C57BL/6J and DBA/2J mice vary in lick rate and ingestive microstructure

Fluid licking in mice is an example of a rhythmic behavior thought to be under the control of a central pattern generator. Inbred strains of mice have been shown to differ in mean or modal interlick interval (ILI) duration, suggesting a genetic-based variation. We investigated water licking in the commonly used inbred strains C57BL/6J (B6) and DBA/2J (D2), using a commercially available contact lickometer. Results from 20-min test sessions indicated that D2 mice lick at a faster rate than B6 mice (10.6 licks/s vs. 8.5 licks/s), based on analysis of the distribution of short-duration ILIs (50-160 ms). This strain difference was independent of sex, extent of water deprivation or total number of licks. D2 mice also displayed a faster lick rate when the strains were tested with a series of brief (5 s) trials. However, when ingestion over the entire 20-min session was analyzed, it was evident that D2 mice had an overall slower rate of ingestion than B6 mice. This was because of the tendency for D2 mice to have more very long pauses (>30 s) between sequences of licking bursts. Overall, it appeared that D2 mice licked more efficiently, ingesting more rapidly during excursions to the spout that were fewer and farther between.     

A High Throughput In Vivo Assay for Taste Quality and Palatability

Taste quality and palatability are two of the most important properties measured in the evaluation of taste stimuli. Human panels can report both aspects, but are of limited experimental flexibility and throughput capacity. Relatively efficient animal models for taste evaluation have been developed, but each of them is designed to measure either taste quality or palatability as independent experimental endpoints. We present here a new apparatus and method for high throughput quantification of both taste quality and palatability using rats in an operant taste discrimination paradigm. Cohorts of four rats were trained in a modified operant chamber to sample taste stimuli by licking solutions from a 96-well plate that moved in a randomized pattern beneath the chamber floor. As a rat’s tongue entered the well it disrupted a laser beam projecting across the top of the 96-well plate, consequently producing two retractable levers that operated a pellet dispenser. The taste of sucrose was associated with food reinforcement by presses on a sucrose-designated lever, whereas the taste of water and other basic tastes were associated with the alternative lever. Each disruption of the laser was counted as a lick. Using this procedure, rats were trained to discriminate 100 mM sucrose from water, quinine, citric acid, and NaCl with 90-100% accuracy. Palatability was determined by the number of licks per trial and, due to intermediate rates of licking for water, was quantifiable along the entire spectrum of appetitiveness to aversiveness. All 96 samples were evaluated within 90 minute test sessions with no evidence of desensitization or fatigue. The technology is capable of generating multiple concentration–response functions within a single session, is suitable for in vivo primary screening of tastant libraries, and potentially can be used to evaluate stimuli for any taste system.     

Disruptive effects of lateral hypothalamic stimulation on the lick-interrupt cycle in rats

Consummatory licking at a water spout was compared with licking at a dry spout maintained by electrical hypothalamic stimulation in the same rats. Both forms of licking, recorded photoelectrically, were maintained on a fixed ratio 8 schedule. Duration of reinforcement delivery was equated [300 ms]. A computer analysis of the temporal distribution of licks in each 1024 ms period from onset of reinforcement revealed that lateral hypothalamic stimulation decreased the occurrence of licking and disrupted the normally synchronous pattern of this behaviour. An analysis of the effects of delivering lateral hypothalamic stimulation contingent on water-maintained licking revealed that this effect of stimulation was clearly current-dependent. It is proposed that differences in licking rates maintained by water and by electrical hypothalamic stimulation, respectively, are due to response interference in the latter case. This interference effect is also proposed to be a major factor underlying higher reward thresholds for self-stimulation when licking is the operant response.     

Excessive habit formation in schedule‐induced polydipsia: Microstructural analysis of licking among rat strains and involvement of the orbitofrontal cortex

Schedule‐induced polydipsia (SIP) is an animal model of compulsive drinking that selects for individual differences and varies across rat strains. The aim of this study was to investigate excessive habit formation by analyzing the SIP licking microstructure among rat strains, and to compare the brain areas activated by SIP in different populations. Wistar, Long Evans and Roman High‐ and Low‐Avoidance rat strains were compared using a cluster analysis of 2 main variables, that is, frequency of licking (percentage of interpellet intervals with drinking episodes) and intensity of licking (mean number of licks per interpellet interval), and were found to exhibit high intensity and frequent licking (compulsive drinkers, CD), low intensity but frequent licking (habitual drinkers, HD), and low intensity and low‐frequency licking (low drinkers, LD). The Wistar strain showed a higher frequency and intensity of licking, and had the largest group of CD rats when compared with the other strains. Regarding the acquisition of SIP, CD rats showed a higher intensity of licking when compared with the HD and LD rats. Moreover, c‐Fos quantification revealed that rats in the CD group showed hyperactivity in the lateral orbitofrontal cortex and basolateral amygdala when compared with the LD group. Analyzing the SIP microstructure could be a valuable tool for understanding the role of excessive habit formation in the development of compulsive drinking and its underpinning neurobiological mechanisms.     

A computational model for motor pattern switching between taste-induced ingestion and rejection oromotor behaviors

The mechanism of switching activity patterns in a central pattern generator is fundamental to the generation of diverse motor behaviors. Based on what is known about a brainstem substrate mediating the oral components of ingestion and rejection, we use computational techniques to construct a hypothetical multifunctional network that switches between the motor outputs of ingestion (licking) and rejection (gaping). The network was constructed using single-compartment conductance-based models for individual neurons based on Hodgkin-Huxley formalism. Using a fast-slow reduction and geometric analysis we describe a mechanism for pattern switching between licks and gapes. The model supports the hypothesis that a single configuration of network connections can produce both activity patterns. It further predicts that prolonged inhibition of some network neurons could lead to a switch in network activity from licks to gapes. Action Editor: Frances K. Skinner     

Multiple processes underlie benzodiazepine-mediated increases in the consumption of accepted and avoided stimuli

Hyperphagia is a reported side effect of anxiolytic benzodiazepines such as chlordiazepoxide (CDP). Prior research has focused primarily on the ingestive responses to sweet or solid foods. We examined CDP effects on licking for normally accepted and avoided taste solutions across a range of concentrations. The effect of CDP (10 mg/kg) versus saline on the licking patterns of water-restricted rats for water and 3 concentrations of sucrose, saccharin, NaCl, monosodium glutamate (MSG), citric acid, and quinine (Q-HCl) solutions was evaluated during 1 h tests. CDP increased meal size for all tastants except citric acid. Analysis of licking microstructure revealed 3 dissociable effects of CDP. CDP affected oromotor coordination as indicated by a uniform increase in the modal interlick interval for all stimuli. CDP increased meal size as indicated by shorter pauses during consumption of water, MSG, and weaker saccharin concentrations, and by fewer long interlick intervals (250-2000 ms) for normally avoided tastants. CDP also increased meal size by increasing burst size, burst duration, and the initial rate of licking for most solutions, suggesting increased hedonic taste evaluation. CDP did not affect variables associated with postingestive feedback such as meal duration or number of bursts, and the results also suggest that CDP did not enhance the perceived taste intensity. We hypothesize that the reduction of pause duration is consistent with an increased motivation to sample the stimulus that synergizes with changes in taste-mediated responsiveness to some but not all stimuli to yield increases in the consumption of both normally accepted and avoided taste stimuli.     

Within-litter variance in rat maternal behaviour

As a first step in determining the influence of maternal behaviour on sibling behavioural variance, we tested whether rat mothers differentially interact with neonates within the same litter. We also tested whether fading of an ink-mark on individual pups could provide an index of within-litter variance in maternal licking in laboratory rats. In Study 1, during the first postnatal week we distinguished individual Sprague-Dawley rat pups across 4 litters by placing an ink-mark on the skin and quantified variance in maternal licking frequency toward each pup and compared fading of individual pup marks to the frequency of maternal licks received and to four pup characteristics that could influence mark-fading. In Study 2, neonate mark-fading (a proxy for maternal licking) was compared to adolescent and adult offspring behaviour across 8 litters. Results indicated that: (1) there are substantial and consistent differences in how much rat mothers lick same-sex siblings within a litter, (2) differential licking rates can be documented with a non-observational method (ink-mark-fading), and (3) within-litter variance in maternal behaviour may relate to sibling behavioural variance. The findings indicate a viable research model for future experimental studies on causes and consequences of differential maternal investment within families.     

Mercaptoacetate inhibition of fatty acid beta-oxidation attenuates the oral acceptance of fat in BALB/c mice

We investigated the effect of beta-oxidation inhibition on the fat ingestive behavior of BALB/c mice. Intraperitoneal administration to mice of mercaptoacetate, an inhibitor of fatty acid oxidation, significantly suppressed intake of corn oil but not intake of sucrose solution or laboratory chow. To further examine the effect of mercaptoacetate on the acceptability of corn oil in the oral cavity, we examined short-term licking behavior. Mercaptoacetate significantly and specifically decreased the number of licks of corn oil within a 60-s period but did not affect those of a sucrose solution, a monosodium glutamate solution, or mineral oil. In contrast, the administration of 2-deoxyglucose, an inhibitor of glucose metabolism, did not affect the intake or short-term licking counts of any of the tasted solutions. These findings suggest that fat metabolism is involved in the mechanism underlying the oral acceptance of fat as an energy source.     

Role of licking and concomitant changes in the excitability of nerve centers in the elaboration of gustatory information in rats

The mechanism of nervous control of licking was studied in the frame work of the electrophysiological analysis of conditioned taste aversion. The licking rhythm is so stable that the rats are unable to slow it down by 20 percent. If the solution which was previously used for establishing the conditioned taste aversion, appears in the drinking spout, the rat stops drinking after one or two licks. Analysis of temporal relationships between afferent and efferent impulse activity of licking shows that the comparison of gustatory signals with memory traces and the corresponding decision occurs within 80 to 120 ms after lick onset. Electrical stimulation of the hypothalamic centres through implanted electrodes has shown that the amplitude of evoked responses and the impairment of licking increases, in proportion to the delay between lick onset and stimulus application. Isolated discharges of epileptic foci in the frontal cortex and the hypothalamus cause an omission of one or several licks, without interfering with the activity of the licking generator.     

Front Cover

Courtship displays are typically comprised of the same behavioral pattern, or patterns, repeated several times by males. Both the quantity and quality of the displays produced by a given male bird are not, however, constant. The number and/or quality of displays can decrease over time, indicating fatigue, or males can increase the number and/or quality as they display more, indicating a warm-up period. Although there is evidence for fatigue or warm-up periods for many types of courtship displays, data on motor components of avian courtship are scant, despite how commonly they are used. Here, we test whether drumming, a non-vocal motor display, in male ruffed grouse (Bonasa umbellus) changes in relation to the number of displays executed. Using a large number of recordings, our linear mixed models yielded a significant effect of cumulative number of drumming displays on the number of wingbeats per second, referred to as pulse rate. Across males, pulse rate is slowest when males begin drumming each day and increases until approximately 50 drumming displays have been produced. The rate of increase is also modulated by the nighttime low temperature such that cooler conditions are associated with lower pulse rates and a slower increase in pulse rate relative to the cumulative number of displays. Further, the maximum pulse rate recorded and average pulse rate after 50 displays is inversely correlated with body mass such that larger males are slower than smaller males. We suggest that the daily changes in pulse rate likely reflect a warm-up period based upon the effects of cumulative drumming count and temperature on pulse rate. Whether these dynamic changes in the production of a motor display are informative to female grouse is unknown. However, we propose that daily changes in how motor displays are performed may be a common feature of avian courtship that has gone relatively unnoticed, despite the potential for motor performance to be a trait that is important for female mate choice.     

Temporal patterns of visitation of birds and mammals at mineral licks in the Peruvian Amazon

Mineral licks are key ecological resources for many species of birds and mammals in Amazonia, providing essential dietary nutrients and clays, yet little is known about which species visit and their behaviors at the mineral licks. Studying visitation and behavior at mineral licks can provide insight into the lives of otherwise secretive and elusive species. We assessed which species visited mineral licks, when they visited, and whether visits and the probability of recording groups at mineral licks were seasonal or related to the lunar cycle. We camera trapped at 52 mineral licks in the northeastern Peruvian Amazon and detected 20 mammal and 13 bird species over 6,255 camera nights. Generalized linear models assessed visitation patterns and records of groups in association with seasonality and the lunar cycle. We report nocturnal curassows (Nothocrax urumutum) visiting mineral licks for the first time. We found seasonal trends in visitation for the black agouti (Dasyprocta fuliginosa), red howler monkey (Alouatta seniculus), blue‐throated piping guan (Pipile cumanensis), red brocket deer (Mazama americana), collared peccary (Pecari tajacu), and tapir (Tapirus terrestris). Lunar trends in visitation occurred for the paca (Cuniculus paca), Brazilian porcupine (Coendou prehensilis), and red brocket deer. The probability of recording groups (>1 individual) at mineral licks was seasonal and related to lunar brightness for tapir. Overall, our results provide important context for how elusive species of birds and mammals interact with these key ecological resources on a landscape scale. The ecological importance of mineral licks for these species can provide context to seasonal changes in species occupancy and movement.

Many species of animals in the Amazon visit mineral licks, consuming soil to supplement their diet. Using camera trap data, we show that visits to mineral licks are seasonal for many species. These trends in visitation are likely due to species‐specific factors such as reproduction, predator avoidance, seasonal diet shifts, and resource use.     

A quantitative ethogram of behavior of yearling ewes during two hours post-parturition

A total of 2130 behaviors representing 24 behavioral categories were observed during the period from parturition to lamb nursing for seven ewe-lamb pairs. Both individual behaviors and behavioral sequences were counted. Ewes showed a high frequency of licking, especially directed at the head (41% of licks) and anterior body (30% of licks) of their lambs. Licking followed a specific, non-random pattern. Ewes licked from front to back of their lambs. Ewes licked their lamb's anterior body and head immediately before their lambs stood. When the lambs sought teats, ewes either presented their udder to their lamb which facilitated nursing, or moved away from the lamb which delayed nursing. This research identified behaviors and behavioral sequences that may be used in future studies seeking to select for improved maternal ability or to design improved lambing environments. Finally, this work may lead to the generation of hypotheses which may be tested in future work examining behaviors involved as causal factors in lamb survival.     

Amiloride alters lick rate responses to NaCl and KCl in rats

The role-of cation channels on taste cell membranes to salttaste sensation was assessed in rats. We measured the numberof licks during multiple 10-s presentations of NaCl and KClconcentrations (0.05, 0.09, 0.16, 0.28, 0.5 M) dissolved ineither water or in 100 µM amiloride, a sodium-channelblocker. The number of licks to water and 0.3 M sucrose wasalso measured. The number of licks to NaCl was significantlylower and the number of licks to KCl was significantly higherwhen these test solutions were dissolved in amiloride than inwater. There were no differences in lick responses to waterand sucrose. These results suggest that amiloride may have alteredthe taste of NaCl and KCl. The results are discussed in relationshipto prior electrophysiological studies characterizing the effectof amiloride in blocking salt responses of the chorda tympaninerve.     

Interference competition and group size effect in sika deer (<Emphasis Type="Italic">Cervus nippon</Emphasis>) at salt licks

Competition has long been considered as a confounding factor of group size effect but the understanding of interference competition is rudimentary for the difficulty in disentangling interference competition from scramble competition adequately. Here, we analysed remote-camera video records of wild sika deer (Cervus nippon) at salt licks in southern China from March 1, 2006 to November 30, 2008 to investigate how interference competition and predation risk interacted on vigilance behaviour. Scramble competition is negligible at salt licks; therefore, we could focus our interest in interference competition. We used linear mixed model to compare vigilance, licking and aggression behaviours among females with and without fawn as well as males with different group sizes to identify the primary role of vigilance behaviour in sika deer. In total, 168 individuals were recorded and observation time was 2,733.04 min. We found that deer spent much time on vigilance and scanned frequently in spring and winter, and females with fawn spent more time on vigilance than females without fawn, suggesting vigilance for predation risk. Aggression ratio increased first and then decreased, while scan frequency continued to decline and then slightly increased when group size increased from two to seven, implying vigilance for interference competition. Our results suggested vigilance in sika deer was influenced by both predation risk and interference competition, but was mainly driven by predation risk even at sites with intense interference competition. Our results of interference competition shed some light on finding the underlying mechanism of group size effect in wild populations.     

Importance of natural licks for the mammals in Bornean inland tropical rain forests

An intensive camera-trapping study and a nutrient analysis were carried out to understand how natural licks are important for mammals in inland tropical rain forests where soil cations are usually depleted. Using camera traps, we investigated the fauna, food habits, and the frequency of visitation by species at five natural licks in the Deramakot forest reserve, Sabah, Malaysia. All food-habit types of mammals (carnivore, herbivore-frugivore, insectivore, and omnivore), which included 29 (78.4%) of 37 species known in Deramakot, were recorded at the natural licks. The sambar deer, followed by the bearded pig, the lesser mouse-deer, the Malay badger, and the orangutan were the most commonly recorded species and represented 77.5% in terms of the frequency of appearance in all photographs taken throughout the year. These results indicated that, although the proportion of species recorded at the natural licks relative to the whole mammalian fauna of the forest was high, the frequency of visitation greatly varied among the species, and only a few species dominated. The frequency of visitation seemed to reflect both the density of species and the demand for the minerals, because some endangered, low-density species were more frequently recorded by cameras than expected—for example, the orangutan which was one of the top five species among natural-lick users. The natural licks with greater concentrations of minerals in seepage soil water were significantly preferred by the sambar deer and the bearded pig than those with lower concentrations of minerals. This result suggests that the chemical properties of soil water in natural licks determine the frequency of visitation of these herbivorous species that have strong demand for minerals.     

Quantification of the honeybee's proboscis reflex by electromyographic recordings

The proboscis-extension reflex of the honeybee, which is subject to classical conditioning was examined by monitoring the proboscis movements with a video camera and recording extracellularly from the muscle which retracts the labium.

• 1.1. The movements of the proboscis during feeding are divided into three phases: extension, repeated licking phases (licks) and retraction.
• 2.2. Recordings from both symmetrically arranged muscles show a one to one relationship of muscle potentials during licks, but they are less synchronized during the extension phase, where the muscle ipsilateral to the stimulated antenna is activated first.
• 3.3. In an odour conditioning paradigm, the latency between the onset of the conditioned stimulus and the conditioned response shortens with acquisition and lengthens with extinction.
• 4.4. During extinction, the motor programme is gradually shortened by means of a decreased number of licks between the extension and retraction phases.

     

Changes in muscle activation can prolong the endurance time of a submaximal isometric contraction in humans.

Fourteen young subjects (7 men and 7 women) performed a fatiguing isometric contraction with the elbow flexor muscles at 20% of maximal voluntary contraction (MVC) force on three occasions. Endurance time for session 3 [1,718 +/- 1,189 (SD) s] was longer than for session 1 (1,225 +/- 683 s) and session 2 (1,410 +/- 977 s). Five men and four women increased endurance time between session 1 and 3 by 60 +/- 28% (responders), whereas two men and three women did not (-3 +/- 11%; nonresponders). The MVC force was similar for the responders and nonresponders, both before and after the fatiguing contraction. Fatiguing contractions were characterized by an increase in the electromyogram (EMG) amplitude and number of bursts during the fatiguing contractions. The responders achieved a similar level of EMG at exhaustion but a reduced rate of increase in the EMG across sessions. The rate of increase in EMG across sessions declined for the nonresponders, but it remained greater than that of the responders. The increase in burst rate during the contractions declined across sessions with a negative relation between burst rate and endurance time (r = -0.42). Normalized force fluctuations increased during the fatiguing contractions, and there was a positive relation (r = 0.60) between the force fluctuations and burst rate. Changes in mean arterial pressure and heart rate during the fatiguing contraction were similar for the responders and nonresponders across the three sessions. The results indicate that those subjects who increased the endurance time of a submaximal contraction across three sessions did so by altering the level and pattern of muscle activation.     

Electrophysiological analysis of retrieval of conditioned taste aversion in rats. Unit activity changes in critical brain regions.

Gustatory discrimination testing shows that rats with an overtrained conditioned taste aversion (CTA) to isotonic LiCl stop salt intake after 1 to 2 licks at the LiCl spout and move to the adjacent water spout within 0.7 s. Activity of 526 neurones from the nucleus of the solitary tract, gustatory thalamus, gustatory cortex, lateral and ventromedial thalamus, and amygdala was recorded in naive or CTA trained rats during the above gustatory discrimination. Post-stimulus histograms (PSH) triggered by water or salt licks or by spout switching were plotted for single units. Population responses of various regions were obtained by integration of the statistically significant excitatory and inhibitory intervals in the individual PSHs. Lick related changes of unit activity were orserved in 52% and 65% of neurones in control and CTA trained rats, respectively. The CTA training increased the incidence of units in which salt licking influenced the activity less than water licking. Presentation of the aversive fluid induced inhibition of unit activity in the gustatory cortex, ventromedial hypothalamus, and amygdala and excitation in the lateral hypothalamus. The changes started 100 to 150 ms after spout switching and culminated 100 ms later. Activity of the solitary tract nucleus and gustatory thalamus was affected less consistently. The results indicate that the gustatory cortex, amygdala and hypothalamus participate in CTA retrieval but a more specific identification of the electrical correlates of memory readout and of drinking control was not possible.     

Rapid post-oral stimulation of intake and flavor conditioning by glucose and fat in the mouse

Although widely assumed to have only satiating actions, nutrients in the gut can also condition increases in intake in some cases. Here we studied the time course of post-oral nutrient stimulation of ingestion in food-restricted C57BL/6J mice. In experiment 1, mice adapted to drink a 0.8% sucralose solution 1 h/day, rapidly increased their rate of licking (within 4-6 min) when first tested with an 8% glucose solution and even more so in tests 2 and 3. Other mice decreased their licking rate when switched from sucralose to 8% fructose, a sugar that is sweet like glucose but lacks positive post-oral effects in mice. The glucose-stimulated drinking is due to the sugar's post-oral rather than taste properties, because sucralose is highly preferred to glucose and fructose in brief choice tests. A second experiment showed that the glucose-stimulated ingestion is associated with a conditioned flavor preference in both intact and capsaicin-treated mice. This indicates that the post-oral stimulatory action of glucose is not mediated by capsaicin-sensitive visceral afferents. In experiment 3, mice consumed flavored saccharin solutions as they self-infused water or glucose via an intragastric (IG) catheter. The glucose self-infusion stimulated ingestion within 13-15 min in test 1 and produced a conditioned increase in licking that was apparent in the initial minute of tests 2 and 3. Experiment 4 revealed that IG self-infusions of a fat emulsion also resulted in post-oral stimulation of licking in test 1 and conditioned increases in tests 2 and 3. These findings indicate that glucose and fat can generate stimulatory post-oral signals early in a feeding session that increase ongoing ingestion and condition increases in flavor acceptance and preference revealed in subsequent feeding sessions. The test procedures developed here can be used to investigate the peripheral and central processes involved in stimulation of intake by post-oral nutrients.