perforated-patch-clamp-recording-mouse-olfactory-sensory-neurons

Perforated Patch-clamp Recording of Mouse Olfactory Sensory Neurons in Intact Neuroepithelium: Functional Analysis of Neurons Expressing an Identified Odorant Receptor

Analyzing the physiological responses of olfactory sensory neurons (OSN) when stimulated with specific ligands is critical to understand the basis of ...

To find a mating partner, moths rely on pheromone communication. Released in very low amounts, female sex pheromones are used by males to identify and localize females. Depending on the physiological state (i.e. age, reproductive state), the olfactory system of the males of the noctuid moth Agrotis ipsilon is 'switched on or off'. To understand the neural basis of this behavioural plasticity, we performed a detailed characterization of the qualitative, quantitative and temporal aspects of pheromone coding in the primary centre of integration of pheromonal information, the macroglomerular complex (MGC) of the antennal lobe. MGC neurons were intracellularly recorded and stained in sexually mature virgin males. When stimulating antennae of males with the three main components of the female pheromone blend, most of the neurons showed a biphasic excitatory-inhibitory response. Although they showed different preferences, 80% of the neurons responded at least to the main pheromone component (Z-7-dodecenyl acetate). Six stained neurons responding to this component had their dendrites in the largest MGC glomerulus. Changes in the stimulus intensity and duration affected the excitatory phase but not the inhibitory phase properties. The stimulus intensity was shown to be encoded in the firing frequency, the number of spikes and the latency of the excitatory phase, whereas the stimulus duration only changed its duration. We conclude that the inhibitory input provided by local interneurons following the excitatory phase might not contribute directly to the encoding of stimulus characteristics. The data presented will serve as a basis for comparison with those of immature and mated males.

Quantitative analysis of sex-pheromone coding in the antennal lobe of the moth Agrotis ipsilon: a tool to study network plasticity.

Male moths use sex pheromones to find their mating partners. In the moth, Agrotis ipsilon, the behavioral response and the neuron sensitivity within the primary olfactory centre, the antennal lobe (AL), to sex pheromone increase with age and juvenile hormone (JH) biosynthesis. By manipulating the JH level, we previously showed that JH controls this age-dependent neuronal plasticity, and that its effects are slow (within 2 days). We hypothesized that the hormonal effect might be indirect, and one neuromodulator candidate, which might serve as a mediator, is octopamine (OA). Here, we studied the effects of OA and an OA receptor antagonist, mianserin, on behavioral and AL neuron responses of mature and immature males during stimulation with sex pheromone. Our results indicate that, although OA injections enhanced the behavioral pheromone response in mature males, OA had no significant effect on behavior in immature males. However, mianserin injections decreased the behavioral response in mature males. AL neuron sensitivity increased after OA treatment in immature males, and decreased after mianserin treatment in mature males. Determination of OA levels in ALs of immature and mature males did not reveal any difference. To study the possible interactive effects of JH and OA, the behavioral pheromone response was analyzed in JH-deprived mature males injected with OA, and in immature males injected with fenoxycarb, a JH agonist, and mianserin. Results show that both JH and OA are necessary to elicit a behavioral response of A. ipsilon males to sex pheromone.

Age-dependent plasticity of sex pheromone response in the moth, Agrotis ipsilon: combined effects of octopamine and juvenile hormone.

In the male moth, Agrotis ipsilon, mating induces a transient inhibition of behavioural and central nervous responses to sex pheromone. Newly mated males are not attracted to sex pheromone, and the sensitivity of their antennal lobe (AL) neurons is lower than in virgin males. This rapid transient olfactory inhibition prevents them from re-mating unsuccessfully until they have refilled their sex glands. We hypothesized that this olfactory 'switch off' might be controlled by neuromodulators such as biogenic amines. To test our hypothesis, we studied the effects of octopamine (OA) and serotonin (5-hydroxytryptamine, 5-HT) on the coding properties of pheromone-sensitive AL neurons in virgin and newly mated males. We show that AL neuron sensitivity increased in newly mated males after injection of OA or 5-HT, but only OA treatment affected certain response characteristics of AL neurons in virgin males. Whereas all measured AL neuron response characteristics were different between virgin and newly mated males, amine treatment in newly mated males restored only the latency and spike frequency, but not the duration of excitatory and inhibitory phases, which were initially found in virgin males. Additionally, we investigated the behavioural effects of OA and 5-HT treatments in virgin and mated males. Although OA and 5-HT enhanced the general flight activity of newly mated males, amine treatments did not restore the behavioural pheromone response of mated moths. Altogether, these results show that, although biogenic amines modulate the olfactory system in moths, OA and 5-HT are probably not involved in the post-mating inhibition of responses to sex pheromone in A. ipsilon males.

Mating-induced transient inhibition of responses to sex pheromone in a male moth is not mediated by octopamine or serotonin.

How information is transformed along synaptic processing stages is critically important to understand the neural basis of behavior in any sensory system. In moths, males rely on sex pheromone to find their mating partner. It is essential for a male to recognize the components present in a pheromone blend, their ratio, and the temporal pattern of the signal. To examine pheromone processing mechanisms at different levels of the olfactory pathway, we performed single-cell recordings of olfactory receptor neurons (ORNs) in the antenna and intracellular recordings of central neurons in the macroglomerular complex (MGC) of the antennal lobe of sexually mature Agrotis ipsilon male moths, using the same pheromone stimuli, stimulation protocol, and response analyses. Detailed characteristics of the ORN and MGC-neuron responses were compared to describe the transformation of the neuronal responses that takes place in the MGC. Although the excitatory period of the response is similar in both neuron populations, the addition of an inhibitory phase following the MGC neuron excitatory phase indicates participation of local interneurons (LN), which remodel the ORN input. Moreover, MGC neurons showed a wider tuning and a higher sensitivity to single pheromone components than ORNs.

Transformation of the sex pheromone signal in the noctuid moth Agrotis ipsilon: from peripheral input to antennal lobe output.

Innate behaviours in animals can be influenced by several factors, such as the environment, experience, or physiological status. This behavioural plasticity originates from changes in the underlying neuronal substrate. A well-described form of plasticity is induced by mating. In both vertebrates and invertebrates, males experience a post-ejaculatory refractory period, during which they avoid new females. In the male moth Agrotis ipsilon, mating induces a transient inhibition of responses to the female-produced sex pheromone. To understand the neural bases of this inhibition and its possible odour specificity, we carried out a detailed analysis of the response characteristics of the different neuron types from the periphery to the central level. We examined the response patterns of pheromone-sensitive and plant volatile-sensitive neurons in virgin and mated male moths. By using intracellular recordings, we showed that mating changes the response characteristics of pheromone-sensitive antennal lobe (AL) neurons, and thus decreases their sensitivity to sex pheromone. Individual olfactory receptor neuron (ORN) recordings and calcium imaging experiments indicated that pheromone sensory input remains constant. On the other hand, calcium responses to non-pheromonal odours (plant volatiles) increased after mating, as reflected by increased firing frequencies of plant-sensitive AL neurons, although ORN responses to heptanal remained unchanged. We suggest that differential processing of pheromone and plant odours allows mated males to transiently block their central pheromone detection system, and increase non-pheromonal odour detection in order to efficiently locate food sources.

Mating-induced differential coding of plant odour and sex pheromone in a male moth.

In the olfactory system of male moths, a specialized subset of neurons detects and processes the main component of the sex pheromone emitted by females. It is composed of several thousand first-order olfactory receptor neurons (ORNs), all expressing the same pheromone receptor, that contact synaptically a few tens of second-order projection neurons (PNs) within a single restricted brain area. The functional simplicity of this system makes it a favorable model for studying the factors that contribute to its exquisite sensitivity and speed. Sensory information--primarily the identity and intensity of the stimulus--is encoded as the firing rate of the action potentials, and possibly as the latency of the neuron response. We found that over all their dynamic range, PNs respond with a shorter latency and a higher firing rate than most ORNs. Modelling showed that the increased sensitivity of PNs can be explained by the ORN-to-PN convergent architecture alone, whereas their faster response also requires cell-to-cell heterogeneity of the ORN population. So, far from being detrimental to signal detection, the ORN heterogeneity is exploited by PNs, and results in two different schemes of population coding based either on the response of a few extreme neurons (latency) or on the average response of many (firing rate). Moreover, ORN-to-PN transformations are linear for latency and nonlinear for firing rate, suggesting that latency could be involved in concentration-invariant coding of the pheromone blend and that sensitivity at low concentrations is achieved at the expense of precise encoding at high concentrations.

Heterogeneity and convergence of olfactory first-order neurons account for the high speed and sensitivity of second-order neurons.

Behavioral strategies employed for chemotaxis have been described across phyla, but the sensorimotor basis of this phenomenon has seldom been studied in naturalistic contexts. Here, we examine how signals experienced during free olfactory behaviors are processed by first-order olfactory sensory neurons (OSNs) of the Drosophila larva. We find that OSNs can act as differentiators that transiently normalize stimulus intensity-a property potentially derived from a combination of integral feedback and feed-forward regulation of olfactory transduction. In olfactory virtual reality experiments, we report that high activity levels of the OSN suppress turning, whereas low activity levels facilitate turning. Using a generalized linear model, we explain how peripheral encoding of olfactory stimuli modulates the probability of switching from a run to a turn. Our work clarifies the link between computations carried out at the sensory periphery and action selection underlying navigation in odor gradients.

Article	Year
Quantitative analysis of sex-pheromone coding in the antennal lobe of the moth Agrotis ipsilon: a tool to study network plasticity. The Journal of experimental biology\| PubMed ID: 19329752	2009
Age-dependent plasticity of sex pheromone response in the moth, Agrotis ipsilon: combined effects of octopamine and juvenile hormone. Hormones and behavior\| PubMed ID: 19409391	2009
Mating-induced transient inhibition of responses to sex pheromone in a male moth is not mediated by octopamine or serotonin. The Journal of experimental biology\| PubMed ID: 20228346	2010
Transformation of the sex pheromone signal in the noctuid moth Agrotis ipsilon: from peripheral input to antennal lobe output. Chemical senses\| PubMed ID: 20601375	2010
Mating-induced differential coding of plant odour and sex pheromone in a male moth. The European journal of neuroscience\| PubMed ID: 21488987	2011
Heterogeneity and convergence of olfactory first-order neurons account for the high speed and sensitivity of second-order neurons. PLoS computational biology\| PubMed ID: 25474026	2014
Dynamical feature extraction at the sensory periphery guides chemotaxis. eLife\| PubMed ID: 26077825	2015

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