Biomedical subjects

I Wanner

Publications and source records attributed to I Wanner.

9 recordsLinked to original sources

PubMed2000

Changing subcellular distribution and activity-dependent utilization of a dendritically localized mRNA in developing Purkinje cells.

In cerebellar Purkinje neurons, the degree of dendritic segregation of the Purkinje cell-specific mRNA L7/pcp-2 is correlated with their development and synaptic investment. This developmental pattern is also observed in Purkinje cells in primary dissociated culture. Short-term (12-48 h) stimulation of cultured Purkinje cells by potassium-induced depolarization or blockade of their inhibitory GABAergic input results in an increased incidence of Purkinje cells with L7/pcp-2 mRNA-positive dendrites and increased levels of L7 protein expression, the latter by a posttranscriptional mechanism. None of these treatments affected the localization of the mRNA encoding calbindin D28k nor the level of this protein in Purkinje neurons. Protracted exposure to depolarizing levels of potassium or elimination of GABAergic transmission resulted in conspicuous changes of Purkinje cell dendritic morphology. These data suggest a scenario in which activity-driven translation of subcellularly segregated mRNAs may contribute to the developmental and functional plasticity of nerve cells.

Animals↗

PubMed1998

Mapping of D1 dopamine receptor mRNA by non-radioactive in situ hybridization.

In order to improve the identification and characterization of dopaminoceptive neurons, the rat brain was mapped for D1 dopamine receptor mRNA by non-radioactive in situ hybridization (ISH) with a 45mer digoxigenin-labeled oligonucleotide probe. The specificity of the results was controlled with the help of a 396-bp D1 receptor riboprobe. Labeled hybrids were visualized with an alkaline phosphatase-coupled anti-digoxigenin antibody. The high resolution obtained permitted individual labeled cells to be identified and to distinction between cell bodies and processes. D1 mRNA was largely confined to neurons. With the exception of ependymal cells, glial cells were not distinctly labeled. Subcellularly, D1 mRNA was localized to perikarya but not to dendrites or axons. D1 receptor-expressing neurons were present in all of the known terminal fields of mesencephalic or diencephalic dopaminergic neurons. However, D1 message was also detected in brain areas which are not known to contain D1 ligand binding sites or in which the presence or the cellular source of this receptor subtype had previously not been unequivocally established, such as the hippocampus or cerebellar cortex. Moreover, labeled neurons were present in regions not known to receive dopaminergic projections, such as the thalamic and some brainstem nuclei. We conclude that this ISH technique provides a considerable gain in sensitivity and resolution with regard to neurotransmitter receptor mapping.

Animals↗

PubMed1996

Molecular genetics of mammalian olfaction.

Olfaction plays a crucial role in the survival of most animal species; it is remarkable in its ability to recognize and discriminate numerous airborne molecules, yet is one of the least understood senses. The advent of molecular genetic approaches has greatly contributed to disclosing some of the mysteries in olfaction. The identification of olfactory-specific proteins, the discovery of the large receptor gene family, and the first insight into the mechanisms governing chemosensory gene expression hold great promise for an eventually detailed understanding of a sensory system that was previously considered as hardly accessible for research at the molecular level.

Animals↗

PubMed1995

Receptor expression in olfactory neurons during rat development: in situ hybridization studies.

In situ hybridization approaches have been employed to explore the onset and time course of odorant receptor expression during the prenatal development of rats. The critical phase for the maturation of the olfactory system from embryonic day (E) 12 to E18 was analysed. The onset of expression of four receptor genes (OR5, OR14, OR37 and OR124) was found between day E12 and E14. In the early phase of development (E14) a distinct receptor subtype was expressed only in a few hundred neurons; the number increased about two-' to three-fold within a 2 day interval. From the very beginning, spatial segregation of receptor subtypes in distinct expression zones was observed.

Animals↗

PubMed1994

Probing olfactory receptors with sequence-specific antibodies.

Molecular cloning has revealed the structure of several putative odorant receptors. Chemically synthesized peptides, that correspond to a predicted extracellular domain of the encoded proteins, were employed to generate receptor-specific antibodies. Immunohistological approaches as well as Western-blot analysis confirmed the specificity of the antipeptide sera. Furthermore, deglycosylation experiments explained the observed discrepancy between the molecular mass of odorant receptors, as determined by SDS/PAGE and Western-blot analysis of ciliary proteins (M(r) 50,000), and the predicted protein size based on the deduced primary structure from cloned receptor genes (M(r) 30,000-35,000). Receptor proteins become phosphorylated upon odorant stimulation of olfactory cilia preparations; this was demonstrated by immunoprecipitation experiments employing the sequence-directed, receptor-specific antibodies. Functional assays revealed that the receptor-specific antibodies significantly attenuate second messenger signalling elicited by inositol 1,4,5-trisphosphate-inducing odorants, whereas activation of the cAMP cascade by appropriate odorants was not affected. These observation indicate that the sequence-specific antibodies not only recognize odorant receptors, but also discriminate between receptor subtypes coupling to different second-messenger pathways.

Amino Acid Sequence↗

PubMed1994

Rostro-caudal patterning of receptor-expressing olfactory neurones in the rat nasal cavity.

The rostro-caudal extent of odorant receptor expression zones in the rat olfactory epithelium was analysed by means of in situ hybridization. Three broad non-overlapping zones were identified that extended along almost the entire anterior-posterior axis; each zone was composed of several separate bands running anterior to posterior throughout the olfactory epithelium. Super-imposed onto these broad zones was the expression area of a particular receptor subtype (OR37); it was restricted to a small region of the epithelial sheet with a high density of reactive neurones in the centre and declining numbers towards the periphery of the region. A quantitative evaluation of the reactive cells revealed that, despite their different distribution patterns, all receptor subtypes were expressed in an equal number of neurones.

Animals↗

PubMed1994

Olfactory neurones expressing distinct odorant receptor subtypes are spatially segregated in the nasal neuroepithelium.

In situ hybridization techniques have been employed to explore the olfactory epithelium of the rat for the distribution of odorant receptor gene transcripts. We demonstrate that olfactory neurone subpopulations expressing distinct receptor subtypes are spatially segregated within the olfactory epithelium. A compartmentalization of the neuroepithelium into distinct expression zone is apparent, cells expressing a specific receptor are randomly distributed within a given zone. Structurally related receptor subtypes share a common distribution pattern.

Animals↗

PubMed1992

Expression of odorant receptors in spatially restricted subsets of chemosensory neurones.

From a rat olfactory library a cDNA clone (OR37) which is supposed to encode an odorant receptor protein has been isolated and characterized. Specific antisense RNA and in situ hybridization techniques have been employed to monitor the olfactory epithelium for the distribution of olfactory neurones expressing the OR37-gene. The OR37-transcripts were detected only in a subset of receptor cells segregated in two restricted areas of the olfactory epithelium. The clusters of reactive cells appear symmetrically in both nasal cavities. Within a reactive region only a subset of the cells expressed the receptor. The segregation of neurones expressing a distinct receptor supports the notion that a spatial component may be involved in coding odour quality.

Amino Acid Sequence↗

PubMedDate not supplied

Subcellular localization of specific mRNAs and their protein products in Purkinje cells by combined fluorescence in situ hybridization and immunocytochemistry.

In this study we have investigated the subcellular distribution of two mRNAs coding for the Purkinje cell-specific proteins, calbindin D28K and L7 (L7/pcp-2). Whereas calbindin mRNA was found to be in the cell body only, L7 transcripts could be detected within the molecular layer, corresponding to Purkinje cell dendrites. We have now combined a highly sensitive fluorescence-based in situ hybridization protocol with immunofluorescence in conjunction with confocal optical sectioning to analyze the precise localization of these mRNAs in individual Purkinje neurons. We show that L7 mRNA is localized in clusters within the proximal and distal branches of dendrites, but also in the proximal part of Purkinje cell axons. In contrast, calbindin transcripts are restricted to the axonal pole of the perikaryon. Purkinje cells grown in primary cultures reveal similar mRNA distribution patterns for the two transcripts. Thus, the mechanism underlying localization of mRNA within Purkinje cells seems to function in a cell-intrinsic manner, guiding specific transcripts, such as L7 mRNA, to neuronal processes while restricting others, such as calbindin mRNA, to the perikaryon.

Alkaline Phosphatase↗