Temporal Expression of Mouse Glial Fibrillary Acidic Protein mRNA Studied by a Rapid In Situ Hybridization Procedure

A rapid and sensitive in situ hybridization technique is described for the detection of mRNA sequences in 6-8-micron cryostat sections. The method incorporates the use of alpha-thio-35S-labelled nucleoside triphosphates for the generation of high-specific-activity DNA probes and a high-stringency washing procedure that virtually eliminates background without unduly compromising histological integrity. Whereas signal resolution is less than that observed using 3H probes, 35S-labelled probes are well-suited for experiments where resolution at the cellular level is required. The method has been applied to a study of the developmental regulation of glial fibrillary acidic protein (GFAP) mRNA expression in developing mouse brain. GFAP-specific sequences are first detectable after the second postnatal day, and thereafter rise to a level that is maintained throughout development and into adulthood. The distribution of GFAP-encoding sequences broadly reflects the known distribution of astrocytes, but the levels of mRNA within these cells vary by a surprisingly large amount depending on their location. For example, in adult animals, the astrocytes of the glial limitans contain an abundance of GFAP-specific mRNA that is higher than corresponding levels in astrocytes in the cerebellar white matter, whereas these cells in turn contain considerably more GFAP-specific mRNA than astrocytes in the gray matter of the cerebrum. Unexpectedly, parallel RNA blot transfer experiments show the existence of some GFAP-encoding mRNA size heterogeneity that is restricted to the first postnatal week.     

Suppression of DNA synthesis in mitogen stimulated leukocytes with partial trypsin digests of Staphylococcus aureus protein A

Staphylococcus aureus protein A (SPA) acts as a mitogen stimulating DNA synthesis and is not antagonistic to the mitogenic activity of other mitogens. The mitogenicity of SPA is due to the protein's multivalent Fc antibody receptors. Partial trypsin digestion of SPA yields monovalent Fc antibody receptors which are not mitogenic. Using a thymidine incorporation assay, results indicated that tryptic digests of SPA added to mouse splenic leukocytes 12 hours after stimulation with either concanavalin A, lipopolysaccaride or SPA suppressed the DNA synthesis normally elicited by these mitogens. SPA was digested with 1% trypsin at pH 8 for varying periods of time. As the digestion time increased from 0 to 60 minutes the mitogenicity of the SPA-trypsin digests decreased indicating cleavage from multivalent to monovalent SPA Fc antibody receptors. The decrease in mitogenicity of the SPA digests was directly associated with increased ability to inhibit DNA synthesis in mitogen stimulated leukocytes. A proposed mechanism for the inhibition of DNA synthesis suggests that SPA-monovalent Fc antibody receptors mimic cellular Fc receptor function thereby influencing cellular gene expression.     

Turkey feeding regimes with different ages of introduction of a choice of foods

An experiment was carried out to investigate five feeding regimes for the turkey which involve introducing a choice of wheat grain and a higher-protein food at 1, 14, 28, 56 and 77 days of age. Broken wheat grains were used from days 1 to 14, and whole grains thereafter. Protein intakes per bird, calculated from standard tables of feed composition, were 39.1, 43.0, 41.8, 35.5 and 36.0 g/day, respectively, from 14 to 98 days. Further groups of birds on a complete diet regime consumed a significantly lower 33.3 g/day. Only two comparisons were significantly different for body weight at 98 days: birds which received the choice at 14 days had a significantly higher weight (5.7 kg/bird) than those which received the choice at 56 days (5.5 kg) and than those on the complete diet regime (5.5 kg). $\text{P}$resentation of the wheat and higher-protein food during the period of protein over-consumption, according to a fixed ratio schedule, might reduce protein intake.