Application of modern methods of organic chemistry and recombinant DNA technologies has provided new insights in the field of DNA radiation damage and its repair. An overview of the chemical nature of the lesions inflicted on DNA by ionizing radiation is presented. The structures of 29 different DNA modified base or sugar residues are shown in comprehensive formation schemes. A fraction of radiation-induced modified bases is spontaneously released from the DNA chain during irradiation. Another part remains attached to the DNA chain backbone and for its characterization mild formic acid or enzymatic hydrolysis have been used. Starting from the chemical formulae of the altered base residues, the specific repair enzymes and their modes of action are discussed. Various glycosylases and endonucleases have been purified to homogeneity, and in some cases the gene which encodes the protein cloned. Using methods derived from Maxam and Gilbert sequencing procedures and DNA fragment 32P-labelled at one end, it has been shown that the alkali-labile sites in DNA induced by radiation are strongly dependent on the DNA base sequence. Enzymatic methods have been used to analyse the DNA base defects produced by gamma-irradiation of cells under in vivo conditions. Structures of modified bases were the same as those observed when DNA was irradiated in aqueous solution.

In situ hybridization experiments were performed with brain sections from normal, control and haloperidol-treated rats to identify and map the cells expressing the D2 dopamine receptor gene. D2 receptor mRNA was detected with radioactive or biotinylated oligonucleotide probes. D2 receptor mRNA was present in glandular cells of the pituitary intermediate lobe and in neurons of the substantia nigra, ventral tegmental area, and forebrain, especially in caudate putamen, nucleus accumbens, olfactory tubercle, and piriform cortex. Hybridization with D2 and preproenkephalin A probes in adjacent sections, as well as combined hybridization with the two probes in the same sections, demonstrated that all detectable enkephalin neurons in the striatum contained the D2 receptor mRNA. Large neurons in caudate putamen, which were unlabeled with the preproenkephalin A probe and which may have been cholinergic, also expressed the D2 receptor gene. Haloperidol treatment (14 or 21 days) provoked an increase in mRNA content for D2 receptor and preproenkephalin A in the striatum. This suggests that the increase in D2 receptor number observed after haloperidol treatment is due to increased activity of the D2 gene. These results indicate that in the striatum, the enkephalin neurons are direct targets for dopamine liberated from mesostriatal neurons.Biochemical and pharmacological investigations have demonstrated the existence of two dopamine receptor subtypes, D1 and D2, which differentiate the signal transduction mediated by dopamine as either an activation or an inhibition of adenylate cyclase (1-4). Rat D2 receptor cDNA has been cloned and sequenced, and it appears that the D2 receptor belongs to a family of receptors that are coupled to guanine nucleotide-binding proteins (5). Its mRNA is abundantly represented in rat brain, especially in the mesencephalon and the basal ganglia (5). These areas contain, respectively, the cell bodies and terminals of the dopaminergic mesostriatal system (6). Striatal neuron activities are under the influence of the dopamine neurons of the substantia nigra and ventral tegmental area (7,8). Binding experiments with radiolabeled ligands have demonstrated the presence of striatal dopamine receptors (9-15). Drugs that interact with dopamine at receptor sites significantly affect neuronal activities, including neuropeptide gene expression (16)(17)(18)(19), in the striatum.While the existence of dopamine receptors on striatal neurons and on dopamine terminals in the striatum is commonly accepted (9-15), there is no anatomical information about the characteristics of the cells expressing the dopamine receptor gene in the striatum. To better understand how D2 receptor gene expression contributes to nigrostriatal interactions, we have used in situ hybridization to study the characteristics of cells containing D2 receptor mRNA in adult rat forebrain, under normal conditions and after blockade of dopamine transmission with a dopamine receptor antagonist, haloperidol. We report here that most cells conta...

Phenoxyacetyl (pac) and methoxyacetyl (mac) for adenine and guanine, isobutyryl for cytosine, were successfully applied as amino protecting groups both in phosphotriester and phosphoramidite approaches. As shown by N.M.R. and H.P.L.C. analysis, they are completely deblocked in less than four hours in 29% ammonia at room temperature allowing the preparation of modified DNA containing alkali labile bases such as saturated pyrimidines. The stability of N6-phenoxyacetyl-deoxyadenosine versus depurination in acidic conditions used in the detritylation step was favorably compared with that of the classic N6-benzoyl protected adenine.

A new methodology for the preparation of addressed DNA matrices is described. The process includes an electrochemically directed copolymerization of pyrrole and oligonucleotides bearing on their 5' end a pyrrole moiety introduced by phosphoramidite chemistry. The electro-controlled synthesis of the copolymer (poly-pyrrole) gives, in one step, a solid conducting film deposited on the surface of an electrode. The resulting polymer consists of pyrrole chains bearing covalently linked oligonucleotide. The polymer growth is limited to the electrode surface, so that it is possible to prepare a DNA matrix on a multiple electrode device by successive copolymerizations. A support bearing four oligonucleotides was used to detect three ras mutations on a synthetic DNA fragment.

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