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Marc Spehr

Publications and source records attributed to Marc Spehr.

15 recordsLinked to original sources
PubMed2006

Beta-arrestin2-mediated internalization of mammalian odorant receptors.

Odorant receptors comprise the biggest subfamily of G-protein-coupled receptors. Although the endocytic mechanisms of other G-protein-coupled receptors have been characterized extensively, almost nothing is known about the intracellular trafficking of odorant receptors. The present study describes the endocytic pathway of mammalian odorant receptors, which bind beta-arrestin2 with high affinity and are internalized via a clathrin-dependent mechanism. After prolonged odorant exposure, receptors are not targeted to lysosomal degradation but accumulate in recycling endosomes. Odorant-induced odorant receptor desensitization is promoted by cAMP-dependent protein kinase A phosphorylation and is dependent on serine and threonine residues within the third intracellular loop of the receptor. Moreover, beta-arrestin2 is redistributed into the dendritic knobs of mouse olfactory receptor neurons after treatment with a complex odorant mixture. Prolonged odorant exposure resulted in accumulation of beta-arrestin2 in intracellular vesicles. Adaptation of olfactory receptor neurons to odorants can be abolished by the inhibition of clathrin-mediated endocytosis, showing the physiological relevance of the here described mechanism of odorant receptor desensitization. A better understanding of odorant receptor trafficking and additional insight into the molecular determinants underlying the interactions of odorant receptors with beta-arrestin2 and other trafficking proteins will therefore be important to fully understand the mechanisms of adaptation and sensitization in the olfactory epithelium.

Amino Acid Motifs↗
PubMed2006

Loss of CNGB1 protein leads to olfactory dysfunction and subciliary cyclic nucleotide-gated channel trapping.

Olfactory receptor neurons (ORNs) employ a cyclic nucleotide-gated (CNG) channel to generate a receptor current in response to an odorant-induced rise in cAMP. This channel contains three types of subunits, the principal CNGA2 subunit and two modulatory subunits (CNGA4 and CNGB1b). Here, we have analyzed the functional relevance of CNGB1 for olfaction by gene targeting in mice. Electro-olfactogram responses of CNGB1-deficient (CNGB1-/-) mice displayed a reduced maximal amplitude and decelerated onset and recovery kinetics compared with wild-type mice. In a behavioral test, CNGB1-/- mice exhibited a profoundly decreased olfactory performance. Electrophysiological recordings revealed that ORNs of CNGB1-/- mice weakly expressed a CNG current with decreased cAMP sensitivity, very rapid flicker-gating behavior and no fast modulation by Ca2+-calmodulin. Co-immunoprecipitation confirmed the presence of a CNGA2/CNGA4 channel in the olfactory epithelium of CNGB1-/- mice. This CNGA2/CNGA4 channel was targeted to the plasma membrane of olfactory knobs, but failed to be trafficked into olfactory cilia. Interestingly, we observed a similar trafficking defect in mice deficient for the CNGA4 subunit. In conclusion, these results demonstrate that CNGB1 has a dual function in vivo. First, it endows the olfactory CNG channel with a variety of biophysical properties tailored to the specific requirements of olfactory transduction. Second, together with the CNGA4 subunit, CNGB1 is needed for ciliary targeting of the olfactory CNG channel.

Animals↗
PubMed2006

Essential role of the main olfactory system in social recognition of major histocompatibility complex peptide ligands.

Genes of the major histocompatibility complex (MHC), which play a critical role in immune recognition, influence mating preference and other social behaviors in fish, mice, and humans via chemical signals. The cellular and molecular mechanisms by which this occurs and the nature of these chemosignals remain unclear. In contrast to the widely held view that olfactory sensory neurons (OSNs) in the main olfactory epithelium (MOE) are stimulated by volatile chemosignals only, we show here that nonvolatile immune system molecules function as olfactory cues in the mammalian MOE. Using mice with targeted deletions in selected signal transduction genes (CNGA2, CNGA4), we used a combination of dye tracing, electrophysiological, Ca2+ imaging, and behavioral approaches to demonstrate that nonvolatile MHC class I peptides activate subsets of OSNs at subnanomolar concentrations in vitro and affect social preference of male mice in vivo. Both effects depend on the cyclic nucleotide-gated (CNG) channel gene CNGA2, the function of which in the nose is unique to the main population of OSNs. Disruption of the modulatory CNGA4 channel subunit reveals a profound defect in adaptation of peptide-evoked potentials in the MOE. Because sensory neurons in the vomeronasal organ (VNO) also respond to MHC peptides but do not express CNGA2, distinct mechanisms are used by the mammalian main and accessory olfactory systems for the detection of MHC peptide ligands. These results suggest a general role for MHC peptides in chemical communication even in those vertebrates that lack a functional VNO.

Animals↗
PubMed2006

Odorant receptors and olfactory-like signaling mechanisms in mammalian sperm.

Since their discovery in 1991, members of the odorant receptor (OR) family have been found in various ectopic tissues, including testis and sperm. It took, however, more than a decade for the first mammalian testicular ORs to be functionally characterized and implicated in a reproductively relevant scenario. Activation of hOR17-4 and mOR23 in human and mouse sperm, respectively, mediates distinct flagellar motion patterns and chemotactic behavior in various bioassays. For hOR17-4, receptor function and downstream signal transduction events are shown to be subject to pharmacological manipulation. Further insight into the basic principles that govern sperm OR operation as well as into the molecular logic that underlies OR-mediated signaling could set the stage for pioneering future applications in procreation and/or contraception.

Animals↗
PubMed2006

Pheromonal recognition memory induced by TRPC2-independent vomeronasal sensing.

Detection and transduction of pheromonal signals by the mouse vomeronasal organ (VNO) is critical for the formation of a persistent memory required for mate recognition in the context of selective pregnancy failure (the Bruce effect). This pregnancy block can be mediated by peptide ligands of disparate major histocompatibility complex (MHC) molecules, but little is known about the molecular mechanisms underlying this effect. Given the proposed key role of the transient receptor potential channel, TRPC2, in VNO signal transduction, we tested whether TRPC2 is essential for memory formation in the context of the Bruce effect. Surprisingly, the loss of the TRPC2 channel did not significantly influence memory formation, whereas surgical lesions of the VNO caused a profound deficit. Furthermore, field potential and single-cell patch-clamp recordings showed that TRPC2 is dispensable for the transduction of MHC peptide ligands by sensory neurons in the basal zone of the VNO. This indicates that a previously unrecognized TRPC2-independent signal transduction mechanism in the VNO underlies the sensing of cues required for the formation of this pheromonal recognition memory.

Animals↗
PubMed2005

One neuron--multiple receptors: increased complexity in olfactory coding?

Olfaction--the sense of smell--is responsible for detecting molecules of immense structural variety. Precise recognition of such diverse stimuli requires a massive receptor repertoire. This functional challenge has been met by simultaneous expression of a multitude of odor-detecting receptors that all belong to the superfamily of heterotrimeric GTP-binding protein (G protein)-coupled receptors. Studies conducted over the past decade have led to the assumption that an individual olfactory sensory neuron expresses only a single odorant receptor, consequently giving rise to the "one receptor-one neuron" hypothesis. This idea is attractive because of its simplicity and has served as the basis for models of olfactory coding. However, recent reports regarding Drosophila have found exceptions to the rule that could have important implications for the logic of olfactory coding.

Animals↗
PubMed2005

A potential role of odorant receptor agonists and antagonists in the treatment of infertility and contraception.

In 1992, the identification of odorant receptor expression in mammalian testicular tissue prepared the ground for an ongoing debate about a potential role for these chemoreceptors in significant sperm behaviors, in particular chemotaxis. The identification of hOR17-4, a human testicular odorant receptor that mediates sperm chemotaxis in various bioassays, revealed the first potential key player in this reproductively relevant scenario. Detailed knowledge of the receptor's molecular receptive field, the discovery of a potent receptor antagonist, as well as specific insight into the receptor-linked signaling cascade(s), could establish a basis for pioneering future applications in fertility treatment and/or contraception.

Contraception↗
PubMed2004

Particulate adenylate cyclase plays a key role in human sperm olfactory receptor-mediated chemotaxis.

Human sperm chemotaxis is a critical component of the fertilization process, but the molecular basis for this behavior remains unclear. Recent evidence shows that chemotactic responses depend on activation of the sperm olfactory receptor, hOR17-4. Certain floral scents, including bourgeonal, activate hOR17-4, trigger pronounced Ca(2+) fluxes, and evoke chemotaxis. Here, we provide evidence that hOR17-4 activation is coupled to a cAMP-mediated signaling cascade. Multidimensional protein identification technology was used to identify potential components of a G-protein-coupled cAMP transduction pathway in human sperm. These products included various membrane-associated adenylate cyclase (mAC) isoforms and the G(olf)-subunit. Using immunocytochemistry, specific mAC isoforms were localized to particular cell regions. Whereas mAC III occurred in the sperm head and midpiece, mAC VIII was distributed predominantly in the flagellum. In contrast, G(olf) was found mostly in the flagellum and midpiece. The observed spatial distribution patterns largely correspond to the spatiotemporal character of hOR17-4-induced Ca(2+) changes. Behavioral and Ca(2+) signaling responses of human sperm to bourgeonal were bioassayed in the presence, or absence, of the adenylate cyclase antagonist SQ22536. This specific agent inhibits particulate AC, but not soluble AC, activation. Upon incubation with SQ22536, cells ceased to exhibit Ca(2+) signaling, chemotaxis, and hyperactivation (faster swim speed and flagellar beat rate) in response to bourgeonal. Particulate AC is therefore required for induction of hOR17-4-mediated human sperm behavior and represents a promising target for future design of contraceptive drugs.

Adenine↗
PubMed2004

Subunit-specific P2X-receptor expression defines chemosensory properties of trigeminal neurons.

The facial innervation pattern of trigeminal nerve fibres comprises the innervation of the nasal epithelium, where free trigeminal nerve endings contribute to detection and discrimination of chemical stimuli including odourants. The signal transduction mechanisms in sensory nerve endings underlying perception of chemical stimuli remain widely uncovered. Here, we characterized trigeminal ATP-activated P2X receptors in cultured rat trigeminal neurons and investigated their role in chemoperception. We identified a new subpopulation of neurons lacking typical nociceptive characteristics and expressing homomeric P2X(2) receptors. Using a certain group of chemicals known as trigeminal stimuli we found no direct activation of trigeminal neurons, but a modulation of P2X(2) receptor mediated currents. In contrast, P2X(3) receptor mediated currents of nociceptive trigeminal neurons remained unaffected by the tested chemicals. Therefore, we assume a functional role for the newly identified subpopulation in chemodetection of certain trigeminal stimuli.

Adenosine Triphosphate↗
PubMed2004

hOR17-4 as a potential therapeutic target.

More than a decade ago, the unexpected finding of olfactory receptor expression in human testicular tissue led to speculation about a potential role of these chemoreceptors in various aspects of mature sperm behavior, especially sperm chemotaxis. Recently, first evidence in favor of this hypothesis was provided by the identification of hOR17-4, a testicular olfactory receptor that mediates human sperm chemotaxis in various bioassays. A detailed characterization of the receptor's molecular receptive range as well as the first description of a potent receptor antagonist could provide the basis for future applications in fertility treatment with important consequences in procreation and/or contraception.

Chemotactic Factors↗
PubMed2003

Identification of a testicular odorant receptor mediating human sperm chemotaxis.

Although it has been known for some time that olfactory receptors (ORs) reside in spermatozoa, the function of these ORs is unknown. Here, we identified, cloned, and functionally expressed a previously undescribed human testicular OR, hOR17-4. With the use of ratiofluorometric imaging, Ca2+ signals were induced by a small subset of applied chemical stimuli, establishing the molecular receptive fields for the recombinantly expressed receptor in human embryonic kidney (HEK) 293 cells and the native receptor in human spermatozoa. Bourgeonal was a powerful agonist for both recombinant and native receptor types, as well as a strong chemoattractant in subsequent behavioral bioassays. In contrast, undecanal was a potent OR antagonist to bourgeonal and related compounds. Taken together, these results indicate that hOR17-4 functions in human sperm chemotaxis and may be a critical component of the fertilization process.

Adenylyl Cyclases↗
PubMed2002

Arachidonic acid plays a role in rat vomeronasal signal transduction.

Sensory neurons of the vomeronasal organ (VNO) detect volatile chemicals that are released by conspecific animals and convey information about social and reproductive behavior. The signal transduction pathway in vomeronasal receptor neurons (VRNs) is not known in detail, but is believed to be distinct from that of the sensory neurons of the main olfactory system. Many of the identified olfactory transduction components are not expressed by VRNs. Using Ca2+ imaging and electrophysiological recordings, we investigated the signal transduction pathway of urine perception and the possible role of polyunsaturated fatty acids (PUFAs) as intracellular messengers in freshly dissociated rat VNO neurons. We found that application of urine induced a transient increase in intracellular Ca2+ that was dependent on the activity of phospholipase C and diacylglycerol (DAG) lipase. The Ca2+ transient was not dependent on depletion of intracellular Ca2+ stores but was dependent on the presence of extracellular Ca2+. Furthermore, the urine response was not sensitive to modulators of adenylate cyclase and inhibitors of inositol 1,4,5-trisphosphate receptors. Application of PUFAs (linolenic acid and arachidonic acid, synthesized in living cells from DAG) also elicited Ca2+ transients in fura 2 measurements and inward currents in whole-cell voltage-clamp recordings. Pharmacological inhibition of lipoxygenase and cyclooxygenase induced a transient increase in intracellular Ca2+, possibly by increasing the endogenous level of PUFAs, leading to activation of transduction channels. These data provide evidence for a role of PUFAs in rat vomeronasal signal transduction.

Adenylyl Cyclase Inhibitors↗
PubMed2002

3-phosphoinositides modulate cyclic nucleotide signaling in olfactory receptor neurons.

Phosphatidylinositol 3-kinase (PI3K)-dependent phosphoinositide signaling has been implicated in diverse cellular systems coupled to receptors for many different ligands, but the extent to which it functions in sensory transduction is yet to be determined. We now report that blocking PI3K activity increases odorant-evoked, cyclic nucleotide-dependent elevation of [Ca(2+)](i) in acutely dissociated rat olfactory receptor neurons and does so in an odorant-specific manner. These findings imply that 3-phosphoinositide signaling acts in vertebrate olfactory transduction to inhibit cyclic nucleotide-dependent excitation of the cells and that the interaction of the two signaling pathways is important in odorant coding, indicating that 3-phosphoinositide signaling can play a role in sensory transduction.

Adaptation, Physiological↗