What are olfaction and gustation, and do all animals have them?

Different animals have distinctive anatomical and physiological properties to their chemical senses that enhance detection and discrimination of relevant chemical cues. Humans and other vertebrates are recognized as having 2 main chemical senses, olfaction and gustation, distinguished from each other by their evolutionarily conserved neuroanatomical organization. This distinction between olfaction and gustation in vertebrates is not based on the medium in which they live because the most ancestral and numerous vertebrates, the fishes, live in an aquatic habitat and thus both olfaction and gustation occur in water and both can be of high sensitivity. The terms olfaction and gustation have also often been applied to the invertebrates, though not based on homology. Consequently, any similarities between olfaction and gustation in the vertebrates and invertebrates have resulted from convergent adaptations or shared constraints during evolution. The untidiness of assigning olfaction and gustation to invertebrates has led some to recommend abandoning the use of these terms and instead unifying them and others into a single category-chemical sense. In our essay, we compare the nature of the chemical senses of diverse animal types and consider their designation as olfaction, oral gustation, extra-oral gustation, or simply chemoreception. Properties that we have found useful in categorizing chemical senses of vertebrates and invertebrates include the nature of peripheral sensory cells, organization of the neuropil in the processing centers, molecular receptor specificity, and function.

Understanding responses to chemical mixtures: looking forward from the past.

Our goal in this article is to provide a perspective on how to understand the nature of responses to chemical mixtures. In studying responses to mixtures, researchers often identify "mixture interactions"-responses to mixtures that are not accurately predicted from the responses to the mixture's individual components. Critical in these studies is how to predict responses to mixtures and thus to identify a mixture interaction. We explore this issue with a focus on olfaction and on the first level of neural processing-olfactory sensory neurons-although we use examples from taste systems as well and we consider responses beyond sensory neurons, including behavior and psychophysics. We provide a broadly comparative perspective that includes examples from vertebrates and invertebrates, from genetic and nongenetic animal models, and from literature old and new. In the end, we attempt to recommend how to approach these problems, including possible future research directions.

Reproductive and metabolic state differences in olfactory responses to amino acids in a mouth brooding African cichlid fish.

Olfaction mediates many crucial life-history behaviors such as prey detection, predator avoidance, migration and reproduction. Olfactory function can also be modulated by an animal's internal physiological and metabolic states. While this is relatively well studied in mammals, little is known about how internal state impacts olfaction in fishes, the largest and most diverse group of vertebrates. Here we apply electro-olfactograms (EOGs) in the African cichlid fish Astatotilapia burtoni to test the hypothesis that olfactory responses to food-related cues (i.e. l-amino acids; alanine and arginine) vary with metabolic, social and reproductive state. Dominant males (reproductively active, reduced feeding) had greater EOG magnitudes in response to amino acids at the same tested concentration than subordinate males (reproductively suppressed, greater feeding and growth rates). Mouth brooding females, which are in a period of starvation while they brood fry in their mouths, had greater EOG magnitudes in response to amino acids at the same tested concentration than both recovering and gravid females that are feeding. Discriminant function analysis on EOG magnitudes also grouped the male (subordinate) and female (recovering, gravid) phenotypes with higher food intake together and distinguished them from brooding females and dominant males. The slope of the initial negative phase of the EOG also showed intra-sexual differences in both sexes. Our results demonstrate that the relationship between olfaction and metabolic state observed in other taxa is conserved to fishes. For the first time, we provide evidence for intra-sexual plasticity in the olfactory response to amino acids that is influenced by fish reproductive, social and metabolic state.

Amino acid specificity of fibers of the facial/trigeminal complex innervating the maxillary barbel in the Japanese sea catfish, Plotosus japonicus.

The Japanese sea catfish, Plotosus japonicus, possesses taste and solitary chemoreceptor cells (SCCs) located on the external body surface that detect specific water-soluble substances. Here, we identify two major fiber types of the facial/trigeminal complex that transmit amino acid information to the medulla. Both single and few fiber preparations respond to amino acid stimulation in the 0.1 µM to mM range. One fiber type responds best to glycine and l-alanine (i.e. Gly/Ala fibers) whereas the other fiber type is best stimulated by l-proline and glycine betaine (hereafter referred to only as betaine) (i.e. Pro/Bet fibers). We demonstrate that betaine, which does not alter the pH of the seawater and therefore does not activate the animals' highly sensitive pH sensors (Caprio et al., Science 344:1154-1156, 2014), is sufficient to elicit appetitive food search behavior. We further show that the amino acid specificity of fibers of the facial/trigeminal complex in P. japonicus is different from that in Ariopsis felis (Michel and Caprio, J. Neurophysiol. 66:247-260, 1991; Michel et al., J. Comp. Physiol. A. 172:129-138, 1993), a representative member of the only other family (Ariidae) of extant marine catfishes.

Anatomical and physiological studies of bigheaded carps demonstrate that the epibranchial organ functions as a pharyngeal taste organ.

The epibranchial organ (EO) is an enigmatic tubular organ found in the pharyngeal cavity of many filter-feeding fishes. We investigated whether it might function as a taste organ that mediates aggregation and ingestion of planktonic food within the buccal cavity. The EO and associated structures of bighead and silver carps, two successful and invasive planktivorous fishes, were examined using histological and electrophysiological techniques. Both species possess finely structured gill rakers that extend directly via a series of protrusions into each of the four blind canals which are organized as the muscular EO, suggesting that the gill rakers and EO probably function in an integrated manner. Both the interior and exterior surfaces of the EOs of both species are covered with high densities of taste buds and solitary chemosensory cells (SCCs) as well as mucous cells. Conversely, taste buds are scarce in both the buccal cavities and external portions of the head and mouth of both species. Electrophysiological recordings from a caudal branch of the vagus nerve (cranial nerve X) found to innervate the EO showed it to be sensitive to chemicals found in a planktonic diet. l-Amino acids accounted for some, but not all of the neural activity. We conclude that taste buds and SCCs located on the EO and gill rakers probably serve to chemically detect food particles, which the EO then aggregates by mucus secretion before eventually expelling them onto the floor of the pharynx for ingestion. This specialized, pharyngeal chemosensory structure may explain the feeding success of these, and perhaps other planktivorous, filter-feeding fishes.

Marine teleost locates live prey through pH sensing.

We report that the Japanese sea catfish Plotosus japonicus senses local pH-associated increases in H(+)/CO2 equating to a decrease of ≤0.1 pH unit in ambient seawater. We demonstrated that these sensors, located on the external body of the fish, detect undamaged cryptic respiring prey, such as polychaete worms. Sensitivity is maximal at the natural pH of seawater (pH 8.1 to 8.2) and decreases dramatically in seawater with a pH <8.0.

Sensitivity and specificity of the olfactory epithelia of two elasmobranch species to bile salts.

Odor detection in vertebrates occurs when odorants enter the nose and bind to molecular olfactory receptors on the cilia or microvilli of olfactory receptor neurons (ORNs). Several vertebrate groups possess multiple, morphologically distinct types of ORNs. In teleost fishes, these different ORN types detect specific classes of biologically relevant odorants, such as amino acids, nucleotides and bile salts. For example, bile salts are reported to be detected exclusively by ciliated ORNs. The olfactory epithelium of elasmobranch fishes (sharks, rays and skates) is comprised of microvillous and crypt ORNs, but lacks ciliated ORNs; thus, it was questioned whether the olfactory system of this group of fishes is capable of detecting bile salts. The present investigation clearly indicates that the olfactory system of representative shark and stingray species does detect and respond to bile salts. Additionally, these species detect glycine-conjugated, taurine-conjugated and non-conjugated bile salts, as do teleosts. These elasmobranchs are less sensitive to the tested bile salts than reported for both agnathans and teleosts, but this may be due to the particular bile salts selected in this study, as elasmobranch-produced bile salts are commercially unavailable. Cross-adaptation experiments indicate further that the responses to bile salts are independent of those to amino acids, a major class of odorant molecules for all tested fishes.

Major differences in the proportion of amino acid fiber types transmitting taste information from oral and extraoral regions in the channel catfish.

The present study investigates for the first time in any teleost the amino acid specificity and sensitivity of single glossopharyngeal (cranial nerve IX) fibers that innervate taste buds within the oropharyngeal cavity. These results are contrasted with similar data obtained from facial (cranial nerve VII) fibers that innervate extraoral taste buds. The major finding is that functional differences are clearly evident between taste fibers of these two cranial nerves. Catfish possess the most extensive distribution of taste buds found in vertebrates. Taste buds on the external body surface are exclusively innervated by VII, whereas IX, along with the vagus (X), innervate the vast majority of taste buds within the oropharyngeal cavity. Responses to the l-isomers of alanine (Ala), arginine (Arg), and proline (Pro), the three most stimulatory amino acids that bind to independent taste receptors, were obtained from 90 single VII and 64 single IX taste fibers. This study confirmed a previous investigation that the amino acid responsive VII fibers consist of two major groups, the Ala and Arg clusters containing taste fibers having thresholds in the ηM range. In contrast, the present study indicates the amino acid responsive IX taste system is dominated by taste fibers responsive to Pro and to Pro and Arg, respectively, has a reduced percentage of Ala fibers, and is less sensitive than VII. The present electrophysiological results are consistent with previous experiments, indicating that the extraoral taste system is essential for appetitive behavior, whereas oropharyngeal taste buds are critical for consummatory behavior.

Highly specific olfactory receptor neurons for types of amino acids in the channel catfish.

Odorant specificity to l-alpha-amino acids was determined electrophysiologically for 93 single catfish olfactory receptor neurons (ORNs) selected for their narrow excitatory molecular response range (EMRR) to only one type of amino acid (i.e., Group I units). These units were excited by either a basic amino acid, a neutral amino acid with a long side chain, or a neutral amino acid with a short side chain when tested at 10(-7) to 10(-5) M. Stimulus-induced inhibition, likely for contrast enhancement, was primarily observed in response to the types of amino acid stimuli different from that which activated a specific ORN. The high specificity of single Group I ORNs to type of amino acid was also previously observed for single Group I neurons in both the olfactory bulb and forebrain of the same species. These results indicate that for Group I neurons olfactory information concerning specific types of amino acids is processed from receptor neurons through mitral cells of the olfactory bulb to higher forebrain neurons without significant alteration in unit odorant specificity.

Amplitude modulation patterns of local field potentials reveal asynchronous neuronal populations.

Neural oscillations, which appear in several areas of the nervous system and cover a wide frequency range, are a prominent issue in current neuroscience. Extracellularly recorded oscillations are generally thought to be a manifestation of a neural population with synchronized electrical activity resulting from coupling mechanisms. The vertebrate olfactory neuroepithelium exhibits beta-band oscillations, termed peripheral waves (PWs), in their population response to odor stimulation. Here, we examine PWs in the channel catfish and propose that their properties could be explained as the superposition of asynchronous oscillators. Our model shows that the intriguing random pattern of amplitude-modulated PWs could be explained by Rayleigh fading, an interference phenomenon well known in physics and recognizable using statistical methods and signal analysis. We are proposing a mathematical fingerprint to characterize neural signals generated by the addition of random phase oscillators. Our interpretation of PWs as arising from asynchronous oscillators could be generalized to other neuronal populations, because it suggests that neural oscillations, detected in local field potential recordings within a narrow frequency band, do not necessarily originate from synchronization events.

Responses of olfactory forebrain units to amino acids in the channel catfish.

A paucity of information exists concerning the processing of odorant information by single neurons in any vertebrate above the level of the olfactory bulb (OB). In this report, odorant specificity to four types of L-alpha-amino acids (neutral with long side-chains, neutral with short side-chains, basic and acidic), known biologically relevant odorants for teleosts, was determined for 217 spontaneously active forebrain (FB) neurons in the channel catfish. Group I FB units were identified that were excited by only one of four types of amino acids; no Group I unit was encountered that was excited by an acidic amino acid. The Group I FB units exhibited similar preferences as described previously for OB neurons, suggesting that no major modifications of olfactory information for at least some of these units occurred between the OB and FB. Evidence, however, for the convergence of odor information between the OB and FB was suggested by Group II FB units that exhibited a broader excitatory molecular receptive range (EMRR) than those of previously recorded types of OB units or the Group I FB units. Group II FB units were excited by both neutral and basic amino acids and a few also by acidic amino acids, EMRRs not observed previously in OB units. Stimulus-induced inhibition, likely for contrast enhancement, was also often observed for the many of the FB units encountered. The observed EMRRs of the FB units presently identified and those of the OB units previously studied are consistent with the ability of catfish to behaviorally discriminate these compounds.

Beyond the olfactory bulb: an odotopic map in the forebrain.

We report electrophysiological evidence that a simple odotopy, the spatial mapping of different odorants, is maintained above the level of the olfactory bulb (OB). Three classes of biologically relevant odorants for fish are processed in distinct regions of the forebrain (FB) in the channel catfish. Feeding cues, mainly amino acids and nucleotides, are represented in lateral, pallial portions of the FB, equivalent to the olfactory cortex of amniote vertebrates, whereas social signals mediated by bile salts are represented in medial FB centers, possibly homologous to portions of the amygdala. As in the OB, the different odorant classes map onto different territories; however, the response properties of units of the olfactory areas of the FB do not simply mirror those of the OB. For some units, distinctive response properties emerged, because the FB is the first center where odors subserving a common behavioral function (i.e., food function) converge.

Odorant specificity of single olfactory bulb neurons to amino acids in the channel catfish.

Odorant specificity to l-alpha-amino acids was determined for 245 olfactory bulb (OB) neurons recorded from 121 channel catfish. The initial tests included 4 amino acids representing acidic [monosodium glutamate (Glu)], basic [arginine (Arg)], and neutral [possessing short: alanine (Ala) and long: methionine (Met) side chains] amino acids that were previously indicated to bind to independent olfactory receptor sites. Ninety-one (37%) units (Group I) tested at 1, 10, and 100 microM showed high selectivity and were excited by only one of the 4 amino acids. Odorant specificity for the vast majority of Group I units did not change over the 3 s of response time analyzed. A total of 154 OB units (63%) (Group II) were excited by a second amino acid, but only at >/=10x odorant concentration. An additional 69 Group I units were tested with related amino acids and derivatives from 10(-9) to 10(-5) M to determine their excitatory odorant thresholds and selectivities. Two groups of units originally selective for Met were evident: those most sensitive to neutral amino acids having branched and linear side chains, respectively. OB units originally selective for Ala responded at low concentration to other similar amino acids. Units originally selective for Arg were excited at low concentration by amino acids possessing in their side chains at least 3 methylene groups and a terminal amide or guanidinium group. The specificities of the OB units determined electrophysiologically are sufficient to account for many of the previous results of behavioral discrimination of amino acids in this and related species.

Correlation between olfactory receptor cell type and function in the channel catfish.

The olfactory epithelium of fish contains three intermingled types of olfactory receptor neurons (ORNs): ciliated, microvillous, and crypt. The present experiments were undertaken to test whether the different types of ORNs respond to different classes of odorants via different families of receptor molecules and G-proteins corresponding to the morphology of the ORN. In catfish, ciliated ORNs express OR-type receptors and Galpha(olf). Microvillous ORNs are heterogeneous, with many expressing Galpha(q)/11, whereas crypt ORNs express Galpha(o). Retrograde tracing experiments show that ciliated ORNs project predominantly to regions of the olfactory bulb (OB) that respond to bile salts (medial) and amino acids (ventral) (Nikonov and Caprio, 2001). In contrast, microvillous ORNs project almost entirely to the dorsal surface of the OB, where responses to nucleotides (posterior OB) and amino acids (anterior OB) predominate. These anatomical findings are consistent with our pharmacological results showing that forskolin (which interferes with Galpha(olf)/cAMP signaling) blocks responses to bile salts and markedly reduces responses to amino acids. Conversely, U-73122 and U-73343 (which interfere with Galpha(q)/11/phospholipase C signaling) diminish amino acid responses but leave bile salt and nucleotide responses essentially unchanged. In summary, our results indicate that bile salt odorants are detected predominantly by ciliated ORNs relying on the Galpha(olf)/cAMP transduction cascade. Nucleotides are detected by microvillous ORNs using neither Galpha(olf)/cAMP nor Galpha(q)/11/PLC cascades. Finally, amino acid odorants activate both ciliated and microvillous ORNs but via different transduction pathways in the two types of cells.

Odorant-induced olfactory receptor neural oscillations and their modulation of olfactory bulbar responses in the channel catfish.

Peripheral waves (PWs) in the channel catfish are odorant-induced neural oscillations of synchronized populations of olfactory receptor neurons (ORNs) that appear after the initial approximately 500 msec of the response. The mean dominant frequency during the initial 2 sec of PW activity is approximately 28 Hz, declining to approximately 20 Hz in the last sec of a 5 sec stimulus. Recordings of PWs from different regions of a single olfactory lamella and simultaneously from widely separated lamellae within the olfactory organ suggest that PWs are initiated in the sensory epithelium within each olfactory lamella. Simultaneous recordings in vivo from the olfactory organ [electro-olfactogram (EOG) or integrated neural activity], local field potentials (LFPs) from the olfactory bulb (OB), and single and few-unit activity from OB neurons were performed. Cross-correlation analysis of simultaneously recorded odor-induced OB LFPs and either EOG or ORN neural activity showed that oscillations occurring within the OB were lower (<20 Hz) than those of PWs; however, during PW activity, OB LFPs increased both their magnitude and dominant frequencies and became correlated with the PWs. Also during odorant-induced PW activity, the responses of different OB neurons with similar odorant specificity became phase locked to each other and to both the PWs and OB LFPs. PWs are hypothesized to function to strengthen the synaptic transfer of olfactory information at specific glomeruli within the OB.

FINE STRUCTURE AND VITAL STAINING OF OSPHRADIUM OF THE SOUTHERN OYSTER DRILL, THAIS HAEMASTOMA CANALICULATA (GRAY) (PROSOBRANCHIA: MURICIDAE).

The morphology of the osphradium of Thais haemastoma canaliculata (Gray) was examined using light microscopy, SEM, and TEM. The osphradium is composed of approximately 150-200 lamellae, each of which is divided into two distinct regions by a groove situated parallel to the dorsal edge of the organ. The dorsal one-fourth of each lamella is covered by dense cilia that are assumed to generate water currents about the osphradium. Ciliary tufts, located in small depressions, and numerous secretory cells are distributed uniformly on the ventral three-fourths of the lamellae. A thin tract of cilia borders the ventral edge of each lamella. The overall cellular organization is less complex than has been reported previously in other marine prosobranchs. Selective staining of putative chemoreceptors was performed using Procion Brilliant Yellow. Individual cells in the ventral region and the ventral edge of each lamella were Procion-positive. Results of this study suggest that ventral interlamellar regions and the ventral edge of each lamella are chemosensory regions, while the dorsal portion of each lamella is indifferent epithelium.