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TESS-based dose-response using pediatric clonidine exposures.
OBJECTIVE: The toxic and lethal doses of clonidine in children are unclear. This study was designed to determine whether data from the American Association of Poison Control Centers Toxic Exposure Surveillance System (TESS) could be utilized to determine a dose-response relationship for pediatric clonidine exposure. METHODS: 3,458 single-substance clonidine exposures in children <6 years of age reported to TESS from January 2000 through December 2003 were examined. Dose ingested, age, and medical outcome were available for 1550 cases. Respiratory arrest cases (n = 8) were classified as the most severe of the medical outcome categories (Arrest, Major, Moderate, Mild, and No effect). Exposures reported as a "taste or lick" (n = 51) were included as a dose of 1/10 of the dosage form involved. Dose ranged from 0.4 to 1980 (median 13) microg/kg. Weight was imputed based on a quadratic estimate of weight for age. Dose certainty was coded as exact (26% of cases) or not exact (74%). Medical outcome (response) was examined via logistic regression using SAS JMP (release 5.1). RESULTS: The logistic model describing medical outcome (P < 0.0001) included Log dose/kg (P = 0.0000) and Certainty (P = 0.045). CONCLUSION: TESS data can provide the basis for a statistically sound description of dose-response for pediatric clonidine poisoning exposures.
Taste reactivity in the hamster.
Taste reactivity, which was first described in the rat, consists of ingestive and aversive response components, the latter seen mostly to bitter-tasting stimuli. The present experiment characterized the hamster's taste reactivity to an array of stimuli (sugars: 1 M sucrose, d-fructose and d-glucose; sodium salts: 1 M NaCl, Na2SO4 and NaNO3; acids: 30 mM HCl, tartaric acid and citric acid; bitter-tasting stimuli: 100 mM quinine hydrochloride and nicotine sulfate and 10 mM denatonium benzoate). These 12 stimuli were chosen to represent 3 examples each of stimuli that taste sweet, salty, sour, or bitter to humans; they were presented in random order via an intraoral fistula, one stimulus each day per animal (n = 10). Infusions of 0.6 ml were delivered over a 1-min period from a syringe pump. Orofacial and somatic motor responses were recorded on videotape for later analysis and were also coded online into a computer. Ingestive responses included forward and lateral tongue protrusions and aversive responses included gaping, chin rubbing, forelimb flailing, fluid rejection, increased locomotion, and aversive posturing. Each stimulus group produced a characteristic pattern of these behaviors, with sugars eliciting only ingestive behaviors and the bitter stimuli evoking predominantly aversive responses. Both sodium salts and acids produced ingestive responses, as seen previously in the rat, although these stimuli also elicited aversive behaviors in the hamster, including apes. The patterns of responses were characterized using multivariate procedures; the stimuli fell into distinct groups that were separated primarily along an hedonic dimension.
Allelic variation in the GABA A receptor gamma2 subunit is associated with genetic susceptibility to ethanol-induced motor incoordination and hypothermia, conditioned taste aversion, and withdrawal in BXD/Ty recombinant inbred mice.
BACKGROUND: Behavioral genomics has made dramatic progress toward mapping quantitative trait loci (QTLs) that contain genes responsible for phenotypic differences in a variety of behavioral responses to alcohol (ethanol). We previously identified a QTL on mouse Chromosome 11 that affects genetic predisposition to acute alcohol withdrawal. Among mice derived from the C57BL/6J (B6) and DBA/2J (D2) inbred strains, this QTL (Alcw3) accounts for 12% of the genetic variability in withdrawal liability. Candidate genes within this QTL encode the gamma-aminobutyric acid type A (GABA A) receptor gamma2, alpha1, alpha6, and beta2 subunits. We recently identified a coding sequence polymorphism between the B6 and D2 strains for the GABA A receptor gamma2 subunit gene (Gabrg2). In this study, we expand our analysis to a panel of BXD strains derived from the B6 and D2 progenitor strains. These BXD strains provide 26 fixed recombinant genotypes that can be used to examine genetic correlations, for example, between a phenotype of interest and allelic variation in a candidate gene. METHODS: Gabrg2 was cloned and sequenced from the 26 BXD recombinant inbred strains. We analyzed genetic correlations between allelic variation in Gabrg2 and alcohol phenotypes previously measured in the BXD strain means. RESULTS: Allelic variation in Gabrg2 is correlated genetically with predisposition to acute alcohol withdrawal and may underlie the Alcw3 locus. In addition, Gabrg2 is associated with ethanol-conditioned taste aversion, ethanol-induced motor incoordination, and ethanol-induced hypothermia. A trend is observed for chronic ethanol withdrawal, ethanol-induced loss of righting reflex, and tolerance to ethanol-induced hypothermia and ataxia. CONCLUSIONS: Functionally relevant variation in Gabrg2, or a closely linked gene, is correlated genetically with some, but not all, behavioral responses to alcohol. The alcohol-related phenotypes associated with Gabrg2 generally may be characterized as debilitating or motivationally negative.
Dynamic and multimodal responses of gustatory cortical neurons in awake rats.
To investigate the dynamic aspects of gustatory activity, we recorded the responses of small ensembles of cortical neurons to tastants administered to awake rats. Multiple trials of each tastant were delivered during recordings made in oral somatosensory (SI) and gustatory cortex (GC). When integrated tastant responses (firing rates averaged across 2.5 sec) were compared with water responses, 14.4% (13/90) of the GC neurons responded in a taste-specific manner. When time was considered as a source of information, however, the incidence of taste-specific firing increased: as many as 41% (37/90) of the recorded GC neurons exhibited taste-specific patterns of response. For 17% of the neurons identified as responding with taste-specific patterns, the stimulus that caused the most significant response was a function of the time since stimulus delivery. That is, a single neuron might respond most strongly to one tastant in the first 500 msec of a response and then respond most strongly to another tastant later in the response. Further analysis of the time courses of GC and SI cortical neural responses revealed that modulations of GC firing rate arose from three separable processes: early somatosensory input (less than approximately 0.2 sec post-stimulus), later chemosensory input ( approximately 0.2-1 sec), and delayed somatosensory input related to orofacial responses (more than approximately 1.0 sec). These data demonstrate that sensory information is available in the time course of GC responses and suggest the viability of views of gustatory processing that treat the temporal structure of cortical responses as an integral part of the neural code.
Conditioned taste aversion with lipopolysaccharide and peptidoglycan does not activate cytokine gene expression in the spleen and hypothalamus of mice.
Several reports show that behavioural and physiological components of the acute phase reaction can be conditioned. However, the mechanisms responsible for these effects remain obscure. The underlying assumption that the changes observed in conditioned animals are dependent on a conditioned production of cytokines has never been demonstrated. In the present study, the possibility of conditioning the production of cytokines or molecules implicated in their signalling pathways was tested by submitting mice to conditioned taste aversion with a new saccharin taste paired with intraperitoneal (i.p.) injections of lipopolysaccharide (LPS, 0.83 microg/g) or peptidoglycan (PGN, 20 microg/g). After two conditioning sessions, conditioned mice developed a clear aversion to saccharine that was not associated with activation of genes of the cytokine network either at the periphery, or in the hypothalamus, as demonstrated by a macroarray approach and confirmed by real time RT-PCR. In contrast, there was an activation of the genes coding for nuclear factor kappa B (NFkappaB) and mitogen activated protein kinase (MAPK) signalling pathways in the spleen and to a lesser extent in the hypothalamus. This modulation of the NFkappaB and MAPK signalling pathways is interpreted in terms of a possible conditioned sensitisation of the immune system.
Differential encoding of behavior and spatial context in deep and superficial layers of the neocortex.
Rodent hippocampal activity is correlated with spatial and behavioral context, but how context affects coding in association neocortex is not well understood. The cellular distribution of the neural activity-regulated immediate-early gene Arc was used to monitor the activity history of cells in CA1, and in deep and superficial layers of posterior parietal and gustatory cortices (which encode movement and taste, respectively), during two behavioral epochs in which spatial and behavioral context were independently manipulated while gustatory input was held constant. Under conditions in which the hippocampus strongly differentiated behavioral and spatial contexts, deep parietal and gustatory layers did not discriminate between spatial contexts, whereas superficial layers in both neocortical regions discriminated well. Deep parietal cells discriminated behavioral context, whereas deep gustatory cortex neurons encoded the two conditions identically. Increased context sensitivity of superficial neocortical layers, which receive more hippocampal outflow, may reflect a general principle of neocortical organization for memory retrieval.
Why do so many drug users smoke?
To better understand why most persons in drug treatment smoke, we explored patients' views of the relationship between their smoking, methadone, and drug use. Recruiting from four methadone clinics, we held seven focus groups with 68 current smokers, and 10 individual interviews with former smokers. Sessions were audio-taped, transcribed, and coded. Participants were > or =18 years old, smoked > or =5 cigarettes per day, and had used prescription methadone for > or =2 years. Most patients linked smoking, methadone, and drug use in three ways. First, patients said smoking and drug use were complementary. Examples included smoking to reduce methadone after taste and using drugs to reduce smoking-related throat pain. Second, smoking and drug use were similar because they shared cues and withdrawal symptoms. Third, smoking differed from drug use because it had fewer acute consequences. Because smoking, methadone, and drug use are closely interrelated, future addictions research and treatment protocols should address them in combination.
Ontogenesis and taste bud cell turnover in the chicken. I. Gemmal cell renewal in the hatchling.
Taste bud cell turnover rate was examined in oral epithelium of the precocial chick, which at hatching contains the adult complement of taste buds. Forty newly hatched chicks received single or double pulse injections of tritiated thymidine (specific activity was 6.7 Curies/millimole; dosage was 0.5 microCuries/g body weight, intraperitoneally). Anterior mandibular epithelium was processed for light microscopic autoradiography at 2 and 16 hours, as well as 1, 2, 3, 4, 6, 8, 10, 12, 14, 16, 18, and 20 days after the initial pulse. In a coded and randomized procedure, the section (7 microns) through the bud's center was selected for counting > or = 6 silver grains over round-clear and gracile-dense gemmal cell nuclei. The mean number of labelled cells/bud varied significantly (P < or = 0.01) during the first four posthatch days, yielding the fastest gemmal cell turnover rates (3.4-4.4 days) yet reported in vertebrates. Average bud diameter also significantly changed during the first four posthatch days, and was reflected in shifts of the distribution of 40-69 microns and > or = 70 microns diameter buds. Both an increase in labelled bud cells and bud diameter during the first two posthatch days may reflect high proliferation rates in initially maturing buds. Subsequent decrease in bud diameter between 2 and 3 days postinjection may indicate splitting of large-diameter (> or = 70 microns) buds and/or normal bud cell death due to failure of sensory afferentation. Bud-splitting alone, however, cannot account for significant decreases in bud cell label which did not occur before 4-6 days postinjection.
Worms taste bitter: ASH neurons, QUI-1, GPA-3 and ODR-3 mediate quinine avoidance in Caenorhabditis elegans.
An animal's ability to detect and avoid toxic compounds in the environment is crucial for survival. We show that the nematode Caenorhabditis elegans avoids many water-soluble substances that are toxic and that taste bitter to humans. We have used laser ablation and a genetic cell rescue strategy to identify sensory neurons involved in the avoidance of the bitter substance quinine, and found that ASH, a polymodal nociceptive neuron that senses many aversive stimuli, is the principal player in this response. Two G protein alpha subunits GPA-3 and ODR-3, expressed in ASH and in different, nonoverlapping sets of sensory neurons, are necessary for the response to quinine, although the effect of odr-3 can only be appreciated in the absence of gpa-3. We identified and cloned a new gene, qui-1, necessary for quinine and SDS avoidance. qui-1 codes for a novel protein with WD-40 domains and which is expressed in the avoidance sensory neurons ASH and ADL.
Neural responses to bitter compounds in rats.
To determine whether the idiosyncratic distribution of transduction mechanisms for bitter tastants in rat taste receptor cells (TRCs) could be inferred from the neural activity they evoke, single neuron responses to ten bitter-tasting compounds were recorded from rat glossopharyngeal (n = 30) and chorda tympani (n = 22) neurons. Responses to several 'bitter' alkaloids were obtained: 10 mM quinine-HCl, 50 mM caffeine, and 1 mM each nicotine, yohimbine, and strychnine, plus a number of non-alkaloid bitter-tasting compounds: 0.1 M KCl, 0.01 M MgCl2, and 1 mM each phenylthiocarbamide (PTC), L-tyrosine, and denatonium benzoate. To obtain some distinctions with other stimuli NaCl (0.1 M), HCl (pH 2.0), and capsaicin (10 microM) were also tested. It was found that individual neurons in both glossopharyngeal and chorda tympani nerves differed in their relative sensitivities to the various bitter stimuli. To determine relationships among these stimuli, the differences in the evoked responses between each stimulus pair were summarized in a multi-dimensional scaling space. In these analyses neither nerve showed any obvious similarity between the placements of quinine and the other bitter stimuli. Such data suggest that first-order gustatory neurons can discriminate among the above bitter stimuli. For glossopharyngeal neurons, some similarity to quinine was found only for nicotine and denatonium, and for chorda tympani neurons, some similarity to quinine was found only for KCl and MgCl2. Of the bitter compounds tested, quinine evoked the greatest response from glossopharyngeal neurons. We propose this arises because quinine can activate TRCs by more transduction mechanisms than other bitter stimuli. The results from these studies were summarized in a qualitative model for the coding of bitter tastants where the variety of transduction mechanisms for bitters are distributed among various TRCs to account for the heterogeneous responses among the neurons.
Temporal codes and computations for sensory representation and scene analysis.
This paper considers a space of possible temporal codes, surveys neurophysiological and psychological evidence for their use in nervous systems, and presents examples of neural timing networks that operate in the time-domain. Sensory qualities can be encoded temporally by means of two broad strategies: stimulus-driven temporal correlations (phase-locking) and stimulus-triggering of endogenous temporal response patterns. Evidence for stimulus-related spike timing patterns exists in nearly every sensory modality, and such information can be potentially utilized for representation of stimulus qualities, localization of sources, and perceptual grouping. Multiple strategies for temporal (time, frequency, and code-division) multiplexing of information for transmission and grouping are outlined. Using delays and multiplications (coincidences), neural timing networks perform time-domain signal processing operations to compare, extract and separate temporal patterns. Separation of synthetic double vowels by a recurrent neural timing network is used to illustrate how coherences in temporal fine structure can be exploited to build up and separate periodic signals with different fundamentals. Timing nets constitute a time-domain scene analysis strategy based on temporal pattern invariance rather than feature-based labeling, segregation and binding of channels. Further potential implications of temporal codes and computations for new kinds of neural networks are explored.
Artificial neural network analysis of gustatory responses in the thalamic taste relay of the rat.
We used an artificial neural network (ANN) as a model for analyzing single-neuron responses from the thalamic taste relay of rats. The network consisted of: (1) a layer of 44 input units, representing the responses of the 44 thalamic taste cells; (2) a layer of hidden units of varying numbers; and (3) a layer of four output units. We used the back-propagation algorithm to train the output units to discriminate among tastants based on inputs from the thalamic neurons. As the network became fully trained, we found that: (1) only two hidden units were necessary to provide nearly the full discriminative capacity of the network; (2) the loss of even a few of the input units that had the highest impact on hidden units caused a drastic reduction of discriminative power, implying that not all neurons contribute equally to the neural code; and (3) adding a temporal component to the input, by representing each 100-ms time bin as a separate input unit, increased the accuracy with which output units were able to identify tastants. We used data from behavioral discrimination tasks as a measure of the capacity of the network to identify stimuli correctly. A network with two hidden units was about as effective as an across-pattern analysis in accounting for the rat's discriminative ability.
Responses of single lingual nerve fibers to thermal and chemical stimulation.
The goals of this study were to characterize the responses of: (1) thermally-sensitive fibers of the lingual branch of the trigeminal nerve to cooling from 35 degrees to 10 degrees C at a rate of 1 degrees C/s; and (2) these neurons to a mid-range concentration of NaCl (150 mM), glucose (150 mM), citric acid (0.3 mM), and quinine-HCl (3 mM) at 35 degrees and 25 degrees C. A cluster analysis of 47 neurons' responses to cooling revealed two major groups and one minor group. Group 1 neurons (n=19) had a shorter latency, exhibited faster time-to-peak activity, and responded over a smaller range of temperature compared to Group 2 neurons (n=22). Group 3 neurons (n=6) exhibited the longest response latency and responded over a wider cooler range of temperature. Twenty-five out of thirty-one thermally-sensitive, non-tactile lingual neurons responded weakly to at least one chemical stimulus, with some neurons responding to 2, 3, or all 4 chemical stimuli. Group 1 neurons responded to more chemical stimuli at 35 degrees C, while Group 2 neurons responded more at 25 degrees C. Under their optimal temperature conditions, Group 1 and Group 2 neurons responded most often to citric acid and least often to glucose, with NaCl and Q-HCl eliciting an intermediate number of responses. As a whole, the responses of thermally-sensitive fibers to chemical stimulation were modest at best with an absence of chemical specificity. There was no evidence of a 'best' stimulus, although there was a suggestion of temporal coding.
Chemosensory proteins from the proboscis of mamestra brassicae.
Soluble, low molecular weight proteins were immunodetected in proboscis extracts of Mamestra brassicae males by Western blot, using antibodies raised against the general odorant-binding protein of the moth Antheraea polyphemus. The same antibodies weakly labelled the sensillum lymph and subcuticular space of sensilla styloconica on ultrathin sections of the proboscis. The morphology of sensilla styloconica is described. The immunodetected proteins yielded several N-terminal sequences, three of which showed strong affinity for tritiated analogues of pheromonal compounds of M. brassicae in binding assays. The cDNAs coding for these sequences were cloned and it was shown that the new proteins are related to the OS-D protein of Drosophila. They are named chemosensory proteins (CSP-MBRA:A1-CSP-MBRA:A5 and CSP-MBRA:B1 and CSP-MBRA:B2) and may have an odorant-binding protein-like function. A common localization in both olfaction and taste organs suggests a physiological role depending on the cellular environment.
Natural selection and molecular evolution in PTC, a bitter-taste receptor gene.
The ability to taste phenylthiocarbamide (PTC) is a classic phenotype that has long been known to vary in human populations. This phenotype is of genetic, epidemiologic, and evolutionary interest because the ability to taste PTC is correlated with the ability to taste other bitter substances, many of which are toxic. Thus, variation in PTC perception may reflect variation in dietary preferences throughout human history and could correlate with susceptibility to diet-related diseases in modern populations. To test R. A. Fisher's long-standing hypothesis that variability in PTC perception has been maintained by balancing natural selection, we examined patterns of DNA sequence variation in the recently identified PTC gene, which accounts for up to 85% of phenotypic variance in the trait. We analyzed the entire coding region of PTC (1,002 bp) in a sample of 330 chromosomes collected from African (n=62), Asian (n=138), European (n=110), and North American (n=20) populations by use of new statistical tests for natural selection that take into account the potentially confounding effects of human population growth. Two intermediate-frequency haplotypes corresponding to "taster" and "nontaster" phenotypes were found. These haplotypes had similar frequencies across Africa, Asia, and Europe. Genetic differentiation between the continental population samples was low (FST=0.056) in comparison with estimates based on other genes. In addition, Tajima's D and Fu and Li's D and F statistics demonstrated a significant deviation from neutrality because of an excess of intermediate-frequency variants when human population growth was taken into account (P<.01). These results combine to suggest that balancing natural selection has acted to maintain "taster" and "nontaster" alleles at the PTC locus in humans.
Trehalose sensitivity in Drosophila correlates with mutations in and expression of the gustatory receptor gene Gr5a.
Drosophila taste gene Tre is located on the distal X chromosome and controls gustatory sensitivity to a subset of sugars [1, 2]. Two adjacent, seven-transmembrane domain genes near the Tre locus are candidate genes for Tre. One (CG3171) encodes a rhodopsin family G protein receptor [3, 4], and the other (Gr5a) is a member of a chemosensory gene family encoding a putative gustatory receptor [5-7]. We carried out molecular analyses of mutations in Tre to elucidate their involvement in the gustatory phenotype. Here, we show that Tre mutations induced by P element-mediated genomic deletions disrupt Gr5a gene organization and the expression of Gr5a mRNA, while disruption of the CG3171 gene or its expression was not always associated with mutations in Tre. In flies with the spontaneous mutation Tre(01), both CG3171 and Gr5a mRNAs are transcribed. Coding sequences of these two candidate genes were compared among various strains. A total of three polymorphic sites leading to amino acid changes in CG3171 were not correlated with the gustatory phenotype. Among four nonsynonymous sites in Gr5a, a single nucleotide polymorphism leading to an Ala218Thr substitution in the predicted second intracellular loop cosegregated with Tre(01). Taken together, the mutation analyses support that Gr5a is allelic to Tre.
Marketing to America's youth: evidence from corporate documents.
OBJECTIVE: To evaluate the claim that the tobacco industry does not market its products to youth. DESIGN: The data for this study come from tobacco industry documents collected from the tobacco industry's document websites, presently linked at http://www.tobaccoarchives.com. The websites were searched using "request for production" (RFP) codes, specified keyword searches, and serendipitous terms identified in document citations found with RFP and keyword searches. RESULTS: Industry documents show that the cigarette manufacturers carefully monitored the smoking habits of teenagers over the past several decades. Candid quotes from industry executives refer to youth as a source of sales and as fundamental to the survival of the tobacco industry. The documents reveal that the features of cigarette brands (that is, use of filters, low tar, bland taste, etc), packaging (that is, size, colour and design), and advertising (that is, media placements and themes and imagery) were developed specifically to appeal to new smokers (that is, teenagers). Evidence also indicates that relevant youth oriented marketing documents may have been destroyed and that the language used in some of the more recent documents may have been sanitised to cover up efforts to market to youth. CONCLUSIONS: The tobacco industry's internal documents reveal an undeniable interest in marketing cigarettes to underage smokers. The industry's marketing approaches run counter to and predicate methods for tobacco prevention: (1) keep the price of the product high; (2) keep product placements and advertising away from schools and other areas with a high volume of youth traffic; (3) make cigarette advertising (that is, themes and visual images) unappealing to youth; (4) make product packaging unappealing to youth; and (5) design the product so it is not easy to inhale.