Hindbrains Research Articles

Effect of glutathione depletion by buthionine sulfoximine on rat embryonic development in vitro

The intracellular thiol glutathione has many functions within cells including protection against xenobiotic and oxidative damage, and a role in protein and DNA synthesis and amino acid transport. Consequently, glutathione might be an important substance for normal growth and development. In this study the extent of glutathione depletion by buthionine sulfoximine, an agent which depletes glutathione by inhibiting its synthesis, and the subsequent effects of the depletion on rat embryonic growth and development were assessed. Day 10.5 rat embryos were cultured in rat serum medium in the presence of l-buthionine -S,R- sulfoximine (0.01 to 2.0 mM) and examined for viability, malformations, growth and development 45 hr later. The glutathione concentrations of the cultured embryos and their yolk sacs were also determined. Exposure to buthionine sulfoximine produced marked and significant (P ⩽ 0.05) depletion of glutathione at a buthionine sulfoximine concentration of 0.10 mM in the embryos and 0.05 mM in the yolk sacs. Exposure to 1 mM buthionine sulfoximine depleted glutathione to less than 7% of control in both of these tissues. None of the concentrations of buthionine sulfoximine tested had a significant effect on embryo viability; however, buthionine sulfoximine caused a significant (P ⩽ 0.05) incidence of malformed embryos at concentrations of 0.25, 0.5, 1.0 and 2.0 mM. The types of defects induced by buthionine sulfoximine were blebs of the maxillary or nasal processes, prosencephalon or forelimb buds, small or misshapen heads, small prosencephalons and swollen hind brains, and tail defects. Embryonic growth was the most sensitive, of the variables assessed, to the effects of buthionine sulfoximine. Significant (P ⩽ 0.05) growth retardation was observed at buthionine sulfoximine concentrations as low as 0.01 mM. At 2.0 mM buthionine sulfoximine, the yolk sac diameter, embryo crown-rump length, head length, number of somites and morphological score were reduced to 65, 72, 77, 90 and 80% of control levels respectively. We propose that the embryotoxic effects of buthionine sulfoximine are due to glutathione depletion and, consequently, that a certain basal level of endogenous glutathione is essential to allow for normal development.

Adrenocortical hormones and brain growth: Reversibility and differential sensitivity during development

The brain growth that follows adrenalectomy can be arrested by corticosterone replacement. Administered daily in combination with deoxycorticosterone, the brain and body conditions of intact rats are closely matched. Using this combination dose (corticosterone, 7.0 mg/kg; deoxycorticosterone, 1.0 mg/kg), hormonal replacement was systematically administered and withheld in a balanced design in order to assess the relative sensitivity of brain tissue to corticosteroids across time, and to determine if brain growth suppressed during one phase could recover upon hormone withdrawal during a second phase. Female forebrains were less sensitive to the hormones during an early phase which spanned ages 27 to 46 days. In contrast, 70 to 80% of potential growth was suppressed by replacement during a later period (ages 46 to 65 days). Brain size differences included weight and forebrain length, width, and depth. Hind brains were more sensitive to hormone replacement during phase 1. However, this suppressed growth was completely regained after hormone withdrawal during phase 2. On the other hand, growth that had occurred in the absence of adrenal steroids was not reversed, only arrested, by subsequent administration. We conclude that the rat brain became increasingly sensitive to the suppressive influence of adrenal steroids as females matured from juveniles to adults. Coupled with this sensitivity was the ability—apparently complete—to recover from steroid-induced brain growth suppression.

Partial purification of rabbit hind brain tryptophan hydroxylase by affinity chromatography

Chromatography of crude homogenates of rabbit hind brains on ε-amino caproyl-D-tryptophan methyl ester-agarose gels provide enzyme fractions with specific activity 7–10 times higher than the starting material. The activity was found to be associated with two distinct components. While nearly forty-fold increase in specific activity can be achieved by purification of the homogenate on calcium phosphate gels prior to affinity chromatography, only a single active component was noted in such prepurified extracts.

Characteristics of a coronavirus causing vomition and wasting in pigs.

SummarySome characteristics of a virus, isolated from the tonsils of 2 pigs with clinical signs of inappetence and vomition and designated VW572 were examined. It induced the formation of syncytia in primary pig kidney cell cultures, caused hemadsorption and hemagglutination using chicken, turkey, rat and mouse erythrocytes. In growth curve experiments, infectious virus was produced intracellularly starting at 6 hours after inoculation and was followed by rapid release of the virus from the infected cells. The virus contains ribonucleic acid, is ether sensitive and has a size between 100 and 220 nm. At 37° C, the infectivity titer decreased about 2 log10TCID50 per 24 hours. The viral population was heterogeneous as indicated by the rate of inactivation by U.V. irradiation and acid pH. Some hemagglutinating activity was left after complete loss of infectivity by ether-, temperature- and U.V. treatment. The VW 572 isolate is antigenically related if not identical to isolates from Canada, U.S.A. and England which were classified as a porcine coronavirus.Oronasal and intracerebral inoculation of the VW572 isolate in colostrum deprived pigs resulted in clinical disease after an incubation period of 6 and 4 days, respectively. Signs were characterized by depression, vomition, loss of appetence with rapid weakness in young pigs and vomition with progressive wasting in older pigs. Virus was isolated from nasal and pharyngeal swabs, nasal mucosa, tonsils, lungs and hind brains but not from rectal swabs or other organs tested. Virus could not be isolated later than 8 days after inoculation. Hemagglutination inhibiting and neutralizing antibodies were detected in sera at 6 days and 9 days after inoculation, respectively.