Administration of 5-bromo-2'-deoxyuridine interferes with neuroblast proliferation and promotes apoptotic cell death in the rat cerebellar neuroepithelium.

The current study was conducted to assess whether a single administration of 5-bromo-2'-deoxyuridine (BrdU) interferes with cell proliferation and leads to the activation of apoptotic cellular events in the prenatal cerebellum. BrdU effects across a wide range of doses (25-300 µg/g b.w.) were analyzed using immunohistochemical and ultrastructural procedures. The pregnant rats were injected with BrdU at embryonic day 13, and their fetuses were sacrificed from 5 to 35 hr after exposure. The quantification of several parameters such as the density of mitotic figures, and BrdU and proliferating cell nuclear antigen (PCNA)-reactive cells showed that, in comparison with the saline injected rats, the administration of BrdU impairs the proliferative behavior of neuroepithelial cells. The above-mentioned parameters were significantly reduced in rats injected with 100 µg/g b.w. of BrdU. The reduction was more evident using 200 µg/g b.w. The most severe effects were found with 300 µg/g b.w. of BrdU. The present findings also revealed that high doses of BrdU lead to the activation of apoptotic cellular events as evidenced by both terminal deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL) assay and immunohistochemistry for active caspase-3. In comparison with saline rats, many apoptotic cells were found in rats injected with 100 µg/g b.w. of BrdU. The number of dying cells increased with 200 µg/g b.w. The most important number of apoptotic cells were observed in animals injected with 300 µg/g b.w. of BrdU. Ultrastructural studies confirmed the presence of neuroblasts at different stages of apoptosis.

Comprehensive expression analysis of mRNA and microRNA for the investigation of compensatory mechanisms in the rat kidney after unilateral nephrectomy.

Compensation is a physiological response that occurs during chemical exposure to maintain homeostasis. Because compensatory responses are not usually considered adverse effects, it is important to understand compensatory mechanisms for chemical risk assessment. Although the kidney is a major target organ for toxicity, there is controversy over whether hyperplasia or hypertrophy contributes to the compensatory mechanism, and there is limited information to apply for chemical risk assessment. In the present study, compensatory mechanisms of the kidney were investigated in a unilateral nephrectomy (UNx) model using adult male and female F344 rats. In residual kidneys of male and female rats after UNx, 5-bromo-2'-deoxyuridine-labeling indices and mRNA expression of cell cycle-related genes were increased, although there were no fluctuations in mRNA expression of transforming growth factor-ß1, which contributes to hypertrophy in renal tubules. Pathway analysis using mRNA expression data from a complementary DNA (cDNA) microarray revealed that canonical pathways related to cell proliferation were mainly activated and that forkhead box M1 (FOXM1) was an upstream regulator of compensatory cell proliferation in residual kidneys of male and female rats. cDNA microarray for microRNAs (miRNAs) demonstrated that nine miRNAs were downregulated in residual kidneys, and mRNA/miRNA integrated analysis indicated that miRNAs were associated with the expression of factors downstream of FOXM1. Overall, these results suggested that FOXM1-mediated hyperplasia rather than hypertrophy contributed to compensatory mechanisms in the kidney and that miRNAs regulated downstream FOXM1 signaling. These results will be beneficial for evaluating nephrotoxicity in chemical risk assessment and for developing new biomarkers to predict nephrotoxicity.

Separating host and microbiome contributions to drug pharmacokinetics and toxicity.

The gut microbiota is implicated in the metabolism of many medical drugs, with consequences for interpersonal variation in drug efficacy and toxicity. However, quantifying microbial contributions to drug metabolism is challenging, particularly in cases where host and microbiome perform the same metabolic transformation. We combined gut commensal genetics with gnotobiotics to measure brivudine drug metabolism across tissues in mice that vary in a single microbiome-encoded enzyme. Informed by these measurements, we built a pharmacokinetic model that quantitatively predicts microbiome contributions to systemic drug and metabolite exposure, as a function of bioavailability, host and microbial drug-metabolizing activity, drug and metabolite absorption, and intestinal transit kinetics. Clonazepam studies illustrate how this approach disentangles microbiome contributions to metabolism of drugs subject to multiple metabolic routes and transformations.

Mutagen Synergy: Hypermutability Generated by Specific Pairs of Base Analogs.

UNLABELLED: We tested pairwise combinations of classical base analog mutagens in Escherichia coli to study possible mutagen synergies. We examined the cytidine analogs zebularine (ZEB) and 5-azacytidine (5AZ), the adenine analog 2-aminopurine (2AP), and the uridine/thymidine analog 5-bromodeoxyuridine (5BrdU). We detected a striking synergy with the 2AP plus ZEB combination, resulting in hypermutability, a 35-fold increase in mutation frequency (to 53,000 × 10(-8)) in the rpoB gene over that with either mutagen alone. A weak synergy was also detected with 2AP plus 5AZ and with 5BrdU plus ZEB. The pairing of 2AP and 5BrdU resulted in suppression, lowering the mutation frequency of 5BrdU alone by 6.5-fold. Sequencing the mutations from the 2AP plus ZEB combination showed the predominance of two new hot spots for A·T→G·C transitions that are not well represented in either single mutagen spectrum, and one of which is not found even in the spectrum of a mismatch repair-deficient strain. The strong synergy between 2AP and ZEB could be explained by changes in the dinucleoside triphosphate (dNTP) pools. IMPORTANCE: Although mutagens have been widely studied, the mutagenic effects of combinations of mutagens have not been fully researched. Here, we show that certain pairwise combinations of base analog mutagens display synergy or suppression. In particular, the combination of 2-aminopurine and zebularine, analogs of adenine and cytidine, respectively, shows a 35-fold increased mutation frequency compared with that of either mutagen alone. Understanding the mechanism of synergy can lead to increased understanding of mutagenic processes. As combinations of base analogs are used in certain chemotherapy regimens, including those involving ZEB and 5AZ, these results indicate that testing the mutagenicity of all drug combinations is prudent.

A CO-FISH assay to assess sister chromatid segregation patterns in mitosis of mouse embryonic stem cells.

Sister chromatids contain identical DNA sequence but are chiral with respect to both their helical handedness and their replication history. Emerging evidence from various model organisms suggests that certain stem cells segregate sister chromatids nonrandomly to either maintain genome integrity or to bias cellular differentiation in asymmetric cell divisions. Conventional methods for tracing of old vs. newly synthesized DNA strands generally lack resolution for individual chromosomes and employ halogenated thymidine analogs with profound cytotoxic effects on rapidly dividing cells. Here, we present a modified chromosome orientation fluorescence in situ hybridization (CO-FISH) assay, where identification of individual chromosomes and their replication history is achieved in subsequent hybridization steps with chromosome-specific DNA probes and PNA telomere probes. Importantly, we tackle the issue of BrdU cytotoxicity and show that our method is compatible with normal mouse ES cell biology, unlike a recently published related protocol. Results from our CO-FISH assay show that mitotic segregation of mouse chromosome 7 is random in ES cells, which contrasts previously published results from our laboratory and settles a controversy. Our straightforward protocol represents a useful resource for future studies on chromatid segregation patterns of in vitro-cultured cells from distinct model organisms.

Selective initiation and transmission of disseminated neoplasia in the soft shell clam Mya arenaria dependent on natural disease prevalence and animal size.

Disseminated neoplasia, a diffuse tumor of the hemolymph system, is one of the six most destructive diseases among bivalve mollusk populations, characterized by the development of abnormal, rounded blood cells that actively proliferate. Though the specific etiology of disseminated neoplasia in Mya arenaria remains undetermined, the involvement of viral pathogens and/or environmental pollutants has been suggested and considered. The current study used 5-bromodeoxyuridine (BrDU) known to induce the murine leukemia virus and filtered neoplastic hemolymph to initiate disseminated neoplasia in clams from different populations and size classes respectively. M. arenaria from three locations of different natural neoplasia occurrences were divided into a control and three experimental treatments and injected with 200 µl of sterile filtered seawater or 50-200 µg/ml BrDU respectively. In a concurrent experiment, animals from different size classes were injected with 2.5% total blood volume of 0.2 µm filtered blood from a fully neoplastic animal. Animals were biopsied weekly and cell neoplasia development was counted and scored as 0-25, 26-50, 51-75 and 76-100% neoplastic hemocytes (stages 1-4) in 50 µl samples. BrDU injection demonstrated that neoplasia development in M. arenaria was dose dependent on BrDU concentration. In addition, natural disease prevalence at the source location determined initiation of neoplasia induction, with animals from the area of the highest natural disease occurrence displaying fastest neoplasia development (p=0.0037). This could imply that depending on the natural disease occurrence, a potential infectious agent may remain dormant in normal (stage 1) individuals in higher concentrations until activated, i.e. through chemical injection or potentially stress. The size experiment demonstrated that only M. arenaria between 40 and 80 mm developed 26-100% neoplastic hemocytes when injected with filtered neoplastic hemolymph, indicating that individuals smaller than 20mm or larger than 80 mm were not or no longer susceptible to disease development. So far neoplasia studies have not considered natural disease prevalence or size involvement in neoplasia development and our results indicate that these should be future considerations in neoplasia examinations.

A mammalian-like DNA damage response of fission yeast to nucleoside analogs.

Nucleoside analogs are frequently used to label newly synthesized DNA. These analogs are toxic in many cells, with the exception of the budding yeast. We show that Schizosaccharomyces pombe behaves similarly to metazoans in response to analogs 5-bromo-2'-deoxyuridine (BrdU) and 5-ethynyl-2'-deoxyuridine (EdU). Incorporation causes DNA damage that activates the damage checkpoint kinase Chk1 and sensitizes cells to UV light and other DNA-damaging drugs. Replication checkpoint mutant cds1Δ shows increased DNA damage response after exposure. Finally, we demonstrate that the response to BrdU is influenced by the ribonucleotide reductase inhibitor, Spd1, suggesting that BrdU causes dNTP pool imbalance in fission yeast, as in metazoans. Consistent with this, we show that excess thymidine induces G1 arrest in wild-type fission yeast expressing thymidine kinase. Thus, fission yeast responds to nucleoside analogs similarly to mammalian cells, which has implications for their use in replication and damage research, as well as for dNTP metabolism.

Organ differences in the impact of p27(kip1) deficiency on carcinogenesis induced by N-methyl-N-nitrosourea.

To evaluate the impact of p27 on carcinogenesis in various organs, N-methyl-N-nitrosourea (MNU), a direct-acting alkylating agent, was given to p27 knock-out mice. Groups of 20-40 male and female mice with null, hetero- or wild-type p27 alleles were given drinking water containing 240 ppm MNU or distilled water every other week for five cycles. The incidence and multiplicity of the induced proliferative lesions were then histologically evaluated at weeks 14 and 20. MNU treatment induced various lesions including squamous hyperplasia and squamous cell carcinoma in the forestomach, atypical hyperplasia and adenocarcinomas in the fundic and pyloric glands, adenomas and adenocarcinomas in the duodenum, malignant lymphomas in the thymus, liver, kidney and spleen and alveolar hyperplasia, adenomas, adenocarcinomas and malignant lymphomas in the lung. Although the incidences of the lesions in the forestomach, fundic and pyloric glands did not differ among the p27 genotypes, those of alveolar hyperplasia of the lung and malignant lymphoma of the thymus were significantly increased in p27-null males as compared with both wild- and hetero-type animals. Moreover, in both p27(+/+) and p27(+/-) cases, the rates for p27-positive cells were obviously increased in proliferative lesions of the pyloric gland and the lung. However, an increased rate of p27-positive cells was not observed in malignant lymphoma of the thymus. These findings suggest that p27 does not control the cell cycle equally in all organs affected by MNU-induced carcinogenesis.

Abnormal brain function of the rat neonate in a prenatal 5-bromo-2'-deoxyuridine (BrdU)-induced developmental disorder model.

Neonatal brain function was investigated in a prenatal BrdU-induced developmental disorder model, which has been reported to exhibit behavioral abnormalities such as locomotor hyperactivity, impaired learning and memory, and lower anxiety in offspring. After 1h home cage deprivation we observed an increase in the number of c-Fos (neuronal activity marker) immunoreactive cells in several brain regions of the olfactory and stress-related areas in normal neonates at 11 days. Next, pregnant rats were exposed to 50mg/kg of BrdU from gestation days 9-15, and their offspring at 11 days were home-cage deprived. Compared to vehicle control, the number of c-Fos immunoreactive cells in BrdU group was found to be decreased in the piriform cortex and locus coeruleus, which are known to play an important role in neonatal learning and memory. We also analyzed Pearson product-moment correlation coefficient of the number of c-Fos immunoreactive cells, focusing on the piriform cortex and locus coeruleus versus numerous other brain areas (11 areas including amygdala). Numerous significant correlations were observed in the vehicle control group, however, correlations of the locus coeruleus disappeared in the BrdU group. By observing c-Fos immunoreactivity after home cage deprivation our study uncovers abnormal brain functions as early as postnatal day 11 in this disorder model. Based on these results, we propose a new histological approach for functional characterization of developmental disorder models.

Unusual morphological damage of Purkinje cells following postnatal BrdU administration in the cerebellar cortex of mouse.

Postnatal development of the cerebellum lasts for weeks in rodents and can be disturbed by systemic 5-bromo-2'-deoxyuridine (BrdU) administration. This thymidine analogue incorporates into the DNA of proliferating cells, and result in more or less serious damage or death granule cells, the most actively dividing neuronal population in the developing cerebellar cortex. Further consequences of postnatal BrdU administration are the interrupted postnatal migration and integrations as well as partial loss of cerebellar Purkinje cells. In the present study, C57B16 mice were administered with 50 µg/g body weight BrdU, one sc. injection daily, between P0 and P11 postnatal days, respectively.Large "cavities" appeared in the cytoplasm of a subpopulation of Purkinje cells by P7 in about one-third of administered animals, their number are size of the cavities (and PCs exhibiting unusual morphology) decreased. EM studies revealed that the unusual Purkinje cells received numerous axonal inputs of unknown origin, first of all on their somatic and dendritic spines. The transitory appearance of a subpopulation of Purkinje cells possessing unusual morphology refers to the influence of other (neuronal, glial, or both) cells on their regular differentiation.

Somite unit chronometry to analyze teratogen phase specificity in the paraxial mesoderm.

Phase specificity, the temporal and tissue restriction of teratogen-induced defects during embryonic -development, is a poorly understood but common property of teratogens, an important source of human birth defects. Somite counting and somite units are novel chronometric tools used here to identify stages of paraxial mesoderm development that are sensitive to pulse-chase exposure (2 to >16 h) to 5-bromodeoxyuridine (BrdU). In all cases, it was the presomitic mesoderm (PSM) that was sensitive to BrdU induced segmentation anomalies. At high concentration (1.0 × 10(-2) M BrdU), PSM presegment stages ss-IV and earlier were irreversibly inhibited from completing segmentation. At low concentration (2.6 × 10(-6) M), BrdU induced periodic focal defects that predominantly trace back to PSM presegments between ss-V and ss-IX. Phase specificity is characteristic of both types of segmentation anomalies. Focal segmentation defects are phase-specific because they result from disruption of 2-3 presegments in the PSM while adjacent -rostral and caudal presegments are (apparently) unaffected. Irreversible inhibition of segmentation is also phase-specific because only PSM presegments ss-IV or earlier were affected while older segments (ss-III to ss-I) were able to complete segmentation. The presegments predominantly affected have not yet passed the determination front, the point at which the segmentation clock establishes somite rostro-caudal -polarity. Somite unit chronometry provides a means to identify specific PSM presegment stages that are susceptible to induced segmentation defects and the biological processes that underlie that vulnerability.

Hyperactivity induced by prenatal BrdU exposure across several experimental conditions.

Behavioral results are sometimes not reproducible even in the positive controls of developmental neurotoxicity (DNT) tests. Effects of several factors on the results should be considered. In the present paper, we examined the effects of strain-, gender-, and test-condition differences on BrdU-induced hyperactivity. The results showed that BrdU-induced hyperactivity was reproducible in two rat strains (SD and F344 rats), rodent species (rat and mouse), and both sexes. When the level of background sound in a test room was increased, the hyperactivity was persistent, resulting in no effect of background sound on BrdU-induced hyperactivity. Thus, we have demonstrated that the BrdU-animal model is a useful positive control via prenatal exposure to validate the entire DNT test process.

Systemic 5-bromo-2-deoxyuridine induces conditioned flavor aversion and c-Fos in the visceral neuraxis.

5-bromo-2-deoxyuridine (BrdU) is often used in studies of adult neurogenesis and olfactory learning, but it can also have toxic effects on highly proliferative tissue. We found that pairing Kool-Aid flavors with acute systemic injections of BrdU induced strong conditioned flavor aversions. Intermittent injections during Kool-Aid-glucose conditioning interfered with learning of a conditioned flavor-nutrient preference. Acute injection of BrdU also elevated plasma corticosterone levels and induced c-Fos in the visceral neuraxis. Thus, acute or intermittent systemic injections of BrdU (50-200 mg/kg) have aversive effects that may interfere with learning.

Acute hepatitis due to brivudin: a case report.

BACKGROUND/AIMS: Brivudin is licensed in several European countries for the treatment of herpetic infections, and is considered safe (approximately 1% of patients with transient elevation of liver enzymes) in large multicenter trials. METHODS: We report a case of acute brivudin hepatitis documented with a liver biopsy in detail. RESULTS: Liver biopsy demonstrated acute liver injury with a predominant cytolytic pattern and features suggestive of a drug-induced immunoallergic hepatitis. Elevated ALT levels returned to normal within weeks. CONCLUSIONS: This is the first published case of acute immunoallergic hepatitis due to brivudin.

Extracellular ADP regulates lesion-induced in vivo cell proliferation and death in the zebrafish retina.

Regeneration and growth that occur in the adult teleost retina by neurogenesis have been helpful in identifying molecular and cellular mechanisms underlying cell proliferation and differentiation. In this report, we demonstrate that endogenous purinergic signals regulate cell proliferation induced by a cytotoxic injury of the adult zebrafish retina which mainly damages inner retinal layers. Particularly, we found that ADP but not ATP or adenosine significantly enhanced cell division as assessed by 5-bromo-2'-deoxyuridine incorporation following injury, during the degenerative and proliferative phase of the regeneration process. This effect of ADP occurs via P2Y1 metabotropic receptors as shown by intra-ocular injection of selective antagonists. Additionally, we describe a role for purinergic signals in regulating cell death induced by injury. Scavenging of extracellular nucleotides significantly increased cell death principally seen in the inner retinal layers. This effect is partially reproduced by blocking P2Y1 receptors suggesting a neuroprotective function for ADP, which is derived from extracellular ATP probably released by dying cells as a consequence of the ouabain treatment. This study demonstrates a crucial role for ADP as a paracrine signal in the repair of retinal tissue following injury.

Cell type specific applicability of 5-ethynyl-2'-deoxyuridine (EdU) for dynamic proliferation assessment in flow cytometry.

Using the nucleoside analogue EdU (5-ethynyl-2'-deoxyuridine) for thymidine substitution instead of BrdU (5-bromo-2'-deoxyuridine) in cell proliferation assays has recently been proposed. However, the effect of EdU on cell viability, DNA synthesis, and cell cycle progression and consequently its usability for dynamic cell proliferation analysis in vitro has not been explored. We compared the effect of EdU and BrdU incorporation into SK-BR-3 and BT474 breast cancer cells and the impact on cell cycle kinetics, cell viability, and DNA damage. We found that EdU can be used not only for pulse but also for continuous cell labeling and henceforth in high resolution EdU/Hoechst quenching assays. BrdU and EdU proliferation assays based on click chemistry revealed comparable results. However, cell viability of SK-BR-3 breast cancer cells was highly affected by long term exposure to EdU. Both SK-BR-3 as well as BT474 cells show cell cycle arrests upon long term EdU treatment whereas only SK-BR-3 cells were driven into necrotic cell death by long term exposure to EdU. In contrast BT474 cells appeared essentially unharmed by EdU treatment in terms of viability. Consequently using EdU enables highly sensitive and quantitative detection of proliferating cells and facilitates even continuous cell cycle assessment. Nevertheless, potential cellular susceptibility needs to be individually evaluated.

Does 5-bromo-2'-deoxyuridine (BrdU) disrupt cell proliferation and neuronal maturation in the adult rat hippocampus in vivo?

5-Bromo-2'-deoxyuridine (BrdU) is frequently used as a mitotic marker in studies of cell proliferation. Recent studies have reported cytotoxic effects of BrdU on neural progenitor cells in embryonic and neonatal brains in vivo and in adult tissue studied in vitro. The present study was conducted to assess whether BrdU interferes with cell proliferation and neuronal maturation in the rat adult hippocampus in vivo. BrdU effects across a wide range of doses (40-480 mg/kg) on cell proliferation and the population of immature neurons in the adult hippocampus were investigated using immunohistochemical labeling methods for the cell cycle marker Ki67 and a marker for immature neurons, doublecortin. BrdU did not influence cell proliferation in the dentate gyrus or the population of immature neurons observed in the adult hippocampus relative to those observed in saline treated controls. Thus, in contrast with reports of deleterious effects of BrdU in embryonic and neonatal tissue and adult tissue studied in vitro, BrdU does not appear to have cytotoxic effects on proliferating hippocampal cells or immature neurons in vivo in rats.

The oxidative stress response is region specific in organogenesis stage mouse embryos exposed to 5-bromo-2'-deoxyuridine.

BACKGROUND: The exposure of mouse embryos to 5-bromo-2'-deoxyuridine (BrdU), a model teratogen, generates oxidative stress, induces c-Fos dependent activator protein-1 (AP-1) DNA binding activity, and causes skeletal malformations (Sahambi and Hales, 2006). The goal of this study was to test the hypothesis that the ability of BrdU to induce oxidative stress, rather than its incorporation into DNA per se, is responsible for triggering the ensuing malformations. To test this hypothesis, we examined the regional localization of BrdU incorporation, 8-oxoguanine (8-oxoG), a marker of oxidative stress, and c-Fos immunoreactivity in organogenesis stage mouse embryos exposed to a teratogenic dose of BrdU. METHODS: Timed pregnant CD1 mice received vehicle (saline) or BrdU (600 or 1000 mg/kg body weight) on gestation day 9. N-acetyl-L-cysteine, a glutathione precursor, was administered to GD 9 mice 2 hours before treatment with vehicle or BrdU to determine the impact of inhibiting the oxidative stress response. Embryos were excised 3 hours after BrdU treatment and processed for staining for BrdU incorporated into DNA, 8-oxoG, and c-Fos. RESULTS: BrdU incorporation, 8-oxoG adduct formation and c-Fos immunoreactivity were highest in the rostral and caudal developing tissues of BrdU-exposed embryos. Although preadministration of N-acetyl-L-cysteine at a dose that decreased BrdU teratogenicity dampened the oxidative stress response, it did not affect the incorporation of BrdU into DNA. CONCLUSIONS: These data show that rostral and caudal neuroepithelial cells are particularly susceptible to oxidative stress in the organogenesis-stage embryo.

Characteristic behavioral anomalies in rats prenatally exposed to 5-bromo-2'-deoxyuridine.

The aim of the present study was to characterize behavioral anomalies in rats prenatally exposed to 5-bromo-2'-deoxyuridine, a useful model of hyperactive disorder. Rats were treated with BrdU at 50mg/kg IP or carboxymethylcellulose, its vehicle, on gestational Days 9 through 15, and their offsprings were subjected to behavioral tests. Rats prenatally exposed to 5-bromo-2'-deoxyuridine showed higher locomotor activity levels when the lights were turned off, and these levels kept increasing throughout the dark cycle. In an elevated plus maze, the rats prenatally exposed to 5-bromo-2'-deoxyuridine exhibited decreased anxiety-related behavior, including higher open arm entries and a longer time spent per one open arm entry when compared with rats prenatally exposed to carboxymethylcellulose. Methylphenidate, a psychostimulant that suppresses hyperactivity in humans with attention-deficit hyperactivity disorder, increased locomotor activity in both rats, with a greater sensitivity in rats prenatally exposed to 5-bromo-2'-deoxyuridine. Desipramine, a specific noradrenaline uptake inhibitor, normalized the hyperactivity of rats prenatally exposed to 5-bromo-2'-deoxyuridine. Paroxetine, a selective serotonin reuptake inhibitor, also normalized the hyperactivity and the low anxiety-related behavior in the elevated plus maze. These results suggest that rats prenatally exposed to 5-bromo-2'-deoxyuridine are hyperactive and exhibit a lower anxiety level. Dysfunctional monoaminergic neurons may be, at least in part, the cause of the behavioral anomalies.

Retardation in somatosensory cortex development induced by postnatal BrdU treatment in mice.

Cerebral dysgeneses are in the background of several neurological and mental disturbances. The aim of the present study was to investigate structural and activity changes following disturbed postnatal neuronal development in mice. Newborn C57Bl6 mice were exposed to 5-bromo-2'-deoxyuridine (BrdU: daily 50 microg/g body weight) during a period between postnatal days P0-P5 or P0-P11, respectively, and neuronal malformation and malfunctioning of somatosensory (barrel field) cortex was analyzed in adolescent animals. Alterations in histological architecture of interneuronal and glial elements were studied and correlated with electrophysiological modifications. Between P30 and P35 days litters underwent ex vivo electrophysiological experiments to examine the changes in basic excitability and in synaptic efficacy. Parallel immunohistochemistry was performed to detect BrdU, GABA and GFAP. There were no BrdU immunopositive cell nuclei in control animals, but marked staining was observed in both BrdU treated groups. Lessening in the number of GABAergic neurons was observed in the treated groups. GFAP immunohistochemical analysis has shown an increased number of activated astroglial cells in treated animals. Reduction of the number of GABAergic neurons was observed in the treated groups. Electrophysiological recordings on cortical slices showed increased excitability in the treated groups.