Genetic analysis of dopaminergic system development in zebrafish.

Zebrafish have become an important model organism to study the genetic control of vertebrate nervous system development. Here, we present an overview on the formation of dopaminergic neuronal groups in zebrafish and compare the positions of DA neurons in fish and mammals using the neuromere model of the vertebrate brain. Based on mutant analysis, we evaluate the role of several signaling pathways in catecholaminergic neuron specification. We further discuss the prospect of identifying novel genes involved in dopaminergic development through forward genetics mutagenesis screens.

Genetic disruption of Ptgs-1, as well as Ptgs-2, reduces intestinal tumorigenesis in Min mice.

Two isoforms of cyclooxygenase (COX) are known, and to date most studies have implicated COX-2, rather than COX-1, as the isoform involved in colon carcinogenesis. In the present study, we show that homologous disruption of either Ptgs-1 or Ptgs-2 (genes coding for COX-1 or COX-2, respectively) reduced polyp formation in Min/+ mice by approximately 80%. Only COX-1 protein was immunohistochemically detected in normal intestinal tissue, whereas both COX-1 and variable levels of COX-2 protein were detected in polyps. Prostaglandin E2 was increased in polyps compared with normal tissue, and both COX-1 and COX-2 contributed to the PGE2 produced. The results indicate that COX-1, as well as COX-2, plays a key role in intestinal tumorigenesis and that COX-1 may also be a chemotherapeutic target for nonsteroidal anti-inflammatory drugs.

Mapping synaptic glutamate transporter dysfunction in vivo to regions surrounding Aß plaques by iGluSnFR two-photon imaging.

Amyloid-ß (Aß) plaques, a hallmark of Alzheimer's disease (AD), are surrounded by regions of neuronal and glial hyperactivity. We use in vivo two-photon and wide-field imaging of the glutamate sensor iGluSnFR to determine whether pathological changes in glutamate dynamics in the immediate vicinity of Aß deposits in APPPS1 transgenic mice could alter neuronal activity in this microenvironment. In regions close to Aß plaques chronic states of high spontaneous glutamate fluctuations are observed and the timing of glutamate responses evoked by sensory stimulation exhibit slower decay rates in two cortical brain areas. GLT-1 expression is reduced around Aß plaques and upregulation of GLT-1 expression and activity by ceftriaxone partially restores glutamate dynamics to values in control regions. We conclude that the toxic microenvironment surrounding Aß plaques results, at least partially, from enhanced glutamate levels and that pharmacologically increasing GLT-1 expression and activity may be a new target for early therapeutic intervention.

Normalization of blunted lymphocyte beta-adrenergic responsivity in melancholic inpatients by a course of electroconvulsive therapy.

Electroconvulsive therapy has been reported to desensitize brain beta-adrenergic receptors in rodents, but this effect has not been studied in man. We examined the effect of a course of electroconvulsive therapy on lymphocyte beta-adrenergic responsivity in 19 inpatients with melancholia. Before treatment, beta-adrenergic cyclic adenosine monophosphate response to isoproterenol was significantly blunted in the patients compared with controls. Following a course of electroconvulsive therapy, beta-adrenergic responsivity increased such that patients no longer differed from controls. Thus, blunted lymphocyte beta-adrenergic responsivity is a state-dependent effect of melancholia that can be corrected by a therapeutic course of electroconvulsive therapy. The effect of electroconvulsive therapy on this beta-adrenergic system is in the opposite direction to that reported for rodent forebrain, where electroconvulsive therapy causes desensitization, and may reflect differences between peripheral and central effects, species differences, or disease effects.

Safe biotechnology 9: values in risk assessment for the environmental application of microorganisms. The Safety in Biotechnology Working Party of the European Federation of Biotechnology.

Risk assessment for the deliberate release of microorganisms into the environment is traditionally carried out on a case-by-case basis. In a similar approach to that used when assessing human pathogenicity, we propose an alternative approach by introducing risk classes to facilitate or complement this type of risk assessment. These consider several sets of scenarios that address the different values that need to be protected. Examples of this approach include risk-class definitions for soil fertility and biodiversity.

Photocarcinogenesis and susceptibility to UV radiation in the v-Ha-ras transgenic Tg.AC mouse.

The v-Ha-ras transgenic Tg.AC mouse line has proven to be a useful model for the study of chemical carcinogenic potential. We undertook experiments designed to study the effect of the physical carcinogen, UV radiation, on tumorigenesis in this mouse strain. Following a total of three exposures on alternating days to a radiation source covering a cumulative UVR exposure range of 2.6-42.6 kJ per m2, squamous papillomas developed by 4 wk after initial exposure in a dose-dependent manner. Malignancies developed within 18-30 wk following the initial UVR exposure and were all diagnosed as squamous cell carcinoma or spindle cell tumors. In contrast to other mouse stains used in photocarcinogenesis studies, few p53 mutations were found in Tg.AC malignancies upon polymerase chain reaction-single stranded conformational polymorphism analysis of exons 4-8 followed by sequencing of suspicious bands; however, all tumors analyzed by in situ hybridization expressed the v-Ha-ras transgene. Immunohistochemical analysis of UVR-exposed skin taken 24 h after the last of three exposures (13.1 kJ per m2 total UVR) showed expression of p53 in hair follicles and in interfollicular epidermis, which indicates that the gene was functional. Thus, although there are some differences between the Tg.AC and other mouse models, these results suggest that the Tg.AC mouse may be a useful model for the study of acute exposure photocarcinogenesis.

Undetected HIV infection among patients admitted to an alcohol rehabilitation unit.

Of 300 patients admitted to an alcohol rehabilitation unit, serological testing of discarded admission blood samples revealed that 31 (10.3%) had HIV infection. Chart records indicated that four patients were known to be HIV infected on admission, and HIV infection was detected in another three patients during hospitalization; however, 24 (77.4%) of the HIV-positive patients were discharged with their HIV infection still undetected.

Cell type-specific lectin staining of the tracheobronchial epithelium of the rat: quantitative studies with Griffonia simplicifolia I isolectin B4.

We compared lectin staining patterns to cell population densities, as determined by morphological criteria in rat airways. Eight lectins were studied: Griffonia simplicifolia I isolectin B4 (GSI-B4), Arachis hypogaea (PNA), Wisteria floribunda (WFA), Glycine maximus (SBA), Dolichos biflorus (DBA), Helix pomatia (HPA), Ulex europaeus (UEA-1), and Maclura pomifera (MPA). Two of the lectins strongly stained morphologically distinct cell subpopulations. GSI-B4 stained basal cells, and MPA stained non-ciliated bronchiolar (Clara) cells. The specificity and sensitivity of GSI-B4 as a marker for basal cells was examined. In the trachea, 35% of all cells were GSI-B4 positive; 84% of these were basal cells, 7% were unidentified cells, 5% were serous/mucous cells, and 4% were ciliated, brush, or inflammatory cells. Comparison to cell population density data strongly suggested that all basal cells were GSI-B4 positive. The segmental bronchus was a transitional area; GSI-B4 positive basal cells were present in the region closest to the lobar bronchus but were absent in the distal region; instead, MPA-positive Clara cells appeared. When dissociated tracheal cells were obtained by pronase digestion, 43% were GSI-B4 positive. These results show that GSI-B4 is a sensitive and relatively specific marker for basal cells in the rat trachea which can be used to study dissociated epithelial cells.

Argon laser trabeculotomies in primates: evaluation by histological and perfusion studies.

The continuous argon laser beam has been applied to the trabeculum at the anterior chamber angle of one eye of nine primates. The treatment was evaluated by light and scanning electron microscopy and constant rate infusion studies at different intervals following the laser treatment. This histological studies revealed coagulative necrosis, with trabeculotomies extending into Schlemm's canal in the immediate postoperative period. However, complete closure of the laser-induced trabeculotomies by healing was observed within 1 to 3 weeks following the treatment. Outflow facility data as measured by the perfusion studies revealed somewhat increased values of the treated eye during the early 2 weeks after laser treatment. On longer follow-up, no significant outflow differences were measured between the treated and untreated eyes.

Characterization of hepatocellular resistance and susceptibility to styrene toxicity in B6C3F1 mice.

Short-term inhalation exposure of B6C3F1 mice to styrene causes necrosis of centrilobular (CL) hepatocytes. However, in spite of continued exposure, the necrotic parenchyma is rapidly regenerated, indicating resistance by regenerated cells to styrene toxicity. These studies were conducted to test the hypothesis that resistance to repeated styrene exposure is due to sustained cell proliferation, with production of hepatocytes that have reduced metabolic capacity. Male mice were exposed to air or 500 ppm styrene (6 h/day); hepatotoxicity was evaluated by microscopic examination, serum liver enzyme levels, and bromodeoxyuridine (BrdU)-labeling index (LI). Metabolism was assessed by measurement of blood styrene and styrene oxide. Both single and repeated exposures to styrene resulted in mortality by Day 2; in mice that survived, there was CL necrosis with elevated BrdU LI at Day 6, and complete restoration of the necrotic parenchyma by Day 15. The BrdU LI in mice given a single exposure had returned to control levels by Day 15. Re-exposure of these mice on Day 15 resulted in additional mortality and hepatocellular necrosis, indicating that regenerated CL cells were again susceptible to the cytolethal effect of styrene following a 14-day recovery. However, in mice repeatedly exposed to styrene for 14 days, the BrdU LI remained significantly increased on Day 15, with preferential labeling of CL hepatocytes with enlarged nuclei (karyomegaly). If repeated exposures were followed by a 10-day recovery period, CL karyomegaly persisted, but the BrdU LI returned to control level and CL hepatocytes became susceptible again to styrene toxicity as demonstrated by additional mortality and acute necrosis after a challenge exposure. These findings indicated a requirement for continued styrene exposure and DNA synthesis in order to maintain this resistant phenotype. Analyses of proliferating-cell nuclear-antigen (PCNA) labeling were conducted to further characterize the cell cycle kinetics of these hepatocytes. The proportion of cells in S-phase was increased by repeated exposure. However, PCNA analysis also revealed an even larger increase in the G1 cell compartment with repeated exposures, without a concurrent increase in G2 phase or in mitotic cell numbers. These data indicate that resistance to styrene-induced necrosis under conditions of repeated exposure is not due to sustained cell turnover and production of new, metabolically inactive cells, but rather is due to some other, as yet unknown, protective phenotype of the regenerated cells.

Characterization of inhaled alpha-methylstyrene vapor toxicity for B6C3F1 mice and F344 rats.

alpha-Methylstyrene (AMS) is a chemical intermediate used in the synthesis of specialty polymers and copolymers. Inhalation studies of AMS were conducted because of the lack of toxicity data and the structural similarity of AMS to styrene, a toxic and potentially carcinogenic chemical. Male and female B6C3F1 mice were exposed to 0, 600, 800, or 1000 ppm AMS 6 h/day, 5 days/week, for 12 days. After 1 exposure, 21% (5/24) of female mice were found dead in the 1000-ppm group, 56% (10/18) in the 800-ppm group, and 6% (1/18) in the 600-ppm concentration group. After 12 exposures, relative liver weights were significantly increased and relative spleen weights were significantly decreased in both male and female mice at all concentrations. No microscopic treatment-related lesions were observed. A decrease in hepatic glutathione (GSH) was associated with AMS exposure for 1 and 5 days. Male and female F344 rats were exposed to 0, 600 or 1000 ppm AMS for 12 days. No mortality or sedation occurred in AMS-exposed rats. Relative liver weights were significantly increased in both males and females after 12 exposures to 600 or 1000 ppm. An increased hyaline droplet accumulation was detected in male rats in both concentration groups; no significant microscopic lesions were observed in other tissues examined. Exposure of male and female F344 rats and male NBR rats to 0, 125, 250 or 500 ppm AMS, 6 h/day for 9 days resulted in increased accumulation of hyaline droplets in the renal tubules of male F344 rats in the 250 and 500 ppm concentration groups. Although AMS and styrene are structurally very similar, AMS was considerably less toxic for mice and more toxic for male rats than styrene.

New issues and future legislation on biosafety.

The current debate on gene technology in Europe is reviewed with particular emphasis on the role of EFB and science in general. From this debate important lessons have been learned by scientists, industrial companies, legislators, parliamentarians and interest organisations. This gives confidence that the continued debate will eventually lead to public acceptance of gene technology even in the food area.

Enhancing treatment gains in a school-based intervention for children of divorce through skill training, parental involvement, and transfer procedures.

The school-based Children's Support Group procedure teaches skills to cope with divorce-related events and provides strategies for mastering disrupted developmental tasks. Ss were 103 3rd-through 5th-grade children of separated or divorced parents who were assigned to 1 of 3 treatment groups: support; support and skill building; support, skill building, transfer, and parent training procedures; or no-treatment control. Twenty-six children from intact homes served as nonstressed controls. The two skill-building conditions yielded durable improvements in adjustive behaviors in the home. Transfer components yielded additional improvements in affect, but the absence of substantial increments in benefits suggests the need for a closer look at the format and expectations of the transfer vehicle. The benefits of the support-alone condition were experienced most by children who entered the intervention with significant problems, with the greatest reductions in clinical symptomatology at follow-up being found in this group.

Toxicity and carcinogenicity of riddelliine following 13 weeks of treatment to rats and mice.

Toxicity studies of riddelliine, a member of a class of pyrrolizidine alkaloids, were conducted because riddelliine has been found to contaminate human food sources. Groups of male and female Fischer rats were administered riddelliine by gavage in phosphate buffer at doses up to 10 mg/kg, and B6C3F1 mice at doses up to 25 mg/kg, five times a week. The animals were necropsied after 13 weeks of treatment or after a 7 or 14 week recovery period. Body weight gains were inversely related to dose in both rats and mice. Body weight of the 1.0 and 3.3 mg/kg female rats and 10.0 and 25.0 mg/kg mice remained depressed during the 14 week recovery period. At 13 weeks, significant findings included dose-related hepatopathy and intravascular macrophage accumulation in rats and hepatocytomegaly in mice. During the 14 week recovery period these lesions persisted and hepatic foci of cellular alteration in male rats and bile duct proliferation in female rats and male and female mice increased in severity. In the 10 mg/kg group of female rats adenomas of the liver occurred in two of ten at 13 weeks and in one of five at the 14 week recovery period. In separate studies, the frequency of micronucleated erythrocytes in peripheral blood was increased in male mice administered a single dose (150 mg/kg) of riddelliine. Increases in unscheduled DNA and S-phase syntheses were detected in primary hepatocytes from rats and mice treated with riddelliine at doses up to 25.0 mg/kg for 5 or 30 days. In mating trials in rats and mice, pup weights from treated dams at birth and during suckling were lower than controls. Thus, riddelliine is genotoxic and carcinogenic and may cross the placenta and/or be found in milk, causing developmental toxicity in rodents.

Odontogenic tumours in the v-Ha-ras (TG.AC) transgenic mouse.

A line of homozygous transgenic mice (TG.AC) carrying a v-Ha-ras gene fused to the promoter of the zeta globin gene produces a variety of mesenchymal and epithelial neoplasms including odontogenic tumours. The 1-year incidence of odontogenic tumour formation in these mice was approx. 35%. Tumours formed more often in the mandible than maxilla. The various types of tumours frequently presented with: (1) primarily mesenchymal cells in a dense fibrous-like matrix, or (2) loose stroma surrounded by anastomosing cords of epithelial cells that exhibited squamous differentiation, or (3) odontomas forming mineralized tooth structures by well-differentiated odontoblasts and ameloblasts. Some tumours had areas with all three of these characteristics. Mineralized dentine and enamel in the odontomas were morphologically similar to those of normal murine teeth. Odontogenic tumours expressed the v-Ha-ras transgene that was primarily localized to the mesenchymal cells. Proliferating-cell nuclear antigen immunohistochemistry showed that the mesenchymal cells adjacent to the epithelial cords not only expressed the ras transgene but were also actively proliferating. The TG.AC mouse provides an excellent model for the study of odontogenic tumours and tooth development.

The safety assessment of novel foods. Guidelines prepared by ILSI Europe Novel Food Task Force.

The diversity of novel foods and novel ingredients covered by the scope of the EU regulation is such that a check list approach to safety evaluation is inappropriate. Rather, a case-by-case approach is required taking into account the composition of the novel food, its intake, its role in the diet and the intended target group. The SAFEST approach provides a means of targeting the safety evaluation on those aspects, nutritional or toxicological, of a novel food which are of particular concern. Using this approach, novel foods are assigned to one of three classes on the basis of certain background information. For those novel foods which can be shown to be in SAFEST class 1, namely those which are substantially equivalent to a traditional counterpart, no further information is required to demonstrate their safety. For those novel foods in SAFEST class 2, i.e. those sufficiently similar to a traditional counterpart or differing from it only in particular, well defined, characteristics, the evaluation will focus on those differences. Only in the case of novel foods which are not in class 1 or class 2 is extensive testing of the whole food likely to be required. Even in these cases, the testing should follow a scientifically-based hierarchical approach involving: literature reviews; chemical analysis; appropriate in vitro and in vivo tests; and, if necessary, confirmation of safety and nutritional value in humans. Examination of the causes of any adverse effects reported by consumers after the novel food or ingredient has been approved and is introduced into the market may provide additional reassurance of safety.

Addiction to and dependence on benzodiazepines. Diagnostic confusion in clinical practice and research studies.

Considerable confusion continues to surround basic concepts for abuse, addiction, tolerance, and dependence. Clinicians may be making decisions about prescribing these medications without clear definitions and distinctions. The terms are not equivalent in meaning and should not be used interchangeably in clinical application. Moreover, they may occur together or independently and are not etiologically related. Abuse is improper use outside the standard norms. Abuse implies a violation component and a control over the use of the drug. Addiction is a preoccupation with the acquisition and compulsive use of and a pattern of relapse to drugs is spite of adverse consequences. Pervasive to the criteria is a loss of control over drug use and a lack of volitional component in the drug use. In spite of problems in definitions, studies have clearly shown that abuse, addiction, tolerance, and dependence develop commonly in benzodiazepine use.

Racial differences in treatment of psychiatric inpatients.

OBJECTIVE: The study examined several aspects of inpatient psychiatric treatment to determine if differences existed between treatment of African-American and white patients. METHODS: Using a structured chart review, data were collected on 76 African-American and 88 white patients consecutively admitted to an acute inpatient setting with a principal axis I diagnosis of a major mood or psychotic disorder. Racial differences in treatment were examined using analysis of variance and logistic regression to assess the effects of diagnosis and socioeconomic status. RESULTS: Nonpsychotic African-American patients had shorter lengths of stay than white patients with similar disorders. White patients were more likely to be on one-to-one observational status. Clinicians were more likely to order urine drug screens for African-American patients with high socioeconomic status than for comparable white patients. African-American patients with schizophrenic disorders received higher neuroleptic dosages than white patients with similar diagnoses. CONCLUSIONS: Most racial differences cited in earlier studies of psychotic patients were not found or were not statistically significant once socio-economic status and diagnosis were accounted for. However, racial differences related to the detection, phenomenology, treatment, and course of psychotic disorders and the diagnosis and management of alcohol and drug use disorders and personality disorders were found.