Brain science: from the very small to the very large

We still lack a clear understanding of how brain imaging signals relate to neuronal activity. Recent work shows that the simultaneous activity of neuronal ensembles strongly correlates with local field potentials and imaging measurements.     

Response of mouse small intestine to fractionated doses of pions

The effect of single or fractionated doses of stopping pions or 200 kV X-rays on the mouse jejunal crypt cells was used to determine in vivo RBE values. For single fraction, the pion/X-ray RBE was 1.27 and it increased to about 1.31 when two fractions were applied at 3 or 24 h interval. When four fractions were given at 3 h intervals, the RBE was 1.46. This is because the fraction of "dose repaired" was always higher for X-rays than for pions and this difference was enhanced when more dose fractions were used. The data presented is, in general, consistent with the biological effects of pions reported for other in vivo end points.     

Aspects of the adaptive response to very low doses of radiation and other agents

When human lymphocytes and other cells are pre-exposed to very low doses of ionizing radiation and subsequently exposed to a high dose, less genetic damage, i.e., fewer chromosome aberrations, is found than is observed in cells that had not been pre-exposed. This has been termed the adaptive response and has been attributed to the induction of a repair mechanism by the low dose exposure. Several experiments have now been carried out on this adaptive response to better characterize the phenomenon. (A) Experiments with differential display of mRNAs indicate that human lymphocytes exposed to 2 cGy of X-rays have somewhat different mRNAs expressed than do unexposed cells. This is providing access to DNA that might be involved in adaptation. (B) Other experiments with embryonic cells from transgenic mice that are deficient in superoxide dismutase (SOD) have shown that the adaptive response is unrelated to the amount of SOD in the cells, and thus is independent of superoxide radicals. (C) Experiments in which very low doses of various restriction enzymes were electroporated into human lymphocytes have shown that low levels of double-strand DNA breaks alone are able to induce the adaptive response. (D) Experiments in which human male lymphocytes (XY chromosome constitution) and human female lymphocytes (XX chromosome constitution) were cocultivated have shown that adaptation is not caused by a change in the rate of cell progression to mitosis after a challenge dose, and is a further indication that cell stage sensitivity is not a factor in the adaptive response.     

DNA damage and repair in embryonic mouse limbs exposed to teratogenic doses of methylnitrosourea

The alkaline elution assay was used to monitor DNA single-strand breaks in embryonic tissue following exposure to the DNA-damaging teratogen N-methyl-N-nitrosourea (MNU, CAS No. 694-93-5). An animal model was developed in which nearly every fetus exposed to the highest dose of MNU had malformations of the hindlimbs while the fetuses exposed to the lowest dose of MNU had none. Hindlimbs pooled within litters were analyzed for DNA single-strand breaks by alkaline elution conducted at rapid (0.35 ml/min) and slow (0.35 ml/min) speeds. Breaks in the DNA of hindlimbs exposed to teratogenic doses of MNU were readily detected by alkaline elution only if slower speeds were used in the assay. Using the more sensitive procedure, DNA breakage was monitored over a 24-h period. DNA breakage peaked in the MNU-exposed hindlimbs in a dose-dependent manner 4 h after injection. While the elution profiles of hindlimbs exposed to the lower doses of MNU returned to control levels 8 h after injection, single-strand breaks persisted in the hindlimbs exposed to the highest dose of MNU for at least 20 h. These latter data suggest that the highly teratogenic dose of MNU induced DNA damage that was more slowly repaired than that produced at lower doses, possibly by saturation of DNA repair systems. Although some necrosis did occur in hindlimbs exposed at teratogenic dose levels, it was not severe and it did not appear to influence the alkaline elution results. These experiments show that alkaline elution is a sensitive assay for the detection of DNA damage in embryonic tissues.     

Renal damage in the mouse: the effect of d(4)-Be neutrons

A further study on the response of the mouse kidney to d(4)-Be neutrons (EN = 2.3 MeV) is described. The results confirm and augment the work published previously by Stewart et al. [Br. J. Radiol. 57, 1009-1021 (1984)]; the present paper includes the data from a top-up design of experiment which extends the measurements of neutron RBE (relative to 240 kVp X rays) down to X-ray doses of 0.75 Gy per fraction. The mean RBE for these neutrons increases from 5.8 to 7.3 as X-ray dose per fraction decreases from 3.0 to 1.5 Gy in the kidney. This agrees with the predictions from the linear quadratic (LQ) model, based on the renal response to X-ray doses above 4 Gy per fraction. The mean RBE estimate from a single dose group at 0.75 Gy per fraction of X rays is, however, 3.9. This is below the LQ prediction and may indicate increasing X-ray sensitivity at low doses. Data from this study and from those published previously have been used to determine more accurately the shape of the underlying response to d(4)-Be neutrons; an alpha/beta ratio of 20.5 +/- 3.7 Gy was found. The best value of alpha/beta for X rays determined from these experiments was 3.04 +/- 0.35 Gy, in agreement with previous values.     

Ventilatory acclimatization in response to very small changes in PO2 in humans.

Ventilatory acclimatization to hypoxia (VAH) consists of a progressive increase in ventilation and decrease in end-tidal Pco(2) (Pet(CO(2))). Underlying VAH, there are also increases in the acute ventilatory sensitivities to hypoxia and hypercapnia. To investigate whether these changes could be induced with very mild alterations in end-tidal Po(2) (Pet(O(2))), two 5-day exposures were compared: 1) mild hypoxia, with Pet(O(2)) held at 10 Torr below the subject's normal value; and 2) mild hyperoxia, with Pet(O(2)) held at 10 Torr above the subject's normal value. During both exposures, Pet(CO(2)) was uncontrolled. For each exposure, the entire protocol required measurements on 13 consecutive mornings: 3 mornings before the hypoxic or hyperoxic exposure, 5 mornings during the exposure, and 5 mornings postexposure. After the subjects breathed room air for at least 30 min, measurements were made of Pet(CO(2)), Pet(O(2)), and the acute ventilatory sensitivities to hypoxia and hypercapnia. Ten subjects completed both protocols. There was a significant increase in the acute ventilatory sensitivity to hypoxia (Gp) after exposure to mild hypoxia, and a significant decrease in Gp after exposure to mild hyperoxia (P < 0.05, repeated-measures ANOVA). No other variables were affected by mild hypoxia or hyperoxia. The results, when combined with those from other studies, suggest that Gp varies linearly with Pet(O(2)), with a sensitivity of 3.5%/Torr (SE 1.0). This sensitivity is sufficient to suggest that Gp is continuously varying in response to normal physiological fluctuations in Pet(O(2)). We conclude that at least some of the mechanisms underlying VAH may have a physiological role at sea level.     

DNA damage response during mouse oocyte maturation

Because low levels of DNA double strand breaks (DSBs) appear not to activate the ATM-mediated prophase I checkpoint in full-grown oocytes, there may exist mechanisms to protect chromosome integrity during meiotic maturation. Using live imaging we demonstrate that low levels of DSBs induced by the radiomimetic drug Neocarzinostatin (NCS) increase the incidence of chromosome fragments and lagging chromosomes but do not lead to APC/C activation and anaphase onset delay. The number of DSBs, represented by γH2AX foci, significantly decreases between prophase I and metaphase II in both control and NCS-treated oocytes. Transient treatment with NCS increases >2-fold the number of DSBs in prophase I oocytes, but less than 30% of these oocytes enter anaphase with segregation errors. MRE11, but not ATM, is essential to detect DSBs in prophase I and is involved in H2AX phosphorylation during metaphase I. Inhibiting MRE11 by mirin during meiotic maturation results in anaphase bridges and also increases the number of γH2AX foci in metaphase II. Compromised DNA integrity in mirin-treated oocytes indicates a role for MRE11 in chromosome integrity during meiotic maturation.     

Adaptive response to chromosome damage in cultured human lymphocytes primed with low doses of X-rays

Human lymphocytes exposed to 0.02 Gy of X-rays in the G1 but not the G0 phase became less susceptible to the induction of chromosome aberrations of the chromosome type by subsequent exposure to 3 Gy of X-rays. The induction of chromatid-type aberrations was not affected by the pretreatment with the priming dose. The expression of this adaptive-type response was transitory, being maximum at 5 h, and disappeared at 9 h after the initial low-dose exposure. Cell-cycle analysis excluded the possibility of a spurious consequence of differential cell-cycle progression.     

Mutation of mouse p53 Ser23 and the response to DNA damage

Recent studies have suggested that phosphorylation of human p53 at Ser20 is important for stabilizing p53 in response to DNA damage through disruption of the interaction between MDM2 and p53. To examine the requirement for this DNA damage-induced phosphorylation event in a more physiological setting, we introduced a missense mutation into the endogenous p53 gene of mouse embryonic stem (ES) cells that changes serine 23 (S23), the murine equivalent of human serine 20, to alanine (A). Murine embryonic fibroblasts harboring the p53(S23A) mutation accumulate p53 as well as p21 and Mdm2 proteins to normal levels after DNA damage. Furthermore, ES cells and thymocytes harboring the p53(S23A) mutation also accumulate p53 protein to wild-type levels and undergo p53-dependent apoptosis similarly to wild-type cells after DNA damage. Therefore, phosphorylation of murine p53 at Ser23 is not required for p53 responses to DNA damage induced by UV and ionizing radiation treatment.     

Hepatotoxic doses of dioxin do not damage mouse bone marrow chromosomes

We report on a study of the cytogenetic and hepatotoxic effects of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). Mice of the C57B1/6J (with high-affinity TCDD receptor) or DBA/2J (with low-affinity TCDD receptor) strains were given single intraperitoneal injections of 50, 100 or 150 micrograms of TCDD/kg body weight. At various times (8-48 h) after injection, we examined bone marrow cells for cytogenetic effects by performing structural aberration, sister-chromatid exchange, and micronucleus tests. 1 month after exposure, liver sections were studied for hepatotoxic effects. We found no evidence of chromosome damage by TCDD given in doses that cause liver damage in both strains of mice.     

Renal response to secretin.

The effect of Jorpes secretin on the urinary volume, pH, and excretion of sodium, potassium, chloride, bicarbonate, titratable acidity, ammonia and phosphate was studied in five healthy male volunteers with and without simultaneous aspiration of duodenal fluids. A three- to fourfold increase in urinary volume and sodium excretion occurred within the first 30 min after secretin injection and this was accompanied by a significant rise in urinary pH in each instance. Urinary bicarbonate excretion increased from 55 plus or minus 13 to 395 plus or minus 33 mueq/30 min after secretin injection. Aspiration of alkaline duodenal contents was accompanied by an even greater postsecretin increase in urinary bicarbonate excretion. No significant changes in arterial pH or blood gases were detected throughout the study. These observations are compatible with a direct effect of secretin upon the renal tubular reabsorption of water, bicarbonate, and other ions, and could account for the transient alterations in urinary pH occurring in response to a meal.     

Inhibition of the adaptive response of human lymphocytes to very low doses of ionizing radiation by the protein synthesis inhibitor cycloheximide

When human lymphocytes are preirradiated with 1 cGy of X-rays, the cells become less sensitive to subsequent exposures to high doses of about 150 cGy in that approximately one-half as many chromatid aberrations are induced as expected. This adaptation has been attributed to the induction of repair enzymes (proteins) some 4-6 h after the initial low-dose exposure. Experiments have now been carried out showing that application of the protein synthesis inhibitor cycloheximide at this time, but not earlier, prevents the adaptive response.