The course of toxicity in the pregnant mouse after exposure to the cyanobacterial toxin cylindrospermopsin: clinical effects, serum chemistries, hematology, and histopathology.

Cylindrospermopsin (CYN) is a toxin produced by a variety of fresh-water cyanobacterial species worldwide and induces significant adverse effects in both livestock and humans. This study investigated the course of CYN-induced toxicity in pregnant mice exposed daily during either the period of major organogenesis (gestation days [GD] 8-12) or fetal growth (GD13-17). Endpoints include clinical signs of toxicity, serum analyses to evaluate hepatic and renal function, histopathology of liver and kidney, and hematology. Study animals were administered 50 µg/kg CYN once daily by ip route and euthanized 24 h after 1, 2, 3, 4, or 5 consecutive doses, or 6 or 13 d after the dosing period. The course of the CYN-induced effects was determined at all euthanasia times for the endpoints just outlined. Results indicated that CYN is a toxin, producing lethality in dams during the early part of gestation, significant weight loss, and bleeding in the gastrointestinal tract, tail tip, and peri-orbital tissues. Effects also included alterations in serum markers for liver function, histopathological changes in liver and kidney tissues, electrolyte abnormalities, leukocytosis, and posttreatment thrombocytopenia and reticulocytosis. The onset of symptoms was rapid, producing reductions in weight gain in GD8-12 animals, bleeding in the vaginal area in GD13-17 animals, and significant increases in sorbitol dehydrogenase (SDH) in both groups after a single dose. Although the GD8-12 dams displayed a 50% lethality, in GD13-17 animals only a single death occurred. Alterations seen in hepatic and renal function or histopathology do not appear to be of sufficient severity to produce death. Evidence indicates that bleeding may play a critical role in the onset of symptoms and eventually, in the observed lethality.

Toxicity and recovery in the pregnant mouse after gestational exposure to the cyanobacterial toxin, cylindrospermopsin.

Cylindrospermopsin (CYN) is a tricyclic alkaloid toxin produced by fresh water cyanobacterial species worldwide. CYN has been responsible for both livestock and human poisoning after oral exposure. This study investigated the toxicity of CYN to pregnant mice exposed during different segments of gestation. The course of recovery and individual responses to the toxin were evaluated. Adverse effects of CYN were monitored up to 7 weeks post-dosing by clinical examination, histopathology, biochemistry and gene expression. Exposure on gestational days (GD) 8-12 induced significantly more lethality than GD13-17 exposure. Periorbital, gastrointestinal and distal tail hemorrhages were seen in both groups. Serum markers indicative of hepatic injury (alanine amino transferase, aspartate amino transferase and sorbitol dehydrogenase) were increased in both groups; markers of renal dysfunction (blood urea nitrogen and creatinine) were elevated in the GD8-12 animals. Histopathology was observed in the liver (centrilobular necrosis) and kidney (interstitial inflammation) in groups exhibiting abnormal serum markers. The expression profiles of genes involved in ribosomal biogenesis, xenobiotic and lipid metabolism, inflammatory response and oxidative stress were altered 24 h after the final dose. One week after dosing, gross, histological and serum parameters had returned to normal, although increased liver/body weight ratio and one instance of gastrointestinal bleeding was found in the GD13-17 group. Gene expression changes persisted up to 2 weeks post-dosing and returned to normal by 4 weeks. Responses of individual animals to CYN exposure indicated highly significant inter-animal variability within the treated groups.

Reproductive effects of maternal and pre-weaning undernutrition in rat offspring: age at puberty, onset of female reproductive senescence and intergenerational pup growth and viability.

Maternal and/or postnatal undernutrition are widespread in human populations and are components of many experimental developmental and reproductive toxicology bio-assays. This study investigated in utero and/or pre-weaning undernutrition effects on reproductive maturation and senescence in the Sprague-Dawley rat as well as potential intergenerational effects. Pregnant rats were given food ad libitum or at 50% of normal dietary intake throughout pregnancy. Their offspring (control or IUGR) were cross-fostered to control dams with litter sizes of 8 or 16 pups (control and undernourished). Offspring body weights were reduced and onset of male puberty slightly delayed in animals from large postnatal litters. Similar body weight effects were observed in females but there was no difference in the age of vaginal opening. Female reproductive senescence as measured by onset of estrus acyclicity occurred at a younger age in IUGR-8-pup and Control-16-pup groups compared to Control-8-pup or IUGR-16-pup groups. Females were bred to control males and no evidence of adverse reproductive effects was found in any F2 groups. The offspring of the F1 generation did not show an intergenerational effect as documented in humans.

The relationship of maternal and fetal toxicity in developmental toxicology bioassays with notes on the biological significance of the "no observed adverse effect level".

Standard developmental toxicology bioassays are designed to identify agents with the potential to induce adverse effects and include dose levels that induce maternal toxicity. The work reported here was undertaken to evaluate the relationship of maternal and fetal toxicity. It constitutes an analysis of 125 developmental toxicity bioassays in the mouse, rat, and rabbit conducted by the National Toxicology Program. Although varying by species, general findings include: (1) most lowest observable adverse effect levels (LOAELs) were determined by reduced maternal gestational weight gain or fetal weight at term. (2) Maternal weight reductions are associated with reduced food intake for a variety of dissimilar test agents. (3) Lower fetal weights were associated with reduced maternal weight gains late in gestation. (4) The degree of fetal weight reduction is correlated with the extent of the maternal weight loss. In a substantial number of the studies, reduced fetal weights at term may, therefore, be due to maternal undernutrition caused by general toxicity rather than direct developmental insult. Consequently, such test agents may be erroneously classified as primary developmental toxicants. Experimental approaches to test the hypothesis that maternal undernutrition in standard developmental toxicology bioassays may be responsible for significant term fetal weight decrements are discussed.

The cyanobacterial toxin, cylindrospermopsin, induces fetal toxicity in the mouse after exposure late in gestation.

Cylindrospermopsin (cyn) is a cyanobacterial toxin implicated in human and wildlife poisonings. We have completed studies investigating the potential of purified cyn to induce developmental toxicity in mammals. The teratology study involved intraperitoneal injections (8.0-128 microg kg(-1)) on gestational days (GD) 8-12 with subsequent examination of term fetuses for viability, weight and morphological anomalies. Cyn was lethal to a significant portion of the dams receiving > or = 32 microg kg(-1). Surviving pregnant females were killed and fetuses removed for examination. Analysis indicates no adverse effects on litter size, fetal weight, or incidence of anomalies. Subsequently, 50 microg kg(-1) cyn was administered on GD 8-12 or 13-17. Animals were allowed to give birth and litters monitored for growth and viability. A reduction in litter size occurred in treated groups. Avg. pup wt. was only affected in the GD 13-17 group. GD 13-17 dams did not exhibit the toxicity noted in the GD 8-12 group but gave birth significantly earlier than controls. There was a significant number of dead GD 13-17 pups and incidences of blood in the gastrointestinal tract and hematomas in the tips of the tails in survivors. Pups were cross-fostered to control mothers in litters of 10. On postnatal days (PND) 5-6 there were no significant differences in weight gain or viability in GD 8-12 litters, while GD 13-17 litters had significantly reduced weight gain and viability. GD 13-17 exposed male pups still weighed significantly less than the controls after 15 months.

Potential developmental toxicity of anatoxin-a, a cyanobacterial toxin.

Some 2000 species of cyanobacteria (blue-green algae) occur globally in aquatic habitats. They are able to survive under a wide range of environmental conditions and some produce potent toxins. Toxin production is correlated with periods of rapid growth (blooms) and 25%-70% of blooms may be toxic. Anatoxin-a is an alkaloid neurotoxin that acts as a potent neuro-muscular blocking agent at the nicotinic receptor. Acute toxicity, following consumption of contaminated water, is characterized by rapid onset of paralysis, tremors, convulsions and death. Human exposures may occur from recreational water activities and dietary supplements, but are primarily through drinking water. The current studies were conducted to examine the effect of in utero exposure on postnatal viability, growth and neurodevelopment, to evaluate the potential of in vitro embryotoxicity, and to explore the synergistic relationship between anatoxin-a and the algal toxin microcystin-LR by the oral route. The results of preliminary studies on amphibian toxicity are also reported. Time-pregnant mice received 125 or 200 microg kg(-1) anatoxin-a by intraperitoneal injection on gestation days (GD) 8-12 or 13-17. Pup viability and weight were monitored over a 6-day period. Maternal toxicity (decreased motor activity) was observed at 200 microg kg(-1) in both treatment periods. There were no significant treatment-related effects on pup viability or weight on postnatal day (PND) 1 or 6. The GD 13-17 pups were evaluated on PND 6, 12 and 20 for standard markers of neurodevelopmental maturation (righting reflex, negative geotaxis and hanging grip time). No significant postnatal neurotoxicity was observed. In vitro developmental toxicity was evaluated in GD 8 mouse embryos exposed to 0.1-25 microm anatoxin-a for 26-28 h. Perturbations in mouse yolk sac vasculature were noted from the 1.0 microm concentration in the absence of significant embryonic dysmorphology. Potential algal toxin synergism was tested in mice receiving either 0, 500 or 1,000 microg kg(-1) microcystin-LR by gavage and approximately 50 min later receiving either 0, 500, 1,000 or 2,500 microg kg(-1) anatoxin-a by the same route. No deaths occurred at any dose and no definitive signs of intoxication were observed. Stages 17 and 25 toad embryos (Bufo arenarum) were exposed to 0.03-30.0 mg l(-1) of anatoxin-a for 10 days. Adverse effects included a dose-dependent transient narcosis, edema and loss of equilibrium. Most notable was the occurrence of 100% mortality at the high dose in both groups 6-13 days post-exposure. The observed delay between initial exposure and death is highly unusual for anatoxin-a.

Lack of evidence for intergenerational reproductive effects due to prenatal and postnatal undernutrition in the female CD-1 mouse.

The impacts of adverse environments during the prenatal and/or early postnatal periods may be manifested as functional deficits that occur later in life. Epidemiological studies have shown an association of sub-optimal pregnancy outcomes in one generation with similar events in the following one, a phenomenon termed the "intergenerational effect". Data indicate that the incidence of adverse pregnancy outcomes and/or low birth weight infants is more closely correlated with the mother's perinatal environment than with that during her pregnancy. However, epidemiological studies are inherently limited given the variability of lifestyles, ethnicity, nutritional status, and exposures to environmental factors. An appropriate animal model would permit control of parameters that may be impossible to evaluate in human populations. The current studies investigated the mouse as a possible animal model. Pregnant CD-1 mice were placed on an ad libitum or food-restricted diet (50% normal) throughout gestation to generate control (CON) and intrauterine growth retarded (IUGR) litters. At birth (postnatal day (PD) 1) pups (F1) were cross-fostered to control dams in litters of either 8 (CON) or 16 (postnatal food restriction (FR)). The experimental groups thus generated represented adequate nutrition (CON-CON) and undernutrition during the prenatal (IUGR-CON), or postnatal periods (CON-FR), or both (IUGR-FR). Pups of dams on a restricted diet during gestation had significant IUGR (P<0.001) as compared to controls (birth weights of 1.32 g versus 1.63 g). At weaning, the average weight of the pups was dependent on postnatal litter size and the difference in birth weights between IUGR and CON animals was not a significant factor. CON-CON pup weight was 24.1g and IUGR-CON was 22.2 g as compared to the CON-FR (17.0 g) and IUGR-FR (17.3 g) groups. The difference in weaning pup weights between the FR and CON groups was significant (P<0.01). The F1 FR females did not reach CON female weights at any time point through 11 months after weaning. At PD60, a single breeding period for all groups of females with CON males began and continued for 75 days with 17 opportunities for breeding. Animals that became pregnant during this time were removed and allowed to litter. No significant differences were noted in average F2 litter size or average pup weight at birth: (CON-CON 12.2/1.62 g; IUGR-CON 11.9/1.6 2 g; CON-FR 10.9/1.70 g; IUGR-FR 11.3/1.61 g). We conclude that body weight at birth in the CD-1 mouse is not correlated with growth through the period of weaning (PD28). We did not find any evidence for an intergenerational reproductive effect after developmental undernutrition.

Comparative pathogenesis of haloacetic acid and protein kinase inhibitor embryotoxicity in mouse whole embryo culture.

Haloacetic acids (HAs) are embryotoxic contaminants commonly found in drinking water. The mechanism of HA embryotoxicity has not been defined, but may be mediated in part by protein kinase C (PKC) inhibition. This study was conducted to evaluate the pathogenesis of HA embryotoxicity, and to compare these data with those from specific (Bis I) and non-specific (staurosporine) inhibitors of PKC. Embryos were incubated for varying times with several HAs, Bis I, staurosporine, or Bis V (a negative control). Cell cycle analysis was performed by flow cytometry following nuclear staining with propidium iodide; apoptosis was evaluated by fluorescence microscopy following LysoTracker staining. At concentrations producing 100% embryotoxicity with no embryolethality, only staurosporine perturbed the cell cycle. However, flow cytometry revealed accumulation of sub-G1 events (an apoptotic indicator) across time with bromochloroacetic acid, dichloroacetic acid, and staurosporine, but not dibromoacetic acid, Bis I, or Bis V. Sub-G1 events were particularly prominent in the head region, and remained at control levels in the heart. LysoTracker staining confirmed a similar pattern of apoptosis in the intact embryo; BCA and DCA produced intense staining in the prosencephalon, with virtually no staining in the heart. These data indicate that while cell-cycle perturbation may not mediate the pathogenesis of HA embryotoxicity, these agents do induce embryonic apoptosis. In addition, the lack of Bis I-induced apoptosis indicates that PKC inhibition is unlikely to be the sole mediator of HA embryotoxicity.

Dysmorphogenic effects of a specific protein kinase C inhibitor during neurulation.

Protein kinase C (PKC) plays a key role in signal transduction and is an important mediator of events throughout development. However, no information exists regarding the effect of a specific PKC inhibitor on mammalian embryogenesis during neurulation. This investigation was undertaken to examine the effects of a specific inhibitor of PKC, as well as inhibitors of other important kinases, on cultured mouse embryos. CD-1 mouse embryos (3 to 6 somite stage) were exposed to bisindolylmaleimide I (a specific PKC inhibitor) as well as specific inhibitors of PKA, PKG, and MAP kinase kinase for 24 h. The PKC inhibitor was a potent embryotoxicant and elicited malformations at concentrations as low as 0.01 microM. Inhibitors of other kinases also produced malformations but at much higher concentrations than those required to produce similar defects with the PKC inhibitor. These data suggest that PKC plays an important role in mammalian neurulation. Further research is required to clarify the mechanism by which PKC inhibition at this developmental stage produces malformations and the potential effects of environmental toxicants with PKC inhibitory properties on this signal transduction pathway.

Comparative effects of haloacetic acids in whole embryo culture.

A major class of disinfection by-products in drinking water are the haloacetic acids. Both dichloro- and trichloroacetic acids are teratogenic when administered to rats throughout organogenesis. However, there is little information regarding the developmental toxicity of other haloacetic acids. Therefore, 3-6 somite staged CD-1 mouse embryos were exposed to acetic acid (AA) or mono- (M), di- (D), and tri- (T) substituted fluoro- (F), chloro- (C), or bromo- (B) acetic acids in whole embryo culture in order to evaluate the effects of these agents on development. A 24 hour exposure to the haloacetic acids produced dysmorphogenesis. Effects on neural tube development ranged from prosencephalic hypoplasia to non-closure defects throughout the cranial region. Exposure to the haloacetic acids affected optic development, produced malpositioned and/or hypoplastic pharyngeal arches, and resulted in perturbation of heart development. In order to determine the relative toxicities of these agents, benchmark concentrations were calculated as the lower 95% confidence interval of the concentration that produced a 5% increase in neural tube defects. The benchmark concentrations occurred over a wide range with DFA (5912.6 microM) and MBA (2.7 microM) at the extremes. Using the benchmark concentrations to compare the chemicals gives a ranking of the agents in order of increasing potency as: DFA < TFA < DCA < AA < TBA < or = TCA < DBA < MCA < MBA. TCA and DCA have demonstrated ability to disrupt development in vivo but were among the least potent haloacetic acids in vitro. Because of the potential for widespread exposure to haloacetic acids in drinking water and the incomplete toxicity profile of these chemicals, further work on their developmental effects is warranted.

Prenatal alpha-difluoromethylornithine treatment: effects on postnatal renal growth and function in the rat.

DFMO (alpha-difluoromethylornithine) is a specific irreversible inhibitor of ornithine decarboxylase (ODC), a key enzyme in the biosynthesis of polyamines, which in turn control macromolecule synthesis during cell proliferation. The current study was designed to investigate the effects of inhibition of ODC during discrete prenatal periods on renal growth and function. We administered 5 doses of 500 mg/kg DFMO or saline s.c. to timed pregnant Sprague-Dawley rats at 12 hr intervals beginning on gestation days (GD) 11, 14, or 17. Half the dams were killed on GD 20 for fetal morphological analyses and half were allowed to go to term. Renal function was assessed on postnatal days (PD) 3, 6, 10, and 14 by tests of basal renal clearance and urinary concentrating ability, and on PD 42-44 we measured serum chemistries. All three gestational treatment regimens resulted in postnatal deficits in general growth. Only in the GD 11-13 treatment group was there evidence of embryotoxicity and neonatal renal pathophysiology. Fetal weights and urogenital morphology were altered following GD 14-16 treatment and there were persistent deficits of renal growth. GD 17-19 treatment was associated only with transient postnatal deficits of renal growth. Thus, inhibition of ODC during critical prenatal periods induced distinct developmental effects. However, there were no associations between impaired renal growth and function. These data indicate that general tissue growth is not always a predictor of physiological development and support the necessity of multifaceted approaches to the understanding of adverse developmental effects.

Functional teratogens of the rat kidney. I. Colchicine, dinoseb, and methyl salicylate.

Substances known or suspected to cause subtle or transient anatomical alterations in renal development were administered prenatally or neonatally to rats in order to determine whether they are capable of altering renal functional development. Colchicine alters mitotic activity and cytoskeletal structure and is teratogenic in many species. Since the kidney of the newborn rat undergoes extensive cellular proliferation and nephron differentiation, it is possible that neonatal administration of colchicine may affect nephron development. Dinoseb and methyl salicylate have previously been reported to produce a high incidence of dilated renal pelvis in the term rat fetus. Colchicine was injected sc, at 75 micrograms/kg, to Postnatal Day (PD) 1 Sprague-Dawley rats. Dinoseb was administered ip to pregnant Sprague-Dawley rats on Gestation Days 10-12 at doses of 8 or 10.5 mg/kg/day, and methyl salicylate was administered ip at doses of 200, 250, or 300 mg/kg/day on Gestation Days 11-12. Renal function was examined in pups from immediately after birth through weaning. Maximal urine concentrating ability was measured after DDAVP (desmopressin acetate, a vasopressin analog) injection in suckling rats, and after 24 hr of water deprivation in weanlings. Proximal tubule transport was measured in renal cortical slices. Basal urinary parameters, including urine flow, osmolality, pH, and chloride content, were measured. Colchicine treatment had no effect on body weight or kidney weight. There was a significant decrease in maximal urine osmolality in PD 30 rats measured after 24 hr of water deprivation. The urine concentrating deficit detected in functionally mature PD 30 rats suggests that colchicine treatment during renal histogenesis causes a latent deficit in medullary function in the absence of any gross morphological effects. The 10.5 mg/kg/day dose of dinoseb caused a weight reduction in neonates which persisted after weaning. Urine volume after DDAVP challenge was increased over controls in both dose groups on PD 6, but maximal urine concentration was unaffected. On PD 14, maximal urine concentration after DDAVP injection was decreased in the 10.5 mg/kg/day group. By PD 30, urine concentrating ability was comparable to controls. Renal cortical slices from the 10.5 mg/kg/day dose group had an enhanced ability to accumulate organic anions on PD 3 and 31, but opposite effects were observed in the low-dose group. No other renal functional parameters were altered. Urine osmolality after DDAVP challenge was decreased over controls in the 250 mg/kg/day methyl salicylate group on PD 6, and urine volume was increased in this group after DDAVP injection on PD 14.(ABSTRACT TRUNCATED AT 400 WORDS)

The significance of the dilated renal pelvis in the nitrofen-exposed rat fetus: effects on morphology and function.

Previously (R. J. Kavlock, B. F. Rehnberg, and E. H. Rogers, 1987, Teratology 36, 51-58) we reported that gestational exposure of rats to adriamycin induced alterations in development of the fetal renal papilla that persisted postnatally. The morphological effect was associated with functional deficits in neonatal animals as seen by their performance during a test of renal concentrating ability in the second postnatal week. In the present study, we utilized an experimental approach similar to that in the adriamycin study to evaluate the fate of the dilated renal pelvis that is induced in fetal rats following prenatal exposure to nitrofen. Groups of Long-Evans rats were exposed to 0, 6.25, 12.5, or 25 mg/kg of nitrofen on gestation Days 7-16. Renal morphology of the offspring were determined on gestation Day 21 and postnatal Week 5. The postnatal cohort was tested in the second postnatal week for their ability to excrete an osmotically concentrated urine. As was the case with adriamycin, the renal concentrating ability in the neonate was reduced and poor performance in the function test was associated with permanence of the morphological effect. By utilizing a standardized semiquantitative procedure to describe the status of the kidneys and ureters during development in combination with physiological assessment of organ performance, we were able to assess the morphological and functional development of the kidney. In the absence of other anomalies we suggest that offspring be monitored during postnatal development when alterations of the fetal renal papilla are observed in standard teratology bioassays in order to determine whether the effect is transient or permanent.

Fate of adriamycin-induced dilated renal pelvis in the fetal rat: functional and morphological effects in the offspring.

Previously we reported that gestational exposure to Adriamycin, an anthracycline antibiotic used in the treatment of neoplasms, reduced renal function in the neonatal rat, and we suggested that alterations in the development of the renal papilla might be responsible for the dysfunction. In this study we exposed groups of Sprague-Dawley rats to 0, 1.0, 1.25, or 1.5 mg/kg of Adriamycin on gestation days 10-12, a period previously shown to be effective in altering postnatal renal function with this compound. Offspring were evaluated at several developmental periods in order to 1) precisely define the morphological status of the urogenital system in Adriamycin-treated offspring; 2) replicate the finding of a decreased renal concentrating ability in the neonates; 3) determine the transience/permanence of any morphological effect; and 4) correlate any permanent alterations in urogenital morphology with our indicator of neonatal functional competence. Maternal Adriamycin treatment induced alterations in the development of the renal papilla that persisted well into the postnatal life of the offspring. The first appearance of the morphological alteration occurred in the absence of other general indicators of developmental toxicity such as growth retardation. The determination of the ultimate fate and functional consequences of the structural alterations required postnatal evaluations of the renal system. Finally, a relatively simple test of renal function in the neonate proved to be predictive of the permanence of the morphological effect, and the absolute test result showed a strong correlation with the incidence of the morphological effect in the overall population.

Critical prenatal periods for chlorambucil-induced functional alterations of the rat kidney.

Previous studies have demonstrated that exposure of rats to chlorambucil during the period of metanephric differentiation results in morphological and functional alterations of the kidneys after birth. The present study describes the effects of chlorambucil treatment at various gestational ages on neonatal renal function to determine if critical periods other than day 11 exist for inducing functional developmental toxicity of the kidneys. Groups of pregnant Sprague-Dawley rats were exposed to 4.5 mg/kg of chlorambucil on gestation days 9, 11, 13 or 15 and the offspring were evaluated for neonatal growth and viability, gross malformations of the kidneys, and renal physiology. The results demonstrate that the critical period for the induction of specific renal defects and hypoplasia lies on day 11 of gestation, but functional alterations of the kidneys were observed after exposure on day 15 of gestation. In terms of practical application, a combination of the basal clearance test and the renal concentration test together provide an efficient means for detecting prenatally induced functional alterations of the kidneys. When renal malformations or anomalies are observed in standard teratology bioassays, studies using techniques similar to those described here may be extremely useful in determining the biological significance as well as permanence or transience of effects such as renal hypoplasia, dilated renal pelvis, and dilated ureter.

Chlorambucil induced congenital renal hypoplasia: effects on basal renal function in the developing rat.

Administration of chlorambucil to pregnant rats on day 11 of gestation induced dose-related alterations in renal growth and function in the postnatal offspring. These effects occurred above and beyond the reductions in body growth and were evident in animals that displayed no overt malformation of the urogenital tract. Reductions in overall growth amounted to 0, 6 and 15% in the 3,4.5 and 6 mg/kg groups, respectively, while kidney weights were reduced by 7, 15 and 23%. The weights of the kidneys relative to the body were reduced 5, 9 and 10% with increasing dose. Although basal renal function was not affected by the degree of hypoplasia seen in the low dose group, reduced glomerular and tubular function were evident following a basal clearance test in the 2 highest dose groups. The data indicate that chlorambucil induced renal hypoplasia results in reductions in renal function that persist for at least the first 3 weeks after birth in the rat and that physiological assessment of developmental toxicity can provide an extremely useful addendum to the more classical morphological criteria.

Renal functional teratogenesis resulting from adriamycin exposure.

Pregnant Sprague-Dawley rats were exposed to adriamycin and offspring were evaluated for renal functional competence. Exposure consisted of 0, 1.0 or 1.5 mg/kg/day by intraperitoneal injection on either gestational days 7-10 or 10-12. The exposed offspring were evaluated for 1) growth and viability; 2) serum concentrations and renal clearances of creatinine, urea, glucose, sodium, potassium, chloride, and total osmotic particles; 3) the ability to excrete an osmotically concentrated urine following fluid deprivation; 4) the ability to excrete an osmotically dilute urine following isotonic volume expansion; and 5) the ability to secrete hydrogen ions following administration of a fixed acid. Exposure during days 7-10 of gestation produced greater evidence of overt developmental toxicity than did exposure during days 10-12 of gestation. The reverse was true, however, for the effects of adriamycin on renal function, as the majority of effects on these measures were found in the high-dose pups exposed during days 10-12 of gestation. The application of the renal function tests did not lower the observed effect level for adriamycin-induced developmental toxicity, but it did provide confirmatory information on the nature of the effect, on the magnitude of the effect in the exposed population, and on the possible morphological site of observed functional lesion. For reasons discussed in the text, a combination of the basal clearance test and the renal concentrating test appears to provide the most efficient means for detecting the presence of prenatally induced functional alterations of the kidneys.

Amphotericin B- and folic acid-induced nephropathies in developing rats.

The kidneys of newborn rats, which are both morphologically and physiologically immature, have been shown to be relatively insensitive to the nephrotoxic effects of several chemicals. To examine the specificity of these age-related differences, pups received sc injections of either 20 mg/kg of amphotericin B or 250 mg/kg folic acid, two known nephrotoxins in adult animals, on postnatal Day 1, 8, or 15. Renal function was examined by a basal clearance test and a hydropenia challenge at 1, 2, or 5 (6 in the case of amphotericin B) days after treatment. We observed no difference in degree of renal toxicity with age, but repair of renal damage tended to proceed slower at the youngest age. Amphotericin treatment produced uremia, increased fractional excretion of water and sodium, a decreased fractional excretion of urea, and a diminished hydropenia response but no change in creatinine clearance and no renal pathology. The observed pattern of renal toxicity may be attributed to an inability to maintain urea gradients in the distal segment of the nephron. Folic acid treatment resulted in greatly increased kidney weights with marked pathology, uremia with decreased creatinine clearance, increased fractional excretion of water, and a decreased hydropenia response. Unlike the renal toxicity observed following amphotericin treatment, renal toxicity from folic acid appears to be a nonspecific response to cell injury within the renal tubules. The data indicate that, in general, neonates do not possess a relative insensitivity to nephrotoxins and that renal physiological measurements which can be performed in neonatal rats are useful in evaluating and interpreting alterations in renal function.