Characterization of inositol-1,4,5-trisphosphate-gated channels in the plasma membrane of rat olfactory neurons.
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It is generally accepted that inositol-1,4,5-trisphosphate (InsP3) plays a role in olfactory transduction. However, the precise mode of action of InsP3 remains controversial. We have characterized the conductances activated by the addition of 10 microM InsP3 to excised patches of soma plasma membrane from rat olfactory neurons. InsP3 induced current fluctuations in 25 of 121 inside-out patches. These conductances could be classified into two groups according to the polarity of the current at a holding potential of +40 to +60 mV (with Ringer's in the pipette and pseudointracellular solution in the bath). Conductances mediating outward currents could be further divided into large- (64 +/- 4 pS, n = 4) and small- (16 +/- 1.7 pS, n = 11) conductance channels. Both small- and large-conductance channels were nonspecific cation channels. The large-conductance channel displayed bursting behavior at +40 mV, with flickering increasing at negative holding potentials to the point where single-channel currents were no longer discernible. The small-conductance channel did not display flickering behavior. The conductance mediating inward currents at +40 to +60 mV reversed at +73 +/- 4 mV (n = 4). The current traces displayed considerable fluctuations, and single-channel currents could not be discerned. The current fluctuations returned to baseline after removal of InsP3. The power density spectrum for the excess noise generated by InsP3 followed a 1/f dependence consistent with conductance fluctuations in the channel mediating this current, although other mechanisms are not excluded. These experiments demonstrate the presence of plasma membrane InsP3-gated channels of different ionic specificity in olfactory receptor cells. (+info)
Coherent oscillations in membrane potential synchronize impulse bursts in central olfactory neurons of the crayfish.
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Lateral protocerebral interneurons (LPIs) in the central olfactory pathway of the freshwater crayfish Procambarus clarkii reside within the lateral protocerebrum and receive direct input from projection neurons of the olfactory midbrain. The LPIs exhibit periodic (0.5 Hz) changes in membrane potential that are imposed on them synaptically. Acute surgical experiments indicate that the synaptic activity originates from a group of oscillatory neurons lying within the lateral protocerebrum. Simultaneous intracellular recordings from many LPI pairs indicate that this periodic synaptic input is synchronous and coherent among the population of approximately 200 LPIs on each side of the brain. In many LPIs, specific odors applied to antennules in isolated head preparations generate long-lasting excitatory postsynaptic potentials and impulse bursts. The impulse bursts are generated only near the peaks of the ongoing depolarizations, approximately 1 s after stimulus application, and so the periodic baseline activity is instrumental in timing burst generation. Simultaneous recordings from pairs of LPIs show that, when impulse bursts occur in both cells after an odorant stimulus, they are synchronized by the common periodic depolarizations. We conclude that the common, periodic activity in LPIs can synchronize impulse bursts in subsets of these neurons, possibly generating powerful long-lasting postsynaptic effects in downstream target neurons. (+info)
Combinatorial receptor codes for odors.
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The discriminatory capacity of the mammalian olfactory system is such that thousands of volatile chemicals are perceived as having distinct odors. Here we used a combination of calcium imaging and single-cell RT-PCR to identify odorant receptors (ORs) for odorants with related structures but varied odors. We found that one OR recognizes multiple odorants and that one odorant is recognized by multiple ORs, but that different odorants are recognized by different combinations of ORs. Thus, the olfactory system uses a combinatorial receptor coding scheme to encode odor identities. Our studies also indicate that slight alterations in an odorant, or a change in its concentration, can change its "code," potentially explaining how such changes can alter perceived odor quality. (+info)
A spatial map of olfactory receptor expression in the Drosophila antenna.
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Insects provide an attractive system for the study of olfactory sensory perception. We have identified a novel family of seven transmembrane domain proteins, encoded by 100 to 200 genes, that is likely to represent the family of Drosophila odorant receptors. Members of this gene family are expressed in topographically defined subpopulations of olfactory sensory neurons in either the antenna or the maxillary palp. Sensory neurons express different complements of receptor genes, such that individual neurons are functionally distinct. The isolation of candidate odorant receptor genes along with a genetic analysis of olfactory-driven behavior in insects may ultimately afford a system to understand the mechanistic link between odor recognition and behavior. (+info)
Functional identification and reconstitution of an odorant receptor in single olfactory neurons.
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The olfactory system is remarkable in its capacity to discriminate a wide range of odorants through a series of transduction events initiated in olfactory receptor neurons. Each olfactory neuron is expected to express only a single odorant receptor gene that belongs to the G protein coupled receptor family. The ligand-receptor interaction, however, has not been clearly characterized. This study demonstrates the functional identification of olfactory receptor(s) for specific odorant(s) from single olfactory neurons by a combination of Ca2+-imaging and reverse transcription-coupled PCR analysis. First, a candidate odorant receptor was cloned from a single tissue-printed olfactory neuron that displayed odorant-induced Ca2+ increase. Next, recombinant adenovirus-mediated expression of the isolated receptor gene was established in the olfactory epithelium by using green fluorescent protein as a marker. The infected neurons elicited external Ca2+ entry when exposed to the odorant that originally was used to identify the receptor gene. Experiments performed to determine ligand specificity revealed that the odorant receptor recognized specific structural motifs within odorant molecules. The odorant receptor-mediated signal transduction appears to be reconstituted by this two-step approach: the receptor screening for given odorant(s) from single neurons and the functional expression of the receptor via recombinant adenovirus. The present approach should enable us to examine not only ligand specificity of an odorant receptor but also receptor specificity and diversity for a particular odorant of interest. (+info)
Effects of phosphatidylinositol 4,5-bisphosphate and phosphatidylinositol 4-phosphate on a Na+-gated nonselective cation channel.
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Olfactory receptor neurons in the lobster express a nonselective cation channel that is activated by intracellular Na+ and carries a substantial part of the depolarizing receptor current. Here, we show that phosphatidylinositol 4,5-bisphosphate [PI(4,5)P2] and phosphatidylinositol 4-phosphate [PI(4)P] applied to the intracellular face of cell-free patches activate the channel in the absence of Na+ and that antibodies against the respective phospholipids irreversibly inhibit the evoked activity. Further, we show that applying PI(4,5)P2 or PI(4)P in the presence of Na+ decreases the concentration of Na+ required to activate the channel from an EC50 of 74 to 22 mM for PI(4,5)P2 and to 29 mM for PI(4)P, respectively. Na+-gated channel activity was irreversibly inhibited by monoclonal antibodies against PI(4,5)P2 and PI(4)P in patches never exposed to exogenous phosphatidylinositols, suggesting that endogenous inositol phospholipids are required for the activation of the channel by intracellular Na+. Our findings suggest that PI(4,5)P2 and/or PI(4)P may serve as intracellular signaling molecules in these primary sensory neurons and provide a general mechanism to explain how the sensitivity of Na+-gated channels to Na+ could be much greater in intact cells than in excised membrane patches. (+info)
Small stress protein Hsp27 accumulation during dopamine-mediated differentiation of rat olfactory neurons counteracts apoptosis.
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The small stress protein Hsp27 is expressed during mammalian neural development. We have analyzed the role of this protein in immortalized rat olfactory neuroblasts. In the presence of dopamine a fraction of these cells differentiate into neurons while the remaining cells undergo apoptosis. We report here that the dopamine induced differentiation and apoptosis are associated with a transient and specific accumulation of Hsp27. Moreover, transfection experiments have shown that Hsp27 overexpression drastically decreases the fraction of cells undergoing apoptosis. In contrast, reduction of the endogenous level of Hsp27 led to abortion of differentiation and, therefore, drastically increased the number of apoptotic cells. Furthermore, in the normal cell population we show that Hsp27 accumulation takes place only in differentiating cells that were not undergoing apoptosis. We therefore conclude that Hsp27 may represent a key protein that controls the decision of olfactory precursor cells to undergo either differentiation or cell death. (+info)
Variable patterns of axonal projections of sensory neurons in the mouse vomeronasal system.
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The vomeronasal system mediates pheromonal effects in mammals. We have employed gene targeting technology to introduce mutations in a putative pheromone receptor gene, VR2, in the germline of mice. By generating alleles differentially tagged with the histological markers taulacZ and tauGFP, we show that VR2 is monoallelically expressed in a given neuron. Axons of VR2-expressing neurons converge onto numerous glomeruli in the accessory olfactory bulb. The pattern of axonal projections is complex and variable. This wiring diagram is substantially different from that of the main olfactory system. The projection pattern is disrupted by deleting the coding region of VR2, but an unrelated seven-transmembrane protein, the odorant receptor M71, can partially substitute for VR2. (+info)