Taste Hairs Research Articles

Base Recording: A Technique for Analyzing Responses of Taste Neurons in Drosophila.

Insects taste the external world through taste hairs, or sensilla, that have pores at their tips. When a sensillum comes into contact with a potential food source, compounds from the food source enter through the pore and activate neurons within. For over 50 years, these responses have been recorded using a technique called tip recording. However, this method has major limitations, including the inability to measure neural activity before or after stimulus contact and the requirement for tastants to be soluble in aqueous solutions. We describe here a technique that we call base recording, which overcomes these limitations. Base recording allows the measurement of taste neuron activity before, during, and after the stimulus. Thus, it allows extensive analysis of OFF responses that occur after a taste stimulus. It can be used to study hydrophobic compounds such as long-chain pheromones that have very low solubility in water. In summary, base recording offers the advantages of single-sensillum electrophysiology as a means of measuring neuronal activity - high spatial and temporal resolution, without the need for genetic tools - and overcomes key limitations of the traditional tip recording technique.

The Basis of Food Texture Sensation in Drosophila

Food texture has enormous effects on food preferences. However, the mechanosensory cells and key molecules responsible for sensing the physical properties of food are unknown. Here, we show that akin to mammals, the fruit fly, Drosophila melanogaster, prefers food with a specific hardness or viscosity. This food texture discrimination depends upon a previously unknown multidendritic (md-L) neuron, which extends elaborate dendritic arbors innervating the bases of taste hairs. The md-L neurons exhibit directional selectivity in response to mechanical stimuli. Moreover, these neurons orchestrate different feeding behaviors depending on the magnitude of the stimulus. We demonstrate that the single Drosophila transmembrane channel-like (TMC) protein is expressed in md-L neurons, where it is required for sensing two key textural features of food-hardness and viscosity. We propose that md-L neurons are long sought after mechanoreceptor cells through which food mechanics are perceived and encoded by a taste organ, and that this sensation depends on TMC. VIDEO ABSTRACT.

Drosophila Pheromone-Sensing Neurons Expressing the ppk25 Ion Channel Subunit Stimulate Male Courtship and Female Receptivity

As in many species, gustatory pheromones regulate the mating behavior of Drosophila. Recently, several ppk genes, encoding ion channel subunits of the DEG/ENaC family, have been implicated in this process, leading to the identification of gustatory neurons that detect specific pheromones. In a subset of taste hairs on the legs of Drosophila, there are two ppk23-expressing, pheromone-sensing neurons with complementary response profiles; one neuron detects female pheromones that stimulate male courtship, the other detects male pheromones that inhibit male-male courtship. In contrast to ppk23, ppk25, is only expressed in a single gustatory neuron per taste hair, and males with impaired ppk25 function court females at reduced rates but do not display abnormal courtship of other males. These findings raised the possibility that ppk25 expression defines a subset of pheromone-sensing neurons. Here we show that ppk25 is expressed and functions in neurons that detect female-specific pheromones and mediates their stimulatory effect on male courtship. Furthermore, the role of ppk25 and ppk25-expressing neurons is not restricted to responses to female-specific pheromones. ppk25 is also required in the same subset of neurons for stimulation of male courtship by young males, males of the Tai2 strain, and by synthetic 7-pentacosene (7-P), a hydrocarbon normally found at low levels in both males and females. Finally, we unexpectedly find that, in females, ppk25 and ppk25-expressing cells regulate receptivity to mating. In the absence of the third antennal segment, which has both olfactory and auditory functions, mutations in ppk25 or silencing of ppk25-expressing neurons block female receptivity to males. Together these results indicate that ppk25 identifies a functionally specialized subset of pheromone-sensing neurons. While ppk25 neurons are required for the responses to multiple pheromones, in both males and females these neurons are specifically involved in stimulating courtship and mating.

ADrosophilaDEG/ENaC Subunit Functions Specifically in Gustatory Neurons Required for Male Courtship Behavior

Detection of specific female pheromones stimulates courtship behavior in Drosophila melanogaster males, but the chemosensory molecules, cells, and mechanisms involved remain poorly understood. Here we show that ppk25, a DEG/ENaC ion channel subunit required for normal male response to females, is expressed at highest levels in a single sexually dimorphic gustatory neuron of most taste hairs on legs and wings, but not in neurons that detect courtship-inhibiting pheromones or food. Synaptic inactivation of ppk25-expressing neurons, or knockdown of ppk25 expression in all gustatory neurons, significantly impairs male response to females, whereas gustatory expression of ppk25 rescues the courtship behavior of ppk25 mutant males. Remarkably, the only other detectable albeit significantly weaker expression of ppk25 occurs in olfactory neurons implicated in modulation of courtship behavior. However, expression of ppk25 in olfactory neurons is not required for male courtship under our experimental conditions. These data show that ppk25 functions specifically in peripheral taste neurons involved in activation of courtship behavior, an unexpected function for this type of channel. Furthermore, our work identifies a small subset of gustatory neurons with an essential role in activation of male courtship behavior, most likely in response to female pheromones.

The Molecular and Cellular Basis of Bitter Taste in Drosophila

The extent of diversity among bitter-sensing neurons is a fundamental issue in the field of taste. Data are limited and conflicting as to whether bitter neurons are broadly tuned and uniform, resulting in indiscriminate avoidance of bitter stimuli, or diverse, allowing a more discerning evaluation of food sources. We provide a systematic analysis of how bitter taste is encoded by the major taste organ of the Drosophila head, the labellum. Each of 16 bitter compounds is tested physiologically against all 31 taste hairs, revealing responses that are diverse in magnitude and dynamics. Four functional classes of bitter neurons are defined. Four corresponding classes are defined through expression analysis of all 68 gustatory taste receptors. A receptor-to-neuron-to-tastant map is constructed. Misexpression of one receptor confers bitter responses as predicted by the map. These results reveal a degree of complexity that greatly expands the capacity of the system to encode bitter taste.

Fixation conditions affect the immunoreactivity of gustducin in rat vallate taste buds

This study aimed to examine gustducin immunoreactivities when adopting various immunostaining conditions in rat vallate taste buds. The occurrence and intensity of the immunoreactivities exhibited specific patterns in accordance with the fixation time. The immunoreactions were localized to only taste hairs, the upper part of the taste bud, after short fixation periods but then to the cell-body cytoplasm excluding the taste hairs after long fixation periods. These immunohistochemical data suggest that the staining protocols, especially the fixation time, cause discrepancies in gustducin immunoreactivities.

A Drosophila Protein Family Implicated in Pheromone Perception Is Related to Tay-Sachs GM2-Activator Protein

Low volatility, lipid-like cuticular hydrocarbon pheromones produced by Drosophila melanogaster females play an essential role in triggering and modulating mating behavior, but the chemosensory mechanisms involved remain poorly understood. Recently, we showed that the CheB42a protein, which is expressed in only 10 pheromone-sensing taste hairs on the front legs of males, modulates progression to late stages of male courtship behavior in response to female-specific cuticular hydrocarbons. Here we report that expression of all 12 genes in the CheB gene family is predominantly or exclusively gustatory-specific, and occurs in many different, often non-overlapping patterns. Only the Gr family of gustatory receptor genes displays a comparable variety of gustatory-specific expression patterns. Unlike Grs, however, expression of all but one CheB gene is sexually dimorphic. Like CheB42a, other CheBs may therefore function specifically in gustatory perception of pheromones. We also show that CheBs belong to the ML superfamily of lipid-binding proteins, and are most similar to human GM2-activator protein (GM2-AP). In particular, GM2-AP residues involved in ligand binding are conserved in CheBs but not in other ML proteins. Finally, CheB42a is specifically secreted into the inner lumen of pheromone-sensing taste hairs, where pheromones interact with membrane-bound receptors. We propose that CheB proteins interact directly with lipid-like Drosophila pheromones and modulate their detection by the gustatory signal transduction machinery. Furthermore, as loss of GM2-AP in Tay-Sachs disease prevents degradation of GM2 gangliosides and results in neurodegeneration, the function of CheBs in pheromone response may involve biochemical mechanisms critical for lipid metabolism in human neurons.

Two Gr Genes Underlie Sugar Reception in Drosophila

We have analyzed the molecular basis of sugar reception in Drosophila. We define the response spectrum, concentration dependence, and temporal dynamics of sugar-sensing neurons. Using in situ hybridization and reporter gene expression, we identify members of the Gr5a-related taste receptor subfamily that are coexpressed in sugar neurons. Neurons expressing reporters of different Gr5a-related genes send overlapping but distinct projections to the brain and thoracic ganglia. Genetic analysis of receptor genes shows that Gr5a is required for response to one subset of sugars and Gr64a for response to a complementary subset. A Gr5a;Gr64a double mutant shows no physiological or behavioral responses to any tested sugar. The simplest interpretation of our results is that Gr5a and Gr64a are each capable of functioning independently of each other within individual sugar neurons and that they are the primary receptors used in the labellum to detect sugars.

Central gustatory projections and side-specificity of operant antennal muscle conditioning in the honeybee

Gustatory stimuli to the antennae, especially sucrose, are important for bees and are employed in learning paradigms as unconditioned stimulus. The present study identified primary antennal gustatory projections in the bee brain and determined the impact of stimulation of the antennal tip on antennal muscle activity and its plasticity. Central projections of antennal taste hairs contained axons of two morphologies projecting into the dorsal lobe, which is also the antennal motor centre. Putative mechanosensory axons arborised in a dorso-lateral area. Putative gustatory axons projected to a ventro-medial area. Bees scan gustatory and mechanical stimuli with their antennae using variable strategies but sensory input to the motor system has not been investigated in detail. Mechanical, gustatory, and electrical stimulation of the ipsilateral antennal tip were found to evoke short-latency responses in an antennal muscle, the fast flagellum flexor. Contralateral gustatory stimulation induced smaller responses with longer latency. The activity of the fast flagellum flexor was conditioned operantly by pairing high muscle activity with ipsilateral antennal sucrose stimulation. A proboscis reward was unnecessary for learning. With contralateral antennal sucrose stimulation, conditioning was unsuccessful. Thus, muscle activity induced by gustatory stimulation was important for learning success and conditioning was side-specific.

Behavioral and electrophysiological investigation on taste response of the termite Zootermopsis nevadensis to wood extractives

Termite feeding behavior and the chemoreception of plant extracts were evaluated to investigate the water extracts from akamatsu (Pinus densiflora), neem (Azadirachta indica), and their equivalent mixture using pseudoergates of Zootermopsis nevadensis. In behavioral assays, termite preference was akamatsu > akamatsu plus neem > neem. Electrophysiological recordings from the taste hairs on labial palps showed vigorous impulse discharge to akamatsu extract but much lower response to neem extract. The response to akamatsu plus neem was mostly the same as that to neem alone, suggesting the neem extracts inhibited the responses to akamatsu extracts. In the present article, we discuss the correlations between the feeding behaviors and the responses at their taste cells to these different extracts.

Habituation and dishabituation of exploratory and appetitive responses in the honey bee ( Apis mellifera L.)

Sucrose-induced antennal and proboscis motor responses of the honey bee were comparatively studied using muscle responses. The activity of the proboscis muscle responses was correlated with sucrose stimulus concentration, but that of antennal muscle responses was not. Both responses habituated following repetitive stimulation with a low sucrose concentration and dishabituated upon stimulation with a high sucrose concentration. Despite the different response properties of the responses, their habituation kinetics were indistinguishable. The absence of sensory adaptation was confirmed by taste hair recordings. The potential for habituation was related to the functions of the responses studied. The exploratory antennal response used for continuous sampling of the environment habituated to a smaller extent than the appetitive proboscis response.

The functions of antennal mechanoreceptors and antennal joints in tactile discrimination of the honeybee (Apis mellifera L.)

Honeybees learn and discriminate excellently between different surface structures and different forms of objects, which they scan with their antennae. The sensory plate on the antennal tip plays a key role in the perception of mechanosensory and gustatory information. It is densely covered with small tactile hairs and carries a few large taste hairs. Both types of sensilla contain a mechanoreceptor, which is involved in the antennal scanning of an object. Our experiments test the roles of the mechanoreceptors on the antennal tip in tactile antennal learning and discrimination. Joints between head capsule and scapus and between scapus and pedicellus enable the bee to perform three-dimensional movements when they scan an object. The role of these joints in tactile antennal learning and discrimination is studied in separate experiments. The mechanoreceptors on the antennal tip were decisive for surface discrimination, but not for tactile acquisition or discrimination of shapes. When the scapus-pedicellus joint or the headcapsule-scapus joint was fixed on both antennae, tactile learning was still apparent but surface discrimination was abolished. Fixing both scapi to the head capsule reduced tactile acquisition.

Expression of metabotropic glutamate receptor group I in rat gustatory papillae.

Taste-metabotropic glutamate receptor 4 (taste-mGluR4) and the heteromers of T1R1 and T1R3 are candidate receptors involved in the sense of umami (monosodium glutamate) taste. Although the expression of group III mGluRs (taste-mGluR4) has been demonstrated in taste tissues, no mention has been made of the expression of group I mGluRs (mGluR1 and mGluR5) in taste tissues. We examined the expression of mGluR1 and mGluR5 in rat gustatory tissues by using reverse transcription-polymerase chain reaction (RT-PCR), in situ hybridization, immunohistochemistry and immunoelectron microscopy. RT-PCR assay showed that mGluR1 alpha and mGluR1 beta mRNAs were expressed in circumvallate papillae, but mGluR5 mRNA was not expressed. The positive signals of mGluR1 mRNA were detected only in circumvallate taste buds by in situ hybridization analysis. In cryosections of fungiform, foliate and circumvallate papillae, the antibody against mGluR1 alpha gave intense labeling on the taste hairs in all taste pores examined. In the developing taste buds, the positive signals of mGluR1alpha in taste hairs gradually increased with the increase in number of taste bud cells. These results show that, in addition to taste-mGluR4 and the heteromer of T1R1 and T1R3, mGluR1 alpha may function as a receptor for glutamate (umami) taste sensation.

Responsiveness of antennal taste hairs of the apterygotan insect, Thermobia domestica (Zygentoma); an electrophysiological investigation

Antennal taste hairs of Thermobia domestica were investigated by a modified tip recording technique. They are sensitive to aqueous salt solutions but do not show the characteristic features of a water and a salt receptor cell. Sucrose and fructose stimulated approximately half of the tested hairs. As revealed by spike amplitude histograms, in most cases two sense cells responded. Proline stimulated the hairs with a threshold concentration of about 2 mM. Again, in most cases, two sense cells responded. In relation to sucrose the major sensitivities to both fructose and proline were not found in clearly defined receptor cells as in flies, but were realized in different cells of either a higher or smaller spike amplitude. This suggests, that the number of specific binding sites varies among the receptor cells from one taste hair to the other. As in cockroaches, but unlike flies, spike series are mainly generated in the apical dendritic segments. It is suggested that these features of the sense cells in thermobian taste hairs represent a primitive condition in relation to the specification of taste hairs cells in higher insect taxa

Transduction ion channels directly gated by sugars on the insect taste cell.

Insects detect sugars and amino acids by a specialized taste cell, the sugar receptor cell, in the taste hairs located on their labela and tarsi. We patch-clamped sensory processes of taste cells regenerated from the cut end of the taste hairs on the labelum of the flashfly isolated from the pupa approximately 20 h before emergence. We recorded both single channel and ensemble currents of novel ion channels located on the distal membrane of the sensory process of the sugar receptor cell. In the stable outside-out patch membrane excised from the sensory processes, we could repeatedly record sucrose-induced currents for tens of minutes without appreciable decrease. An inhibitor of G-protein activation, GDP-beta-S, did not significantly decrease the sucrose response. These results strongly suggested that the channel is an ionotropic receptor (a receptor/channel complex), activated directly by sucrose without mediation by second messengers or G protein. The channel was shown to be a nonselective cation channel. Analyses of single channel currents showed that the sucrose-gated channel has a single channel conductance of approximately 30 pS and has a very short mean open time of approximately 0.23 ms. It is inhibited by external Ca(2+) and the dose-current amplitude relation could be described by a Michaelis-Menten curve with an apparent dissociation constant of approximately 270 mM. We also report transduction ion channels of the receptor/channel complex type directly gated by fructose and those gated by L-valine located on the sensory process.

Chemosensory control of pollen ingestion in the hoverfly Eristalis tenax by labellar taste hairs.

The labellar gustatory system of the dronefly Eristalis tenax L. (Syrphidae; Diptera) that enables the fly to discriminate between pollen and nectar is investigated, and the triggering of pollen ingestion is examined. In behavioural preference tests, exhaustively extracted pollen of the sunflower Helianthus annuus is consumed in smaller amounts than untreated pollen, indicating that water-soluble substances are important for acceptance. Dry pollen is preferred over moist pollen in which the grains stick together, suggesting that mechanical properties of the pollen also play a role in its sensory assessment. Electrophysiological studies of the labellar taste hairs reveal that aqueous extracts of pollen (2% w/v) stimulate the salt receptor cell, but not the sugar receptor cell. The response of the water receptor cell remains the same as to pure water (or standard electrolyte, 10 mmol.l-1 KCl). Of the 20 amino acids tested, the salt cell is sensitive only to proline in a submillimolar range. Behavioural experiments support the electrophysiological findings. When KCl is applied at concentrations eliciting salt-cell spike frequencies equal to those produced by pollen extract (which is often accepted), the water receptor cell is inhibited and a pronounced rejection behaviour occurs. This rejection of concentrated salt solution in Eristalis is therefore mainly mediated by the inhibition of the water cell.

New aspects of chemoreception in flies.

For studies of insect taste the sensory cells of labellar taste hairs on the proboscis of calliphorid flies (e.g., Phormia) are preferred objects. Three sensory cells are electrophysiologically characterized: a sugar cell, a water cell and a cation or salt cell. Studying hairs on legs and proboscis of other families of flies we recently obtained results that extend our knowledge about the complexity of food detection in flies. The hoverfly Eristalis tenax (Syrphidae family) feeds on nectar and pollen of flowers. While nectar is recognized via the sugar cell, the so-called salt cell is activated by low concentrations of a water-soluble substance in extracts of pollen. From several tested pollen constituents only proline, an essential amino acid for pollen germination, stimulates the salt cell. The discrimination between salt and pollen in behavioral tests can be explained by the finding that the water cell remains active in the presence of pollen extract, but is inhibited by salts. Obviously the water cell activity is involved in the feeding decision. We already described a similar situation in Phormia, where the salt cell activity only elicits feeding behavior when the water cell is active too. In contrast to the sugar cells of species of flies studied so far the sugar cell of the house fly Musca domestica (Muscidae family) is highly sensitive to lactose, a disaccharide (beta-galactoside (1-->4) glucoside) naturally occurring only in the milk of mammals. Thus sugar spectra adapt to special environmental food situations. The fifth tarsomere of the forelegs of Musca bears besides two D-hairs 46 hairs of the B-type. The B-hairs are functionally not uniform in contrast to their structural identity. Only two hairs, the 'water hairs,' contain a water cell beside a spontaneously firing cell, but no sugar cell. Other examples of diversity are hairs that contain apart from the classical sugar cells, additional cells which react either to p-nitrophenyl-beta-galactoside, but not to the chemically related lactose, or to p-nitrophenyl-alpha-glucoside. Therefore the rule established for the labellar taste hairs that each hair type contains the same physiological equipment of sensory cells, does not hold for the taste hairs of the legs.

The interaction of taste and heat on the biting response of the tsetse fly Glossina fuscipes fuscipes

Abstract.Tsetse flies probe more on a heated surface with a trace of uric acid than on one without. Uric acid is one of the components of human sweat and it elicits spike responses from taste hairs on the flies' legs. In this paper it is examined how heat from the surface and taste interact to affect the biting behaviour of Glossina fuscipes fuscipes Newstead 1910 (Diptera: Glossinidae) over successive days of food deprivation. The biting behaviour consists of bouts of probing, both ambulatory and stationary, intercalated with short hops of flight. The number of bouts increases over successive days, whereas the average bout duration does not. Although uric acid alone could not induce the flies to probe, in combination with surface heat it affected the flies greatly. Average bout duration was two‐fold that on a heated surface not treated with uric acid. The frequency of bouts was not affected by uric acid. These experiments and auxiliary ones on mechanoreceptive input and odours lead to the insight that the factors which affect biting behaviour can be viewed as a hierarchy. The hierarchy extends from those that affect the onset of biting to those that affect its course.

Ingestion of sugar diets correlates with spike activity in labellar chemosensilla of the flesh‐fly,Neobellieria(=Sarcophaga)bullata

Abstract.Female 2‐day‐old Neobellieria (= Sarcophaga) bullata (Parker) (Diptera: Sarcophagidae) were exposed to different concentrations of sucrose, glucose and fructose in a single‐choice potometer, and the volume ingested in the first hour was measured. Nerve spike activity in response to the same sugars was recorded from medium labellar taste hairs of similar flies by tip‐recording. Two classes of chemosensory cells responded to sucrose, glucose and fructose. Cell 1 showed an increasing spike activity with sugar concentration, whereas cell 2 did not; cell 1 was identified as the ‘sugar cell'.For both spike activity in cell 1 and feeding, sucrose was the most stimulatory sugar. The dose–response curves for glucose and fructose crossed over at about 200 mm. At higher concentrations, glucose was more stimulatory for both cell 1 and for feeding, and at lower concentrations, fructose. The pattern of spike activity supports a separate location on the sensory cells of receptors for pyranose and fructose forms of sugar. The strong correlation between volume ingested and spike activity indicates that sugar feeding is controlled by sensory input from the ‘sugar' cells of labellar chemosensilla. Moreover, the results suggest that the flies do not distinguish between these sugars except by apparent ‘sweetness'.

Development of specific responses in antennal taste hairs after ecdysis. An electrophysiological investigation of the cockroach, Periplaneta brunnea

Spike generation in cockroach antennal taste hairs of nymphs appears to follow the same rules as in adults analysed previously. After ecdysis, taste hairs on the proximal segments responded earlier than those on distal ones. Shorter hairs became active before longer ones. The mechanoreceptor responded before the chemoreceptors. During a transient period, 3–5 hours after ecdysis, all four chemoreceptors and also the mechanoreceptor were sensitive to 150 mM KCl. Spikes elicited by 75 mM KCl were generated predominantly in the outer dendritic segments. They did not differ in shape and frequency when 60 mM sucrose was added. The earliest sign of a specific reaction to sucrose was observed approximately 4–5 hours after ecdysis, when the mixture of sucrose and KCl elicited more spikes than KCl alone.