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Spikes were evoked in rat olfactory sensory neuron (OSN) populations by electrical stimulation of the olfactory bulb nerve layer in pentobarbital anesthetized rats. The latencies and recording positions for these compound spikes showed that they originated in olfactory epithelium. Dual simultaneous recordings indicated conduction velocities in the C-fiber range, around 0.5 m/s. These spikes are concluded to arise from antidromically activated olfactory sensory neurons. Electrical stimulation at 5 Hz was used to track changes in the size and latency of the antidromic compound population spike during the odor response. Strong odorant stimuli suppressed the spike size and prolonged its latency. The latency was prolonged throughout long odor stimuli, indicating continued activation of olfactory receptor neuron axons. The amounts of spike suppression and latency change were strongly correlated with the electroolfactogram (EOG) peak size evoked at the same site across odorants and across stimulus intensities. We conclude that the curve of antidromic spike suppression gives a reasonable representation of spiking activity in olfactory sensory neurons driven by odorants and that the correlation of peak spike suppression with the peak EOG shows the accuracy of the EOG as an estimate of intracellular potential in the population of olfactory sensory neurons. In addition, these results have important implications about traffic in olfactory nerve bundles. We did not observe multiple peaks corresponding to stimulated and unstimulated receptor neurons. This suggests synchronization of spikes in olfactory nerve, perhaps by ephaptic interactions. The long-lasting effect on spike latency shows that action potentials continue in the nerve throughout the duration of an odor stimulus in spite of many reports of depolarization block in olfactory receptor neuron cell bodies. Finally, strong odor stimulation caused almost complete block of antidromic spikes. This indicates that a very large proportion of olfactory axons was activated by single strong odor stimuli.

Responses of the rat olfactory epithelium were assessed with the electroolfactogram while odorants were presented to the external nares with an artificial sniff or to the internal nares by positive pressure. A series of seven odorants that varied from very polar, hydrophilic odorants to very nonpolar, hydrophobic odorants were used. Although the polar odorants activated the dorsal olfactory epithelium when presented by the external nares (orthonasal presentation), they were not effective when forced through the nasal cavity from the internal nares (retronasal presentation). However, the nonpolar odorants were effective in both stimulus modes. These results were independent of stimulus concentration or of humidity of the carrier air. Similar results were obtained with multiunit recordings from olfactory bulb. These results help to explain why human investigations often report differences in the sensation or ability to discriminate odorants presented orthonasally versus retronasally. The results also strongly support the importance of odorant sorption in normal olfactory processes.

Electroolfactogram recordings were made with a four-electrode assembly from the olfactory epithelium overlying the endoturbinate bones facing the nasal septum. In this study we tested whether odors of different chemical structures produce maximal responses along longitudinally oriented regions following the olfactory receptor gene expression zones described in the literature. The distribution of responses along the dorsal-to-ventral direction of this epithelium (i.e., across the expression zones) was tested in two types of experiments. In one, four electrodes were fixed along the dorsal-to-ventral axis of one turbinate bone. In the other, four electrodes were placed in corresponding positions on four turbinate bones and moved together up toward the top of the bone. These experiments compared the odorants limonene and alpha-terpinene, which are simple hydrocarbons, with carvone and menthone, which differ from the hydrocarbons by the presence of ketone groups. All responses were standardized to an amyl acetate or ethyl butyrate standard. The responses to limonene and alpha-terpinene were often larger for the ventral electrodes. The responses to carvone and menthone were largest for the dorsal electrodes. Intermediate electrodes gave responses that were intermediate in amplitude for these odors. The possibility that direction of air flow caused the observed response distributions was directly tested in experiments with odor nozzles placed in two positions. The relatively larger dorsal responses to carvone and relatively larger ventral responses to limonene were present despite odor nozzle position. We conclude that the responses to this set of odors vary systematically in a fashion parallel to the four gene expression zones. The odorant property that governs this response distribution may be related to the presence of oxygen-containing functional groups. Certain odors evoked larger responses at the intermediate electrode sites than at other sites. Cineole was the best example of this effect. This observation shows that not all oxygen-containing functional groups produce the same effect. Although we cannot exclude other possible mechanisms, these three response gradients may be produced by the four receptor expression zones described for many of the putative olfactory receptor genes. Therefore many of the receptors in each zone may share common properties. It remains to be determined whether this zonal input is significant in central odor processing. However, the correlation of odor chemical properties with the structure of receptor molecules in each zone may provide significant leads to structure-function relationships in vertebrate olfaction.

1. Electroolfactogram (EOG) recordings were made in three configurations from the rat olfactory epithelium. Each configuration compared recordings in the dorsomedial recess of the epithelium with recordings in ventral or lateral parts of the epithelium. Most comparisons were made with simultaneous recordings. The exception was a series in which the dorsal recess and lateral space between the base of two turbinate bones were directly exposed for odor application and recording. The spatial distributions of maximal responses were largely independent of recording configuration. 2. Simultaneous recordings compared dorsomedial and lateral sites in the epithelium during stimulation with a series of 50 odorants. The odorants that evoked larger responses in the lateral sites were usually compounds that lacked oxygen containing functional groups (such as the carbonyl group). This was true for straight chain and cyclic alkanes, for terpine compounds, and for aromatic compounds. The major exception was cineole, a bicyclic compound. All compounds containing ketone groups evoked larger dorsomedial responses. The responses of aldehydes and esters depended upon whether they were attached to aliphatic or aromatic chains. 3. In the three types of preparation, the sites responding best to ketones were in the same expression zone of the epithelium according to published maps for the rat and mouse. The sites responding best to odors without functional groups were in the far lateral or ventral region and corresponded to one of the two most lateral and ventral expression zones. This fact suggests that the receptors in these regions have a preference for particular chemical properties. This level of analysis cannot determine whether all receptors in each zone have a stronger response to certain properties of these odorants or whether each zone contains different proportions of receptors with these properties.

1. Electroolfactorgram (EOG) recordings were made from different regions of the rat olfactory epithelium to test for spatial distribution of odor responses. 2. The EOG recordings showed spatial distribution of the odor responses in the olfactory epithelium. While some odorants (amyl acetate, anisole, and ethyl butyrate) were more effective in evoking responses in the dorsal recess near the septum, other odorants (including limonene, cineole, cyclooctane, and hexane) were more effective in the lateral recesses among the turbinate bones. These differences were seen as statistically significant odorant-by-position interactions in analysis of variance. 3. Comparisons of recordings along the anteroposterior dimension of the epithelium produced smaller differences between the odor responses. These were not significant for 3-mm distances, but were statistically significant for 5- to 6-mm distances along the dorsomedial epithelium. 4. The latencies were significantly longer in the lateral recesses than in the medial region. This probably reflects a more tortuous air path along the turbinate bones to the lateral recesses. 5. The olfactory receptor cells were activated by antidromic stimulation via the nerve layer of the olfactory bulb. The population spikes evoked from the olfactory receptor cells could be suppressed by prior stimulation with odorants that evoked strong EOG responses. This collision of the antidromic action potentials with the odor-evoked action potentials indicates that the same population of receptor cells was activated in both cases. 6. The flow rate and duration of the artificial sniff were varied systematically in some experiments. The differential distribution of response sizes was present at all flow rates and sniff durations. Some odors (e.g., amyl acetate and anisole) produced increased responses in the epithelium of the lateral recesses when flow rates or sniff durations were high. We suggest that these changes may reflect the sorptive properties of the nasal membranes on these odors. The responses to other odors (e.g., hexane or limonene) were not greatly affected by flow rate or sniff duration. 7. Taken with existing anatomic data, the results indicate that the primary olfactory neurons that project axons to glomeruli in different parts of the olfactory bulb are responsive to different odors. The latency differences between responses at medial and lateral sites are large enough to be physiologically significant in the generation of the patterned responses of olfactory bulb neurons.

1. Intracellular recordings were made from the output neurons (mitral and tufted cells) of the rat olfactory bulb during electrical orthodromic stimulation of the olfactory nerve layer (ONL) and antidromic stimulation of the lateral olfactory tract and posterior piriform cortex (pPC) to test for physiological differences among the neuron types. Many of these neurons were identified by intracellular injections of biocytin, and others were identified by their pattern of antidromic activation. 2. Both marked and unmarked mitral cells showed large inhibitory postsynaptic potentials (IPSPs) in response to antidromic stimulation of the pPC, whereas tufted cells exhibited small IPSPs in response to pPC stimulation. Tufted cells, however, showed large IPSPs in response to ONL stimulation. In many cases, these tufted cell responses to ONL stimulation were larger than the mitral cell responses. The marked superficial tufted cells, those with basal dendrites in the superficial sublayer of the external plexiform layer (EPL), had the smallest IPSPs in response to pPC stimulation. These data support anatomic observations suggesting that the granule cell populations responsible for the IPSPs may be different for mitral and for superficial tufted cells. 3. The different types of output cells also showed differences in their responses to orthodromic stimulation. Type I mitral cells, which have basal dendrites confined to the deep sublayer of the EPL, were significantly less excitable by ONL stimulation than were the type II mitral cells, which have basal dendrites distributed within the intermediate sublayer of the EPL. Half of the type I mitral cells could not be excited at all by ONL stimulation. Superficial tufted cells showed even greater orthodromic excitability than type II mitral cells, usually responding to ONL stimulation with two or more spikes. 4. The ionic basis of the IPSPs in the superficial tufted cells appeared similar to those described for mitral cells. These IPSPs could be reversed by chloride injection and were associated with increased membrane conductance. 5. For both mitral and tufted cells, the number of ONL electrodes evoking IPSPs was greater than the number evoking spikes. These data suggest a kind of center-surround organization of inputs to these cells from the ONL, although this does not yet imply that the sensory receptive field of these output cells has a center-surround organization. 6. In conclusion, the properties of rat olfactory bulb output cells correlate with the sublayers of the EPL in which their basal dendrites lie.(ABSTRACT TRUNCATED AT 400 WORDS)

1. Intracellular recordings were made from 28 granule cells and 6 periglomerular cells of the rat olfactory bulb during odor stimulation and electrical stimulation of the olfactory nerve layer (ONL) and lateral olfactory tract (LOT). Neurons were identified by injection of horseradish peroxidase (HRP) or biocytin and/or intracellular response characteristics. Odorants were presented in a cyclic sniff paradigm, as reported previously. 2. All interneurons could be activated from a wide number of stimulation sites on the ONL, with distances exceeding their known dendritic spreads and the dispersion of nerve fibers within the ONL, indicating that multisynaptic pathways must also exist at the glomerular region. All types of interneurons also responded to odorant stimulation, showing a variety of responses. 3. Granule cells responded to electrical stimulation of the LOT and ONL as reported previously. However, intracellular potential, excitability, and conductance analysis suggested that the mitral cell-mediated excitatory postsynaptic potential (EPSP) is followed by a long inhibitory postsynaptic potential (IPSP). An early negative potential, before the EPSP, was also observed in every granule cell and correlated with component I of the extracellular LOT-induced field potential. We have interpreted this negativity as a "field effect," that may be diagnostic of granule cells. 4. Most granule cells exhibited excitatory responses to odorant stimulation. Odors could produce spiking responses that were either nonhabituating (response to every sniff) or rapidly habituating (response to first sniff only). Other granule cells, while spiking to electrical stimulation, showed depolarizations that did not evoke spikes to odor stimulation. These depolarizations were transient with each sniff or sustained across a series of sniffs. These physiological differences to odor stimulation correlated with granule cell position beneath the mitral cell layer for 12 cells, suggesting that morphological subtypes of granule cells may show physiological differences. Some features of the granule cell odor responses seem to correlate with some of the features we have observed in mitral/tufted cell intracellular recordings. Only one cell showed inhibition to odors. 5. Periglomerular (PG) cells showed a response to ONL stimulation that was unlike that found in other olfactory bulb neurons. There was a long-duration hyperpolarization after a spike and large depolarization or burst of spikes (20-30 ms in duration). Odor stimulation produced simple bursts of action potentials, Odor stimulation produced simple bursts of action potentials, suggesting that PG cells may simply follow input from the olfactory nerve.(ABSTRACT TRUNCATED AT 400 WORDS)

Extracellular recordings were made from mitral cells, tufted cells, and presumed glomerular layer and external plexiform layer interneurons of the olfactory bulb of anesthetized rats during odor stimulation. Intensity responses of these cells were studied by presenting a series of six or seven concentrations, spanning a range greater than two log units, in a cyclic artificial sniff paradigm, which produced repeated response measures at each concentration. Experiments focused on obtaining a complete intensity series, including interspersed unstimulated spontaneous activity records, for a single odorant (usually amyl acetate), but concentration responses to other odorants were tested when possible. Odor responses of 46 cells were studied with two approaches. Response form was examined in an attempt to define response classes based on qualitative characteristics of the temporal pattern of response. Assessment of response magnitude was attempted, in order to construct stimulus-response functions for each cell, independent of response form. As previously reported for olfactory bulb cells, the cells in our sample responded to odor stimulation with spike trains of a variety of temporal patterns, consisting of excitatory and inhibitory components that were frequently recognizable in the responses of a cell across a range of concentrations. However, response patterns usually changed significantly with concentration, such that response form across the concentration range could not be predicted from the response at any one concentration. Responses of different cells were sometimes similar to each other in form at one concentration and quite different from each other in the rest of their concentration-response profiles. Classification of response profiles into discrete types, based on consistency of response form throughout the profile, was therefore not feasible. In agreement with other reports, response of a single cell to different odorants sometimes showed similar forms and sometimes showed very different forms across the concentration-response profiles. Since the response form depends on the stimulus intensity as well as the stimulus quality, characterization of response magnitude and of the pattern of response to different odors require testing with a series of stimulus concentrations. Because odor responses consisted of temporally patterned spike trains, whose components changed in complex ways with stimulus intensity, it was not possible to quantify response magnitude by measuring characteristics of particular response components or counting mean frequency.(ABSTRACT TRUNCATED AT 400 WORDS)

1. Intracellular and extracellular recordings were made from rat olfactory bulb mitral and tufted cells during odor stimulation and during electrical stimulation of the olfactory nerve. Neurons were identified by horseradish peroxidase injections and/or antidromic activation. The presentation of multiple concentrations of at least one odorant in a cyclic artificial sniff paradigm, as reported previously (10), allowed the study of odor responses. This approach was extended to multiple odorants to compare their concentration-response profiles. This procedure avoids the problems of interpretation resulting from nonequivalence of the effective concentrations of different odorants used as stimuli that have characterized previous studies of odor quality effects. Comparisons of intracellular events and responses to electrical stimulation with the odor-induced spike train activity allow us to begin to delineate the local circuitry involved in generating odor-induced responses. 2. The concentration-response profiles of the 72 cells in the present study are comparable to those previously reported for output neurons of the olfactory bulb, showing ordered changes in the temporal patterning of spike activity with step changes in odor concentration. However, eight of the neurons exhibited inhibitory responses to lower concentrations, but excitation, at similar latency, to higher concentrations of the same odorant. These data emphasize that to study pattern changes induced by changing odor quality the influence of stimulus intensity must also be carefully examined. The data also provide evidence that the temporal pattern evoked by an odorant is probably not in itself the code for odor quality recognition. 3. Complete concentration-response profiles, including subthreshold concentrations, to more than one odorant show that, although responses to the different odorant can evolve systematically with concentration, the responses to different odorants can evolve through very different patterns. For example, in some cells, the response patterns to different odors were complementary in form. These results demonstrate that the patterned responses of olfactory bulb neurons can reflect changes in odor quality as well as intensity. 4. Intracellular recording was employed to compare the temporal patterning of spikes during odor stimulation with membrane potential changes. In some cases, the spike pattern was closely correlated with apparent postsynaptic potentials. However, there were several clear exceptions. In five cells, a prominent hyperpolarization, seen in the first sniff of a series of 10 consecutive sniffs, was associated with pauses in spike activity. In the following