions of acetazolamide. The time of maximal sensitivity to acetazolamide-induced ectrodactyly in the CBA/J strain was the middle of day 10; the dose response at this time was studied. Comparison of the responses of the two strains and reciprocal hybrids indicated that sensitivity is a property of the embryo and is not maternally mediated. SWV mice were also resistant to dichlorphenamide which suggests they may be resistant to many or even all teratogenic carbonic anhydrase inhibitors.     

Potentiation of acetazolamide induced ectrodactyly in Wistar rats by vasoactive agents and physical clamping of the uterus

The vasoactive agents serotonin, ergotamine, and nicotine potentiate acetazolamide induced forelimb ectrodactyly (missing digits) in Wistar rats. These vasoactive agents administered alone do not produce forelimb ectrodactyly and are not known to be inhibitors of carbonic anhydrase. Additionally, physical clamping of the uterine horns in addition to oral acetazolamide administration increases the frequency of forelimb ectrodactyly, suggesting that decreased uterine blood flow can potentiate acetazolamide teratogenesis. Since the vasoactive agents used in this study are reported to possess uterine vasoconstrictive activity, a decrease in uterine blood flow is a plausible mechanism for the potentiative ability of these agents.     

Acetazolamide teratogenesis in Wistar rats: potentiation and antagonism by adrenergic agents

Acetazolamide, a carbonic anhydrase inhibitor, induced right forelimb ectrodactyly in rat fetuses when the mothers were treated on late day 10 and early day 11 of gestation. Coadministration of the selective alpha-1-adrenergic agonist phenylephrine significantly increased the incidence of acetazolamide-induced right forelimb ectrodactyly while failing to induce the lesion when administered alone. Pretreatment with the alpha-adrenergic antagonists phenoxybenzamine and prazosin prevented the phenylephrine-induced increase in right forelimb ectrodactyly. In addition, treatment with either phenoxybenzamine or prazosin in the absence of stimulation with phenylephrine significantly decreased the incidence of acetazolamide-induced ectrodactyly. The results suggest an adrenergic component in acetazolamide teratogenesis. Alterations in uterine blood flow are discussed as a plausible mechanism for the modification of the incidence of ectrodactyly by these adrenergic agents.     

The effect of administration time on malformations induced by three anticonvulsant agents in C57BL/6J mice with emphasis on forelimb ectrodactyly.

Exposure of C57BL/6J mice to three anticonvulsant derivatives, namely, dimethadione, sodium valproate, and sodium diphenylhydantoin, each induced postaxial forelimb ectrodactyly. The agents were administered at gestational days 9, 9 1/3, 9 2/3, and 10. It was determined that administration at day 9 2/3 induced the highest percentage of forelimb ectrodactyly for each of the three agents. The forelimb ectrodactyly response in the C57BL/6J strain was compared with the A/J strain (Collins et al., Teratology, 41:61-70, 1990); it was found that the C57BL/6J strain was more sensitive to dimethadione and the A/J strain was more sensitive to diphenylhydantoin and sodium valproate. The position of vertebral defects induced by sodium valproate correlated with the time of drug administration. The overall syndrome of malformations induced by the three anticonvulsant agents was relatively similar in the two mouse strains and differed between each of the anticonvulsant agents.     

Potentiating effect of caffeine on the teratogenicity of acetazolamide in C57BL/6J mice

Pregnant C57BL/6J mice were treated with 0 or 50 mg of caffeine (CAFF) per kg, and 0, 200 mg/kg (L) or 1,000 mg/kg (H) of acetazolamide (ACZM) during day 9 of gestation (9DPC). Individual fetuses were examined for gross morphological abnormalities and skeletal variations. The increase in fetal malformations seen, especially right forelimb electrodactyly, was augmented at both dose levels of acetazolamide by concomitant exposure to caffeine. Both frequency and severity of ectrodactyly were potentiated by caffeine. Skeletal examination revealed a reduction of the number of ossified cervical and caudal vertebral centra among litters exposed to ACZM at either dose. In either case (ACZM-H, ACZM-L) that effect was augmented by co-administration of CAFF. The first cervical vertebra (C1) appeared to provide the most sensitive index of teratogenic exposure. This study provides evidence that a subteratogenic dose of caffeine can potentiate the teratogenic effect of acetazolamide in C57BL/6J mice when dams are treated on day 9 of gestation. In addition, skeletal examination provided evidence that simultaneous treatment with both agents delayed fetal development. Many litters exposed to ACZM or both agents displayed a reduction in skeletal ossification even in the absence of gross morphological abnormalities, suggesting that ossification can be used as an indicator of prenatal exposure to potentially harmful substances in the C57BL/6 mouse strain.     

Interactive effects of cadmium and all-trans-retinoic acid on the induction of forelimb ectrodactyly in C57BL/6 mice

BACKGROUND: Most toxicological studies have tested single chemical agents at relatively high doses, and fewer studies have addressed the toxic effects of chemical interactions. It is important to understand the toxicity of chemical mixtures in order to assess the more realistic risks of environmental and occupational exposures. A number of chemicals are known to induce a predominantly postaxial forelimb ectrodactyly in C57BL/6 mice, including acetazolamide, ethanol, cadmium, valproic acid, carbon dioxide, dimethadione, phenytoin, and 13-cis-retinoic acid and all-trans-retinoic acid (RA). In the present study, the interactive effects of coadministration of cadmium and RA on developing limbs were investigated. METHODS: Pregnant C57BL/6 mice were treated with different intraperitoneal (IP) doses of cadmium chloride (CdCl2) and/or RA on gestational day (GD) 9.5, and fetuses were collected on GD 18 and double stained for examination of skeletal defects. RESULTS: When RA was given simultaneously with cadmium, a significant increase in the incidence and severity of forelimb ectrodactyly (predominantly postaxial) was observed compared to the results with corresponding doses of cadmium or RA alone. When mice were exposed to subthreshold doses of both cadmium (0.5 mg/kg) and RA (1 mg/kg), the combined treatment exceeded the threshold, resulting in forelimb ectrodactyly in 19% of the fetuses. Moreover, coadministration of cadmium and RA at doses exceeding the respective thresholds showed a synergistic effect, that is, 92% of fetuses were found with the forelimb defect as opposed to 10% if the response were additive. CONCLUSIONS: The findings demonstrate that concurrent exposure to these teratogens can have a synergistic effect and that subteratogenic doses may combine to exceed a threshold.     

Pathogenesis of the mouse forelimb deformity induced by acetazolamide: an electron microscopic study

Scanning electron microscopic observations after removal of the epidermis from developing limb buds reveal a fine mesenchymal cell process meshwork (CPM). The relationship between apical ectodermal ridge (AER) development and CPM density was investigated and related to the postaxial reduction deformities induced by acetazolamide (AA). AA was given orally to pregnant mice at 9 A.M. and 4 P.M. of day 9 and 9 A.M. of day 10 (VP = 0) in a dose of 1,000 mg/kg. Forelimb ectrodactyly, especially on the right, was the most common deformity observed. Scanning electron microscopic observations showed that the AER in AA-treated right forelimb buds did not extend postaxially as far as that in controls. The postaxial region with the hypoplastic AER became defective. Scanning and transmission electron microscopic observations revealed that in control and treated right forelimb buds, the CPM underneath the typical AER was sparser than that underneath the dorsal or ventral non-ridge epidermis. However, in treated right forelimb buds, the CPM underneath a hypoplastic AER was denser than that underneath the normal AER. These findings suggest that AA-induced deformity results from a disturbance of the AER-mesenchymal interactions.     

Genetic characteristics of the "EEC" syndrome (ectrodactyly, ectodermal dysplasis, cheilognathopalatoschisis)]

The analysis of the literature and author's observations of the "EEC" syndrome (ectrodactyly, ectodermal dysplasia and cleft lip/palate) revealed that this is a disorder with an autosomal-dominant type of inheritance with an incomplete penetrance and varying expressivity. Both sexes are affected with the same frequency. The complete form of the syndrome was mentioned in 27 cases only; all other patients had incomplete forms. The combination of two out of 3 main features is enough for the diagnosis of this syndrome. The most common trait of the "EEC" syndrome is ectrodactyly (73/77), clefts of lip or palate were observed in 53 patients out of 77, the ectodermal dysplasia was mentioned in 44 cases. There is an increase of mutation frequency in older parents.     

Reduced epithelial surface activity is related to a higher incidence of facial clefting in A/WySn mice

The epithelial surfaces of the facial primordia were evaluated by scanning electron microscopy (SEM) during primary palatogenesis in two genetically related mouse strains, the A/J and the A/WySn strains. These two strains were selected because the reported frequency of spontaneous cleft lip with or without cleft palate [CL(P)] in the A/J strain approximates 0%, whereas the spontaneous frequency of CL(P) in the A/WySn strain is 20-30%. The embryos were examined prior to (two to six tail somites), during (seven to ten tail somites), and after (ten to 14 tail somites) primary palate fusion. During fusion, epithelial surface activity (characterized as cellular debris, dissociated cells, cellular projections, and epithelial bridging) was more pronounced in A/J embryos. A/WySn embryos with spontaneous cleft lip exhibited a marked deficiency in epithelial activity when compared to their normal littermates. No discernible differences were detected in the facial morphology, with the exception of the distal end of the medial nasal prominence, which appeared longer in the A/J strain. This study suggests that the degree of epithelial surface activity at the putative site of fusion and the relative length of the medial nasal prominence may account for the observed differences in facial clefting of the two strains. Face shape, related to prominence divergence, was similar in the two strains and could not explain the higher incidence of clefting observed in the A/WySn strain.     

Increased susceptibility to 6-aminonicotinamide-induced cleft lip of heterozygote Dancer mice

Dancer heterozygotes (Dc/+) very rarely have cleft lip and show a dancing behaviour due to inner ear defects while homozygotes (Dc/Dc) have cleft lip. Males of the two genotypes Dc/+ and +/+ were mated to C3H strain and R stock females and Dc/+ males to Dc/+ females. On day 10/8 of gestation females were treated with 6-aminonicotinamide (6AN) at either 19 mg/kg or 28.5 mg/kg followed 3 h later by a protective dose of nicotinamide. Controls were untreated. Both 6AN treatments caused a significant increase in cleft lip to between 25% and 29% for crosses of Dc/+ males to C3H and R females whereas crosses with +/+ males gave 0% cleft lip. In the controls the cleft lip frequency was: for Dc/+ X Dc/+ 14%, Dc/+ X C3H 1.4%, and for the other three crosses 0%. The four crosses given the high dose of 6AN and the +/+ X C3H and Dc/+ X R cross at the low dose showed significantly increased resorption rates to between 23% and 47% over the control rates of from 5% to 11%. The presence of the Dc gene increased the susceptibility to cleft lip caused by 6AN.     

Induction of postaxial forelimb ectrodactyly with anticonvulsant agents in A/J mice

Exposure of A/J mice on day 9.5 of gestation to the derivatives of three acidic anticonvulsant agents, namely dimethadione, sodium valproate, and sodium diphenylhydantoin, each induced postaxial forelimb ectrodactyly predominantly of the right side. This specific malformation has previously been associated with the administration of acetazolamide to rodents; however, several agents can induce this same defect including other carbonic anhydrase inhibitors, carbon dioxide, cadmium, ethanol, ammonium chloride, and 13-cis retinoic acid. The relative potency of the three agents indicates no direct relationship to the pKa of the acid. Other than ectrodactyly, each of the anticonvulsant agents induced a compound-specific spectrum of malformations despite the uniform administration time. This finding suggests that these agents are capable of acting via different mechanisms or by the differential spatial and temporal dynamics of a common mechanism.     

Reduction of uterine blood flow by phenylephrine, an alpha-adrenergic agonist, in the day 11 pregnant rat: relationship to potentiation of acetazolamide teratogenesis

We have demonstrated previously that phenylephrine, a selective postsynaptic alpha-1-adrenergic agonist, significantly potentiates the incidence of acetazolamide-induced right forelimb ectrodactyly in a dose-response manner. As reported herein, phenylephrine also decreases maternal uterine blood flow in a dose-response manner as measured by radioactive microsphere methodology. At the potentiative dose of 12.5 mg/kg phenylephrine decreases uterine blood flow by 86.8% when compared to control. In turn, pretreatment with prazosin, a selective postsynaptic alpha-1-adrenergic antagonist, prevents this large decrease in uterine blood flow and abolishes the potentiation of acetazolamide teratogenesis by phenylephrine. Although the effects of acetazolamide or acetazolamide + phenylephrine on uterine blood flow were not measured the data suggest a correlation between decreased uterine blood flow and potentiation of acetazolamide teratogenesis.     

Unravelling the complex genetics of cleft lip in the mouse model

Nonsyndromic cleft lip in ``A' strain mice and humans is genetically complex and is distinct from isolated cleft palate. Cleft lip embryos recovered in 2.4% of 1485 first backcross (BC₁) segregants from a cross of A/WySnJ (24% cleft lip) and C57BL/6J (no cleft lip) in A/WySnJ mothers, and in testcrosses of 10 recombinant inbred (RI) strains (AXB/Pgn or BXA/Pgn), were used for gene mapping and for inference of genetic architecture. The A/WySnJ maternal genotype increased cleft lip risk in reciprocal crosses; the relevant genetic difference between AXB-6/Pgn (8%) and A/WySnJ (24%) is entirely maternal. A combination of new mapping panels (325 meioses), new markers, and a recombinant cleft lip embryo redefined the location of a recessive factor essential to cleft lip risk, clf1, and candidate genes Itgb3 and Crhr, to between D11Mit146/360 and D11Mit166/147. A screen of 54 YACs for 46 genes and SSLP loci located Wnt15, Wnt3, Crhr, Mtapt, Itgb3, Dlx3, and Dlx7 within the clf1 candidate region. The clf2 locus was newly mapped to Chromosome (Chr) 13 by a genome screen of BC₁ segregants, and further defined to a 4-cM region between D13Mit13/54 and D13Mit231 by strain distribution patterns of cleft lip liability and markers in testcrossed RI strains. Specific combinations of marker genotypes associated with cleft lip risk indicated that high risk in A/WySnJ mice is caused by epistatic interaction between clf1 and clf2 in the context of a genetic maternal effect. Human homologs of clf1 and clf2 are expected to be on 17q and 5q/9q. Received: 17 May 2000 / Accepted: 30 November 2000     

Review of drug-induced limb defects in mammals

The objective of this paper was to illustrate the spectrum of possible limb malformations in mammals resulting from drug exposure. A bibliography of 171 papers from 20 journals was generated from which pertinent data (drug used, limb defects reported, predominant defect location) were tabulated. These data should provide a basis for predictions about types of defects that might be expected in further studies and for judging postulated drug-induced human limb defects. However, direct extrapolation to humans is inappropriate. The following trends were observed: 1) Distal limb defects (autopod) are almost twice as common as proximal limb defects (stylopod and zygopod). 2) Ectrodactyly is the single most common type of limb defect, accounting for over half of the autopod defects. 3) Ectrodactyly is almost twice as common in the hindlimb as in the forelimb. 4) Postaxial ectrodactyly is over twice as common as preaxial ectrodactyly in the forelimb, but preaxial ectrodactyly is four times more common in the hindlimbs. 5) Polydactyly occurs with approximately equal frequency in forelimbs and hindlimbs, and preaxial polydactyly is most common in both fore and hindlimbs. 6) Polymelia (supernumerary limbs) occurred in one case, and may have been a spurious result. 7) Either transverse hemimelia is greatly underreported in teratology studies or it essentially does not occur. We have concluded that, at least in some cases, acetazolamide, adenine, 1,7-dimethylxanthine, and xanthine derivative aminophylline, retinoic acid, acetoxy-methyl-methylnitrosamine, aspirin, and cadmium can all cause unilateral defects.     

Expression of Veratrum alkaloid teratogenicity in the mouse

Jervine, a steroidal alkaloid found as a minor constituent in the teratogenic range plant Veratrum californicum, has produced similar terata in sheep, rabbit, hamster, and chick, although the sensitivity to the alkaloid varies in the different species. Sprague Dawley rats and Swiss Webster mice are relatively insensitive. The aim of this study was to determine the teratogenic potential of jervine in three strains of mice and to ascertain if the response is strain dependent. One strain, Swiss N:GP(S), was retested since a Swiss Webster strain had been found previously to be jervine-resistant. In addition, we tested C57BL/6J and A/J, which are known to differ in their response to the teratogenic action of steroids and vitamin A. Mice were treated by gavage with single doses of jervine (70, 150, or 300 mg/kg body weight) on either day 8, 9, or 10 of gestation. Jervine was teratogenic to C57BL/6J and A/J mice but not to N:GP(S). The induced terata included cleft lip with or without cleft palate, isolated cleft palate, mandibular micrognathia or agnathia, and limb malformations. Fetal teratogenicity and maternal and fetal toxicity were highly correlated. The prevalence of each defect and fetal death was a function of strain, dose, and time of treatment. Maternal death was higher in C57BL/6J than in A/J mice. Although some of the terata were similar, the response pattern between strains was different from corticosteroids and vitamin A for both sensitive period and the strain dose response. An effect on differentiation of chondrocyte precursors may account for many of the defects, but an earlier lethal effect on differentiation of neural crest cells or precordal mesenchyme may also occur.     

Ethanol-induced limb defects in mice: effect of strain and Ro15-4513

It is now thought that ethanol exerts many of its behavioral effects in the CNS by interaction with the gamma-aminobutyric acid (GABA) receptor, and it has been shown that the benzodiazepine reverse agonist Ro15-4513 reverses some of the CNS effects produced by ethanol. The hypothesis was tested that ethanol exerts its teratogenic effects through interaction with a putative embryonic GABA receptor by determining whether Ro15-4513 reverses ethanol-induced forelimb ectrodactyly in C57BL/6 mice. First, pregnant C57BL/6 dams were injected twice i.p. with ethanol (2.9 g/kg body weight, 4 hr apart) on day 10 of gestation: 49% of the fetuses were resorbed or dead and 46% of the survivors showed forelimb ectrodactyly. In contrast, when SWV mice were treated with ethanol, embryolethality was only 11.9% and no forelimb ectrodactyly was observed. In a second experiment, when ethanol (2.6 g/kg x 2) was administered to C57BL/6 mice, 34% resorptions and 31% forelimb ectrodactyly were observed. Ectrodactyly induced by ethanol was primarily of the forelimb and exclusively postaxial. Ethanol produced an unusual forelimb defect in a small number of instances where there was a postaxial autopod reduction defect coupled with a preaxial zeugopod reduction defect. Ro15-4513 administered alone (50 mg/kg x 2) was neither embryolethal nor teratogenic in C57BL/6 mice. To attempt to reverse the teratogenic effect of ethanol, dams that were injected 5 min before each ethanol administration with Ro15-4513 (0.5, 1, 2.5, 5, 10 mg/kg twice) showed no significant change in frequency of forelimb ectrodactyly compared to embryos treated with ethanol alone. However, resorptions increased significantly to 77% and 62% with the 5 and 10 mg/kg doses of Ro15-4513. Thus there appears to be an embryolethal interaction of Ro15-4513 with ethanol. Nevertheless, since Ro15-4513 did not reverse the teratogenic effect induced by ethanol, these results do not support the hypothesis that the teratogenic mechanism of ethanol is mediated through a putative embryonic GABA receptor.     

Counseling dilemmas in EEC syndrome

In this report we describe a prenatally diagnosed case with four-limb ectrodactyly and cleft lip/palate. The family history reveals three-generation oligodontia. The difficulties in counseling of the families with EEC syndrome are discussed.