Detection of DNA by tritiated actinomycin D on ultrathin frozen sections

Ultrathin frozen sections of fresh liver tissue were floated on actinomycin D-³H. Quantitative high resolution radioautography was performed to determine the value of the method for detection of DNA by electron microscopy. A complete series of control experiments involving various treatments of frozen sections with enzymes (pronase, DNase) and 0.1 N HCl were also carried out to determine the specificity of the labeling. The results indicate the value of the method for detection of DNA directly on ultrathin frozen sections. Short treatments with pronase followed by DNase reduce the labeling to zero, whereas removal of chromosomal proteins with HCl increases the amount of radioactivity in the nucleus considerably. The results are discussed in view of the future applications opened by ultracryotomy, since radioautographic detection of various macromolecules and cellular components by labeled compound with specific affinities will now be possible.     

No Evidence of BRCA1/2 genomic rearrangements in high-risk French-Canadian breast/ovarian cancer families

The discovery of deleterious mutations in the breast and ovarian cancer susceptibility genes, BRCA1 and BRCA2, has facilitated the identification of individuals at particularly high risk of these diseases. There is a wide variation between populations in the prevalence and related risks of various types of BRCA1/2 mutations, so estimates cannot be extrapolated to Canadians, especially not founder populations such as French- Canadians. Polymerase chain reaction (PCR)-based methods were used to detect the majority of these mutations. These approaches usually failed to detect large DNA rearrangements, which have been claimed to be involved in other populations in 5% to up to 36% of BRCA1-positive families. There is very little information about the contribution of this type of mutation in BRCA2-positive families. To investigate if our available mutation spectrum of BRCA1 and BRCA2 in high-risk French-Canadian breast/ovarian cancer families has been biased by PCR-based direct sequencing methods, we first used Southern blot analysis to test DNA samples from 61 affected/obligate carrier individuals from 58 families in which no BRCA1/2 deleterious mutation was found. Finally, 154 individuals from 135 BRCA1/2 nonconclusive families, including all those tested previously by Southern blot analysis, were tested with the new multiplex ligation probe amplification (MLPA) technique. These approaches failed to detect any rearrangement. Moreover, if the frequency of MLPA-detectable rearrangements in our cohort of 135 BRCA1/2 nonconclusive families was 2.2% or higher, we would have had a 95% or greater chance of observing at least one such rearrangement. As no rearrangements were identified, such large rearrangements must be quite rare in our population.     

Food restriction affects energy metabolism in rat liver mitochondria

To examine the effect of 50% food restriction over a period of 3 days on mitochondrial energy metabolism, liver mitochondria were isolated from ad libitum and food-restricted rats. Mitochondrial enzyme activities and oxygen consumption were assessed spectrophotometrically and polarographically. With regard to body weight loss (-5%), food restriction decreased the liver to body mass ratio by 7%. Moreover, in food-restricted rats, liver mitochondria displayed diminished state 3 (-30%), state 4-oligomycin (-26%) and uncoupled state (-24%) respiration rates in the presence of succinate. Furthermore, "top-down" elasticity showed that these decreases were due to an inactivation of reactions involved in substrate oxidation. Therefore, it appears that rats not only adapt to food restriction through simple passive mechanisms, such as liver mass loss, but also through decreased mitochondrial energetic metabolism.     

Incidence of cystic fibrosis in Saguenay-Lac-St.-Jean (Quebec, Canada).

The incidence of cystic fibrosis (CF) in Saguenay-Lac-St.-Jean, a geographically isolated region of Quebec, was estimated to be 1 in 902 during the period 1975-1988. The carrier rate was calculated to be 1 in 15 inhabitants. The high incidence of CF in Saguenay-Lac-St.-Jean is probably the result of a founder effect and genetic drift for one or more mutations. Historical, demographic, and social factors also may have contributed to the high incidence.     

Factors involved in the post-copulatory neural inhibition of pheromone production in Choristoneura fumiferana and C. rosaceana females

Normal mating lasts approximately 3 h in Choristoneura fumiferana and C. rosaceana. Data generated from interrupted matings showed that the act of mating did not suppress pheromone production (pheromonostasis) in either species although, in C. rosaceana, pheromone titre declined slightly the night following mating. In both species the migration of sperm to the spermatheca (SP) occurred several hours after mating, and coincided with a significant and permanent depression in pheromone titre, as well as egg fertilisation and oviposition. However, disrupting matings within 2 h of the onset resulted in oviposition patterns similar to virgins in both species, with mostly infertile eggs being laid by C. fumiferana females while oviposition was totally inhibited in C. rosaceana. The transection of the ventral nerve cord (VNC) 1 h post-mating did not result in the depression of pheromone titres the following night in either species but if the VNC was transected 3 h post-mating, pheromonostasis was observed. While 25% of C. fumiferana females had sperm in their SP 2 h after mating, it took at least 4 h in C. rosaceana. This suggests that while the physical presence of sperm in the SP may play some role in the termination of pheromone production in C. fumiferana, other factors must trigger the neural signal that elicits pheromonostasis in both species. A better understanding of the temporal dynamics of both apyrene and eupyrene sperm within the different parts of the female reproductive system might clarify these interspecific differences.