Fine structural analysis of membrane changes during reaggregation culture of chick optic tectum

Thin section and freeze-fracture electron microscopy have been used to characterize the changes in membrane morphology of reaggregating cultures of chick optic tectum. The cells are rounded and freely dispersed at 0 hr after dissociation. Between 2 and 6 hr the cells become closely apposed on all sides by other cells and form small aggregates. At this time punta adhaerentia junctions and focal densities are seen along the membranes of neighboring cells. Between 1 and 5 days in vitro (DIV) neurites containing growth cone regions are present. At 5 DIV the first synaptic contacts are observed. Between 7 and 14 DIV, the number of synaptic contacts increase and fewer growth cone regions are observed. As early as 7 DIV profiles are observed which strongly resemble both astrocytic and oligodendroglial cell somata and processes. Freeze-fracture analysis of aggregates at 0–4 hr reveals a sparse particle distribution on the P and E faces of apposed cells. By 1 DIV small clusters of loosely packed, large sized particles are seen on the P face of apposed cell membranes which may represent junctional contacts. Apparent coated vesicle fusion sites are common on the P face at 1–2 DIV. By 7 DIV, E face particle arrays are seen on cell bodies and neurites which correspond to specializations characteristic of excitatory synaptic junctions. By 8–10 DIV particle arrays are seen on the P face of post-synaptic membrane which may represent inhibitory synaptic contacts. Other types of particle specializations seen in freeze-fracture replicas include: specializations characteristic of gap junctions between cells and orthogonal assemblies of particles thought to be characteristic of astrocytes.     

Genetic and biochemical requirements for chemotaxis to L-proline in Escherichia coli.

Chemotaxis to L-proline was examined by the capillary assay, using a set of Escherichia coli strains bearing well-defined defects in the enzymes of proline transport and utilization. Aspartate taxis was measured as a constitutive, control activity whose receptor and transducer requirements are known. Proline chemotaxis showed a pattern of induction more analogous to that of proline dehydrogenase than of that of proline transport, but chemotaxis to proline was eliminated by mutations eliminating either or both of these activities. No response to proline was observed in the absence of a proline concentration gradient or when succinate was provided as an oxidizable carbon source. These data suggest that the chemotactic response to proline results from a direct impact of proline oxidation on the energy metabolism of the cell.     

Photooxidation products of 7,12-dimethylbenz[a]anthracene.

The principal products of the photooxidation of 7,12-dimethylbenz[a]-anthracene (DMBA) in aqueous solutions by photooxidation induced by laboratory lighting have been characterized by high performance liquid chromatograms (HPLC), ultraviolet and mass spectrograms and by comparisons with authentic samples. The products identified were the 7,12-epidioxy-7,12-dihydro-7-12-dimethyl-, 7,12-dione, 7-hydroxymethyl-12-methyl-, 12-hydroxymethyl-7-methyl-, 7-formyl-12-methyl-, 12-formyl-7-methyl-, and 12-hydroxy-12-methyl-7-one derivatives of benz[a]-anthracene. The HPLC profile of products is similar to that obtained from oxidation of DMBA by 'one-electron' reagents, singlet oxygen, or liver microsomal metabolism. The first points of attack are the 7- and 12- positions. The mechanism of photooxidation appears to be generation of singlet oxygen by photodynamic effect of DMBA. None of the products is photosensitizing, however.     

Needle aspiration of the breast with immediate reporting of material.

Needle aspiration with immediate cytological reporting has been practised in a breast clinic for one year. Patients benefit by receiving immediately the provisional diagnosis and, when indicated, appointments for metastatic surveys. Close co-operation between surgeon and cytologist has resulted in increased skill in aspiration, better preparation of samples, and greater accuracy in interpretation of reports. Since 5% of clinically benign lesions have proved malignant, even on immediate reporting, we would recommend cytological examination of all breast lumps.     

A rapid,enzymatic assay for the measurement of inorganic pyrophosphate in animal tissues

A simple, rapid enzymatic assay for the determination of inorganic pyrophosphate in tissue and plasma has been developed using the enzyme pyrophosphate-fructose-6-phosphate 1-phosphotransferase (EC 2.7.1.90) which was purified from extracts of Propionibacterium shermanii. The enzyme phosphorylates fructose-6-phosphate to produce fructose-1,6-bisphosphate using inorganic pyrophosphate as the phosphate donor. The utilization of inorganic pyrophosphate is measured by coupling the production of fructose-1,6-bisphosphate with the oxidation of NADH using fructose-bisphosphate aldolase (EC 4.1.2.13), triosephosphate isomerase (EC 5.3.1.1), and glycerol-3-phosphate dehydrogenase (NAD⁺)(EC 1.1.1.8). The assay is completed in less than 5 min and is not affected by any of the components of tissue or plasma extracts. The recovery of pyrophosphate added to frozen tissue powder was 97 ± 1% (n = 4). In this assay the change in absorbance is linearly related to the concentration of inorganic pyrophosphate over the cuvette concentration range of 0.1 μm to 0.1 mm.