Labelling Criteria Research Articles

Isolation and characterization of a Chinese hamster ovary cell mutant which accumulates UDP glucuronic acid and requires uridine for growth.

The isolation and characterization of a new pyrimidine requiring mutant of Chinese hamster ovary cells (CHO-Kl, pro-) is described. This mutant (P192) shows an absolute requirement for uridine for growth but is fully capable of pyrimidine nucleotide synthesis. P192 cells accumulate exceedingly large amounts of a compound identified by a variety of chromatographic, chemical, and labelling criteria as UDP glucuronic acid (UDPglcUA). This compound comprises approximately 60% of the acid-soluble nucleotides of P192 cells. Possible genetic mechanisms leading to the phenotype of P192 are examined, and the role of UDPglcUA in cellular metabolism is discussed.

Axotomy causes loss of muscarinic receptors and loss of synaptic contacts in the hypoglossal nucleus.

MUSCARINIC receptors in the brain and ileum can be studied by means of the binding of radiolabelled cholinergic antagonists1,2. The basic criteria for receptor-specific labelling are that there should be a saturable component of binding and that the binding of the radiolabelled ligand should be inhibited by pharmacologically effective concentrations of drugs which are known to act at muscarinic receptors and not by drugs that act at different receptors. In addition, the saturable binding component should be restricted to tissues known from pharmacological experiments to contain muscarinic receptors. These criteria are satisfied by tritiated propylbenzilylcholine mustard (3H-PrBCM)3, a potent irreversible muscarinic antagonist the binding of which to rat brain sections can be demonstrated autoradiographically. The evidence which suggests that the bulk of tissue radioactivity is associated with muscarinic receptors is that in autoradiographs the regional variation of autoradiographic grain density parallels the intensity of specific 3H-PrBCM binding observed by in vitro assay on brain homogenates from selected regions, and that in the homogenates 65–85% of the bound radioactivity is blocked by pretreatment with the classical muscarinic antagonist, atropine (10−6M). In the forebrain the general features of the distribution of 3H-PrBCM binding to thin sections correspond closely with the findings of Kuhar and Yamamura4,5 who used the non-covalently bound antagonist 3H-3-quinuclidinylbenzilate. We report here, however, that in the brain stem the hypoglossal nuclei are heavily labelled with 3H-PrBCM. In light microscopic autoradiographs (Fig. 1a, left side) the grains are distributed throughout the neuropil of the hypoglossal nucleus. The cytoplasm and nuclei of the large hypoglossal neuronal somata (Fig. 1b) are unlabelled. The density of the silver grains falls abruptly at the borders of the hypoglossal neuropil. This clear demarcation of the labelled hypoglossal neuropil is abolished by atropine pretreatment (Fig. 1c).