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A mass of connective tissue, continuous with the neural lamella, develops on the dorsal side of the abdominal region of the nerve cord of Lepidoptera during the pupal stage. The mucosubstances of this tissue in the wax-moth,Galleria mellonella, have been characterized histochemically using various techniques involving Alcian Blue binding, periodic acid-Schiff and high iron diamine reactions, and enzyme digestions. The results indicate that this fibrous tissue contains chondroitin and dermatan sulphates and neutral glycoproteins. Thoracic ganglia of adult stick insects,Carausius morosus, were subjected to the same histochemical tests. The neural lamella possesses chondroitin, dermatan and keratan sulphates, while the glial lacunar system contains only hyaluronic acid.

Our objective was to determine whether subarachnoid haemorrhage modifies cerebral artery smooth muscle cell phenotype and the contractile protein α-actin measured 7 days after haemorrhage. We used a rabbit subarachnoid haemorrhage model and immunofluorescence labelling of α-smooth muscle actin, vimentin and desmin. The paired comparison between the haemorrhage and sham rabbits was performed using confocal laser-scanning microscopy. We found in the haemorrhage group significantly less intense α-actin immunostaining (p = 0.036) and more intense vimentin immunostaining (p = 0.043) but no significant change in the intensity of desmin staining. Our results indicate an absolute decrease after subarachnoid haemorrhage in the amount of functional α-actin and in the light of the literature may suggest a certain degree of dedifferentiation of smooth muscle cells in the cerebral artery wall.

An improved new method for the simultaneous visualization of mRNA and encoded protein in LR White resin-embedded specimens is described. This pre-embedding electron microscopical in situ hybridization (procedure) localized rat growth hormone mRNA specifically as high electron-density products on the polysomes of the rough endoplasmic reticulum. A subsequent post-embedding immunoreaction, using protein A colloidal gold particles, identified growth hormone as gold particles both in the cisternae of the rough endoplasmic reticulum and on the secretory granules. In our previous report, we used Epon resin for tissue embedment, which required an etching process using hydrogen peroxide or sodium periodate for immunoreactivity retrieval. In general, osmification and embedment in Epon resin are reported to decrease the immunoreactivity of the targeted protein, and the etching process using hydrogen peroxide or sodium periodate results in deosmification and shades off the signals of mRNA. To resolve these problems, we have recently used LR White resin for tissue embedment. In LR White resin-embedded tissues, retrieval of immunoreactivity using hydrogen peroxide or sodium periodate is not required, and, therefore, the gradation of the signals of mRNA can be avoided. © 1998 Chapman & Hall

A cholinesterase localization method and a monoamine histofluorescence technique were used to locate nerve fibres in regenerating rat submandibular gland autografts. Experimental rats had a portion of one submandibular gland excised and cut into small fragments which were autografted immediately into the middle one-thrid of the tongue. Control rats had a portion of one submandibular gland removed and discarded, and their tongues were sham-operated. Seven to ten weeks later, the rats were killed and the tongues were removed, frozen and sectioned in a cryostat. A light microscopical study of the tongue sections subjected to the cholinesterase technique showed that the submandibular gland autografts contained many nerve fibres that exhibited cholinesterase activity. These cholinesterase-positive nerve fibres were distributed throughout the autografts. The fibres were associated with the numerous duct-like structures and the less numerous acini. In addition, ultraviolet illumination of tongue sections after treatment with a glyoxylic acid mixture revealed histofluorescent monoaminergic nerves within the autografts. These fibres were less prominent than the cholinesterase-positive fibres and appeared to run primarily along blood vessels within the autografts. The results suggest that autonomic nerves are present within regenerating submandibular gland autografts.

Histamine stimulates acid secretion by the parietal cell and this secretion is inhibited by the histamine H2-receptor antagonists. Whole body autoradiography showed that radioactivity from14C-histamine was localized in the artery walls of the stomach and in the muscularis mucosae, but that the level in the fundic mucosa was the same as the blood. When the H2-receptor antagonists burimamide, metiamide and cimetidine were labelled with35S,14C or3H and dosed to rats, whole body autoradiography showed that the stomach was predominantly labelled in the glandular mucosa from 5 to 120 min after administration. Microautoradiography in the rat and dog after intravenous injection of [3H] metiamide or [3H] cimetidine demonstrated an uptake of tritium in the parietal cell cytoplasm that was 3- to 4-times greater than that found in adjacent peptic cells or areas of muscularis mucosa. The preferential labelling persisted at a low level up to 6 h after injection in the rat. The localization of radioactivity from the H2-antagonists in the parietal cell cytoplasm correlates well with their pharmacological activity in preventing acid secretion from this cell.

The non-enzyme histochemistry of the closely related parathyroid and ultimobranchial glands of the domestic fowl has been studied. The parathyroid cells and the ultimobranchial C cells gave only weakly positive reactions in tests for specific amino-acid residues. The rather stronger reactions found in the C cells were probably related to their content of polypeptide secretory granules. Carbohydrate and lipid stains also gave only negative or weakly positive reactions although the parathyroid cells did contain some free lipid. The lining cells of the ultimobranchial vesicles may be either actively secreting or inactive and cystic. Active cells contain large numbers of granules that are composed of large amounts of sulphated acid mucopolysaccharides, moderate amounts of mucoproteins and small amounts of neutral mucopolysaccharides and glycogen. Both the granular and colloidal secretions of these cells appeared to be identical in composition to the intracellular granules. The vesicular lining cells and their secretions showed strong but variable reactions for sulphydryl, disulphide and amino groups. The intermediate cell strands, which connect the ultimobranchial parathyroid foci with the vesicles, showed a range of reactions intermediate between those exhibited by parathyroid and vesicular cells. The results are discussed in relation to the structure and function of the various cell types.

Cell fractionation, SDS-PAGE, quantitative Western blot, confocal immunolocalization and immunogold labelling were performed to find an interpretation of the physiological response of the unicellular eukaryote Paramecium to β-adrenergic ligands. The 69 kDa polypeptide separated by SDS-PAGE in S2 and P2 Paramecium subcellular fractions cross-reacted with antibody directed against human β2-adrenergic receptor. This was detected by Western blotting followed by chemiluminescent detection. Quantitative image analysis showed that β-selective adrenergic agonist (–)-isoproterenol – previously shown to enhance phagocytic activity – evoked redistribution of the adrenergic receptor analogue from membraneous (P2) to cytosolic (S2) fraction. The relative increase in immunoreactive band intensity in S2 reached 80% and was paralleled by a 59% decrease in P2 fraction. Confocal immunofluorescence revealed β2-adrenergic receptor sites on the cell surface and at the ridge of the cytopharynx – where nascent phagosomes are formed. This localization was confirmed by immunoelectron microscopy. These results indicate that the 69 kDa Paramecium polypeptide immunorelated to vertebrate β2-adrenergic receptor appeared in this evolutionary ancient cell as a nutrient receptor.

Localisation patterns of the transcription factor Stat5b in the udders from pregnant, lactating and involuting ewes were compared with the expression patterns of two major milk protein genes α-lactalbumin and αS1 casein. Stat5b was detected in the cytoplasm and nuclei of epithelial cells at all stages of mammary gland development. A consistent positive relationship between the nuclear localisation of Stat5b in lactating mammary alveolar epithelial cells, and the presence of milk protein gene mRNA was apparent during lactation and early involution. Conversely, there was little evidence of nuclear localisation of Stat5b in non-lactating mammary alveolar epithelial cells during lactation and early involution. This supports the observation that during lactation, Stat5b may play a role in milk protein gene expression. However, during pregnancy and later involution, while Stat5b was observed to be present in mammary epithelial cell nuclei and cytoplasm, no relationship between this and the presence of milk protein gene mRNA was apparent. This suggests that during late pregnancy and in later involution, Stat5b may be involved in processes other than initiation of milk protein gene transcription.