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H Breer

Publications and source records attributed to H Breer.

At least 91 records · Page 5Linked to original sources
PubMed1995

Topographic patterns of odorant receptor expression in mammals: a comparative study.

In a comparative study, molecular probes for various odorant receptor subtypes were employed in in situ hybridization experiments on tissue sections through the nose from different mammalian species. OR37 reactive neurons were found exclusively in the rodent species, where they were clustered in very similar position within the nasal cavities; an OR37-related receptor subtype was not detectable in the rabbit. All other subtypes tested, hybridized across species borders to neurons that were distributed within the distinct zones of the olfactory epithelium. Most receptor types were found in the same zone in all species; however, a few subtypes which are expressed in the medial zone in rat were found in the dorsal zone in guinea pig.

Animals↗
PubMed1995

Presynaptic calcium channels: pharmacology and regulation.

Voltage-dependent Ca2+ channels are considered as molecular trigger elements for signal transmission at chemical synapses. Due to their central role in this fundamental process, function and pharmacology of presynaptic Ca2+ channels have recently been the subject of extensive exploration employing various experimental techniques. Several lines of evidence indicate that, at nerve terminals in higher vertebrates, the evoked influx of Ca2+ -ions is mainly mediated by Ca2+ channels of the P-type. The stringent regulation of presynaptic Ca2+ channels is supposed to be involved in fine-tuning the efficiency of synaptic transmission. Intrinsic control mechanisms, such as voltage- or Ca(2+)-dependent inactivation, or modulation of channel activity, either by G-proteins directly or via phosphorylation by protein kinases, may be of particular functional importance.

Animals↗
PubMed1995

Two classes of olfactory receptors in Xenopus laevis.

Xenopus laevis possess a gene repertoire encoding two distinct classes of olfactory receptors: one class related to receptors of fish and one class similar to receptors of mammals. Sequence comparison indicates that the fish-like receptors represent closely related members of only two subfamilies, whereas mammalian-like receptors are more distantly related, most of them representing a different subfamily. The fish-like receptor genes are exclusively expressed in the lateral diverticulum of the frog's nose, specialized for detecting water-soluble odorants, whereas mammalian-like receptors are expressed in sensory neurons of the main diverticulum, responsible for the reception of volatile odors.

Amino Acid Sequence↗
PubMed1994

Positive feedback between acetylcholine and the neurotrophins nerve growth factor and brain-derived neurotrophic factor in the rat hippocampus.

In the rat hippocampus, nerve growth factor (NGF) and brain-derived neurotrophic factor (BDNF) are synthesized by neurons in an activity-dependent manner. Glutamate receptor activation increases whereas GABAergic stimulation decreases NGF and BDNF mRNA levels. Here we demonstrate that NGF and BDNF mRNA and NGF protein are up-regulated in the rat hippocampus by the activation of muscarinic receptors. Conversely, NGF and BDNF enhance the release of acetylcholine (ACh) from rat hippocampal synaptosomes containing the nerve endings of the septal cholinergic neurons. NGF also rapidly increases the high-affinity choline transport into synaptosomes. The reciprocal regulation of ACh, NGF and BDNF in the hippocampus suggests a novel molecular framework by which the neurotrophins might influence synaptic plasticity.

Acetylcholine↗
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

Olfactory desensitization requires membrane targeting of receptor kinase mediated by beta gamma-subunits of heterotrimeric G proteins.

Olfaction is mediated by G protein-coupled receptors. In isolated rat olfactory cilia, odorants such as citralva stimulate a burst of cAMP, which peaks in 50 ms and returns almost to base-line level within 150 ms in the continuing presence of odorant. This desensitization is mediated by the cAMP dependent protein kinase and a specialized G protein-coupled receptor kinase originally termed beta ARK2 (GRK3). In vitro experiments suggest that the prenylated beta gamma-subunits of heterotrimeric G proteins target the cytosolic beta ARK1 (GRK2) enzyme to its membrane bound receptor substrate by binding to sites in its carboxyl terminus. Here we demonstrate that odorants stimulate translocation of GRK3 from cytosol to membranes in isolated rat olfactory cilia. We introduced a glutathione S-transferase-GRK3ct fusion protein, containing the carboxyl-terminal 222 amino acid residues of GRK3, which includes the beta gamma binding site, or a 28-amino acid peptide derived therefrom, into permeabilized cilia preparations. These reagents block odorant-mediated enzyme translocation and desensitization while markedly attenuating odorant-stimulated phosphorylation of olfactory proteins. These findings suggest that beta gamma-subunits may physiologically regulate a G protein-coupled receptor kinase and that enzyme translocation may be a general and required feature of the activity of some members of this enzyme family.

Animals↗
PubMed1994

Odor recognition and second messenger signaling in olfactory receptor neurons.

The detection of volatile odorants is supposed to begin with their interaction with soluble binding proteins which shuttle the hydrophobic ligands through the aqueous mucus layer towards specific odorant receptors in the ciliary membrane of olfactory neurons. A large family of receptors for odorants has been identified recently; individual receptor types are expressed in subsets of cells distributed in distinct zones of the olfactory epithelium. Ligand-receptor interaction triggers a rapid multistep reaction cascade, ultimately leading to an electrical response of the receptor neuron. Olfactory signaling is terminated by phosphorylation of receptors via a negative feedback reaction catalyzed by two types of kinases.

Animals↗
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↗
PubMed1994

GABAB-receptor-mediated inhibition of calcium signals in isolated nerve terminals.

Time-resolved fluorometric monitoring of rapid changes in intracellular Ca(2+)-concentrations [Ca2+]i was employed to analyze the effect of gamma-aminobutyric acid (GABA) on evoked Ca(2+)-signals in rat hippocampal synaptosomes. The inhibitory action of GABA was mimicked by Baclofen, the selective agonist for GABAB-receptors, and also by the nonhydrolyzable GTP-derivative GTP-gamma-S. Preincubation of synaptosomes with GABA or baclofen for up to 250 ms before depolarization resulted in a maximal inhibition. The GABA-induced attenuation of the evoked rise in [Ca2+]i was maximal during the first milliseconds after depolarization. The inhibitory action of GABA apparently does not involve second messengers, such as cAMP or IP3/DAG; the GABA-induced inhibition of presynaptic voltage-dependent Ca(2+)-channels may be mediated directly by G-proteins.

Animals↗
PubMed1994

Signal recognition and chemo-electrical transduction in olfaction.

The mimicking of olfaction is considered to be a promising approach for the construction of artificial odour-sensing systems. In the nose, the detection of volatile odorants begins when the odorant ligands interact with specific odorant receptors in the ciliary membrane of the olfactory neurons. A large family of genes encoding putative odorant receptors has been identified recently. Individual receptor types are expressed in subsets of cells distributed in distinct zones of the olfactory epithelium. Ligand-receptor interaction triggers a rapid multistep reaction cascade, resulting in a "pulse" of second messengers that initiates an electrical response from the receptor neuron. Olfactory signalling is terminated by phosphorylation of receptors via a negative feedback reaction, catalyzed by specific kinases.

Animals↗
PubMed1994

Ca(2+)-dependent inactivation of P-type calcium channels in nerve terminals.

Rapid Ca2+ signals evoked by K+ depolarization of rat cerebral cortical synaptosomes were measured by dual-channel Ca2+ spectrofluorometry coupled to a stopped-flow device. Kinetic analysis of the signal rise phase at various extracellular Ca2+ concentrations revealed that the responsible voltage-dependent Ca2+ channels, previously identified as P-type Ca2+ channels, inactivate owing to the rise in intracellular Ca2+ levels. At millimolar extracellular Ca2+ concentrations the channels were inactivated very rapidly and the rate was dependent on the high influx rate of Ca2+, thus limiting the Ca2+ signal amplitudes to 500-600 nM. A slower, probably voltage-dependent regulation appears to be effective at lower Ca2+ influx rates, leading to submaximal Ca2+ signal amplitudes. The functional feedback regulation of calcium channels via a sensor for intracellular Ca2+ levels appears to be responsible for the different inhibition characteristics of Cd2+ versus omega-agatoxin IVa.

Animals↗
PubMed1994

Single odors differentially stimulate dual second messenger pathways in lobster olfactory receptor cells.

Quench-flow measurements are used to determine the subsecond kinetics of odor-induced changes in second messenger concentrations in lobster olfactory receptor neurons. Individual odors transiently and differentially increase the production of both adenosine cAMP and inositol 1,4,5-trisphosphate (IP3) within 50 msec of odor stimulation. The ability of two different odors to stimulate cAMP and IP3 correlates with the odors' ability to excite and inhibit receptor cells physiologically. These results strengthen the proposition, heretofore based largely on evidence from cultured cells, that dual second messenger pathways mediate excitatory and inhibitory input to lobster olfactory receptor cells.

Animals↗
PubMed1993

Neurotrophin induced cAMP and IP3 responses in PC12 cells. Different pathways.

NGF and BDNF elevate cAMP and IP3 levels in membranes of PC12 cells within a subsecond time period. The cAMP formation induced by NGF and BDNF pretreatment for 2 s was reduced by GDP-beta-S and PTX, but not the trkNGFR inhibitor K 252a. NGF, but not BDNF, induced IP3 formation. IP3 formation was reduced by K 252a, but not by GDP-beta-S and PTX. Using p75NGFR expressing, but trkNGFR-deficient PCNA cell membranes, NGF and BDNF induced cAMP formation, but not IP3 formation. We suggest that NGF and BDNF induced cAMP formation is mediated via a p75NGFR/G-protein mediated mechanism, and IP3 formation via a K 252a sensitive pathway.

Animals↗
PubMed1993

Transfected cGMP-dependent protein kinase suppresses calcium transients by inhibition of inositol 1,4,5-trisphosphate production.

cGMP is a key regulatory molecule in visual transduction, integration of neuronal response to excitatory neurotransmitters, relaxation of smooth muscle, intestinal secretion of water and salt, and reabsorption of sodium and water in the distal tubules of the nephron. Some of these cellular functions are associated with the activation of cGMP kinase and a decrease in cytosolic calcium levels ([Ca2+]i). The mechanism by which cGMP kinase lowers [Ca2+]i is controversial. We have used CHO cells stably transfected with cGMP kinase to test several of the proposed [Ca2+]i-lowering mechanisms. Thrombin induces a calcium transient in wild-type and cGMP kinase-expressing CHO cells by releasing calcium from intracellular stores. Preincubation of wild-type cells with 8-bromo-cGMP had no effect on the calcium transient, whereas 8-bromo-cGMP prevented the thrombin-stimulated calcium transient in cGMP kinase-expressing CHO cells. In both cell types 8-bromo-cGMP had no effect on [Ca2+]i transients induced by replacing extracellular sodium by tetramethylammonium, ruling out an effect of cGMP kinase on Ca(2+)-ATPases. However, cGMP kinase activation effectively suppressed thrombin-induced stimulation of inositol 1,4,5-trisphosphate production. These results show that cGMP kinase lowers [Ca2+]i by interfering with the inositol 1,4,5-trisphosphate synthesis.

Animals↗
PubMed1993

A beta-adrenergic receptor kinase-like enzyme is involved in olfactory signal termination.

We have previously shown that second-messenger-dependent kinases (cAMP-dependent kinase, protein kinase C) in the olfactory system are essential in terminating second-messenger signaling in response to odorants. We now document that subtype 2 of the beta-adrenergic receptor kinase (beta ARK) is also involved in this process. By using subtype-specific antibodies to beta ARK-1 and beta ARK-2, we show that beta ARK-2 is preferentially expressed in the olfactory epithelium in contrast to findings in most other tissues. Heparin, an inhibitor of beta ARK, as well as anti-beta ARK-2 antibodies, (i) completely prevents the rapid decline of second-messenger signals (desensitization) that follows odorant stimulation and (ii) strongly inhibits odorant-induced phosphorylation of olfactory ciliary proteins. In contrast, beta ARK-1 antibodies are without effect. Inhibitors of protein kinase A and protein kinase C also block odorant-induced desensitization and phosphorylation. These data suggest that a sequential interplay of second-messenger-dependent and receptor-specific kinases is functionally involved in olfactory desensitization.

Animals↗
PubMed1993

Cloning and expression of odorant receptors.

Myriads of odorous molecules that vary widely in structure are nevertheless readily detected and discriminated by the sense of smell, but how this is achieved by the olfactory system has been a long-standing puzzle. Several different models have been proposed, and previous observations indicate that the recognition sites for odorous molecules could be G-protein-coupled receptor proteins, an idea supported by the discovery of a new gene family that probably encodes a diversity of odorant receptors. Here we report the identification of new members of the gene family encoding putative odorant receptors and demonstrate that they are indeed transcribed in olfactory receptor neurons. Furthermore, the receptor-encoding complementary DNA is expressed in non-neuronal surrogate cells, which generate second messenger responses upon stimulation with appropriate odorants, indicating that the receptors recognize odorants and couple to G proteins of the host cells.

Amino Acid Sequence↗