Mechanical Conflict System: A Novel Operant Method for the Assessment of Nociceptive Behavior

A new operant test for preclinical pain research, termed the Mechanical Conflict System (MCS), is presented. Rats were given a choice either to remain in a brightly lit compartment or to escape to a dark compartment by crossing an array of height-adjustable nociceptive probes. Latency to escape the light compartment was evaluated with varying probe heights (0, .5, 1, 2, 3, and 4 mm above compartment floor) in rats with neuropathic pain induced by constriction nerve injury (CCI) and in naive control rats. Escape responses in CCI rats were assessed following intraperitoneal administration of pregabalin (10 and 30 mg/kg), morphine (2.5 and 5 mg/kg), and the tachykinin NK1 receptor antagonist, RP 67580 (1 and 10 mg/kg). Results indicate that escape latency increased as a function of probe height in both naive and CCI rats. Pregabalin (10 and 30 mg/kg) and morphine (5 mg/kg), but not RP 67580, decreased latency to escape in CCI rats suggesting an antinociceptive effect. In contrast, morphine (10 mg/kg) but not pregabalin (30 mg/kg) increased escape latency in naive rats suggesting a possible anxiolytic action of morphine in response to light-induced fear. No order effects following multiple test sessions were observed. We conclude that the MCS is a valid method to assess behavioral signs of affective pain in rodents.     

Masking by light in hamsters with SCN lesions

Inhibition of wheel running by light (masking) was investigated in Syrian hamsters with suprachiasmatic nucleus or sham lesions. Approximately 90% of the wheel revolutions made by hamsters with complete suprachiasmatic nucleus lesions, as judged by histology and power spectrum analysis of their wheel running, occurred during the dark phases of an ultradian light-dark cycle (3.5 h light, 3.5 h dark). This was demonstrated for two illumination levels (380 lx and 6 lx). Similar results were obtained with sham-operated animals. In further tests, the hamsters with lesions also retained a strong preference for the dark side of a box divided into dark and light sides. These results demonstrate that the suprachiasmatic nucleus is not necessary for masking by light or the preference for a dark over a light compartment. Evidently the direct effects of light can substitute for the endogenous control by the suprachiasmatic nucleus to maintain appropriate behaviour in time and space. Accepted: 30 January 1999     

Gender differences in development of exploratory activity in rats

The dynamics of exploratory activity in male and female Wistar rats before weaning and over the course of puberty was investigated. Behavior of animals was examined on the 18th, 35th, and 50th postnatal days in a modified light-dark test and on the 24th postnatal day in an open-field test. It was found that exploratory activity of males considerably changed over the course of puberty period. From the 18th to 35th postnatal days, the number of animals exploring a light compartment increased. From the 35th to 50th postnatal days, the duration of the light compartment investigation increased. The only change in exploratory activity of female rats consisted in an increase in the number of transitions between the light and dark compartments from 35th to 50th postnatal days. The results suggest different mechanisms of exploratory behavior development in male and female rats.     

Effects of Anesthetic Ketamine on Anxiety-Like Behaviour in Rats

There is scarce information regarding the effects of anesthetic doses of the non-competitive N-methyl-d-aspartate receptor antagonist ketamine on anxiety. The current study evaluated the acute effects of intraperitoneally (i.p.) administered anesthetic ketamine (100 mg/kg) i.p. on anxiety in rats. For this purpose, the light/dark and the open field tests were utilized. The effects of anesthetic ketamine on motility were also examined using a motility cage. In the light/dark test, anesthetic ketamine, administered 24 h before testing reduced the number of transitions between the light and dark compartments and the time spent in the light compartment in the rats compared with their control cohorts. In addition, ketamine was found to exert a depressive effect on rats’ motility. In the open field test, animals treated with anesthetic ketamine 24 h before testing spent essentially no time in the central area of the apparatus, decreased horizontal ambulatory activity, and preserved to a certain extent their exploratory behaviour compared to their control counterparts. The results suggest that, in spite of its hypokinetic effect, a single anesthetic ketamine administration apparently induces an anxiety-like state, while largely preserving exploratory behaviour in the rat. These effects were time-dependent they since they were extinguished when testing was carried out 48 h after anesthetic ketamine administration.

     

miR-194 relieve neuropathic pain and prevent neuroinflammation via targeting FOXA1

The emerging role of microRNAs (miRNAs) have been deeply explored in multiple diseases including neuropathic pain. miR-194 was widely reported to be a tumor suppressor and was related to the inflammatory response. The critical role of neuroinflammation on neuropathic pain leads to a thinking about the relationship between miR-194 and neuropathic pain. However, the function of miR-194 in neuropathic pain remains unknown. This study was aimed to explore the relationship between miR-194 and neuropathic pain progression by chronic sciatic nerve injury (CCI). miR-194 abnormally downregulated in the CCI model rat and its overexpression significantly alleviates neuroinflammation in vivo. We predict Forkhead box protein A1 (FOXA1) as a direct target of miR-194, whose restoration can markedly reverse the effects of miR-194 on neuropathic pain. Overall, our study demonstrated a novel mechanism of neuropathic pain progression that miR-194 alleviates neuropathic pain via targeting FOXA1 and preventing neuroinflammation by downregulating inflammatory cytokines containing cyclooxygenase 2, interleukin 6 (IL-6), and IL-10 in vivo, which can be reversed by the overexpression of FOXA1.     

Rat aversion to isoflurane versus carbon dioxide

Some experts suggest that sedation of laboratory rodents with isoflurane before euthanasia with carbon dioxide (CO₂) is a humane alternative to euthanasia with CO₂ alone, but little research has compared aversion with these agents. Albino rats were tested in a light–dark box where they had the choice between remaining in a dark compartment filling with isoflurane or CO₂, or escaping to a lit compartment. Experiment 1 validated the procedure by confirming that rats responded to agent and light intensity. In experiment 2, 9/16 and 0/16 rats remained in the dark compartment until recumbent when initially exposed to isoflurane and CO₂, respectively. In experiment 3, more rats remained in the dark compartment until recumbent during initial (10/16) versus re-exposure (1/16) to isoflurane. These results indicate that initial exposure to CO₂ is more aversive than isoflurane, and that re-exposure to isoflurane is more aversive than initial exposure. We conclude that sedation with isoflurane is a refinement over euthanasia with CO₂ alone for rats that have not been previously exposed to inhalant anaesthetics.     

What is the clinical relevance of different lung compartments?

The lung consists of at least seven compartments with relevance to immune reactions. Compartment 1 - the bronchoalveolar lavage (BAL), which represents the cells of the bronchoalveolar space: From a diagnostic point of view the bronchoalveolar space is the most important because it is easily accessible in laboratory animals, as well as in patients, using BAL. Although this technique has been used for several decades it is still unclear to what extent the BAL represents changes in other lung compartments. Compartment 2 - bronchus-associated lymphoid tissue (BALT): In the healthy, BALT can be found only in childhood. The role of BALT in the development of the mucosal immunity of the pulmonary surfaces has not yet been resolved. However, it might be an important tool for inhalative vaccination strategies. Compartment 3 - conducting airway mucosa: A third compartment is the bronchial epithelium and the submucosa, which both contain a distinct pool of leukocytes (e.g. intraepithelial lymphocytes, IEL). This again is also accessible via bronchoscopy. Compartment 4 - draining lymph nodes/Compartment 5 - lung parenchyma: Transbronchial biopsies are more difficult to perform but provide access to two additional compartments - lymph nodes with the draining lymphatics and lung parenchyma, which roughly means "interstitial" lung tissue. Compartment 6 - the intravascular leukocyte pool: The intravascular compartment lies between the systemic circulation and inflamed lung compartments. Compartment 7 - periarterial space: Finally, there is a unique, lung-specific space around the pulmonary arteries which contains blood and lymph capillaries. There are indications that this "periarterial space" may be involved in the pulmonary host defense. All these compartments are connected but the functional network is not yet fully understood. A better knowledge of the complex interactions could improve diagnosis and therapy, or enable preventive approaches of local immunization.     

New challenges in the study of the evolution of wild animals and their gut microbiome

In this viewpoint, by reviewing the recent findings on wild animals and their gut microbiomes, we found some potential new insights and challenges in the study of the evolution of wild animals and their gut microbiome. We suggested that wild animal gut microbiomes may come from microbiomes in the animals'' living habitats along with animals'' special behavior, and that the study of long‐term changes in gut microbiomes should consider both habitat and special behaviors. Also, host behavior would facilitate the gut microbiome transmission between individuals. We suggested that research should integrate the evolutionary history and physiological systems of wild animals to understand the evolution of animals and their gut microbiomes. Finally, we proposed the Noncultured‐Cultured‐Fermentation‐Model Animal pipeline to determine the function (diet digestion, physiology, and behavior) of these target strains in the wild animal gut.     

Metal Sources for Freshwater Invertebrates: Pertinence for Risk Assessment

Ecological risk assessments are likely to be more effective if they are built upon knowledge of from where and in what manner animals take up contaminants. We discuss the relative importance of various metal sources for aquatic invertebrates. First, we address the question do sediment-dwelling animals take up their metals from the overlying water compartment or the sediment compartment or both (both compartments include water and particles). We find that the overlying water column is more important as a metal source for insects, whereas the sediment compartment is more important for oligochaete worms. We explain this tendency by the behaviors of the animals involved. Second, we ask the question do animals take up their metals from food or water within a given compartment. Through case studies on three widespread freshwater invertebrates, we conclude that for some predatory insects food is their major source of several metals, whereas for the crustacean Hyalella both food and water appear to be important depending on the metal involved and the experimental protocol used to study the question. We conclude that ignoring food as a metal source could severely underestimate metal exposures for some animals. We suggest that integrating these complexities into laboratory tests and risk assessment protocols will improve their meaningfulness and thus their ability to protect aquatic ecosystems.     

A decreased frequency of peeking out from the dark compartment--the only constant index of the effect of anxiogens on the behavior of mice in a "light-darkness" chamber]

Special measurements of the effects of anxiolytics and anxiogens on the commonly used parameters of behavior of mice in a dark-light chamber (the rate of transitions and time spent in a dark and light compartments) demonstrated their low reproducibility in consecutive experiments. Therefore, these indices are not reliable (Lapin, 1992). One more parameter was tested in the present study. A decrease in the rate of leanings out of the dark compartment appeared to be a constant effect of standard anxiety-inducing drugs: caffeine, pentylenetetrazole, yohimbine, and a putative endogenous anxiogen phenylethylamine. Increase in the rate of leanings out, like increase in the rate of transitions and shortening of the time spent in a dark compartment produced by anxiolytics diazepam, chlordiazepoxide, hydroxyzine, phenibut and baclofen were not significant in the majority of experiments. That is why these effects of anxiolytics are not reliable for measuring the activity of anxiolytics in a dark-light chamber.     

Does evaluation of the ligamentous compartment enhance diagnostic utility of sacroiliac joint MRI in axial spondyloarthritis?

IntroductionInflammation of the sacroiliac joints (SIJ) is a fundamental clinical feature of axial spondyloarthritis (SpA). The anatomy of the irregularly shaped SIJ is complex with an antero-inferior cartilaginous compartment containing central hyaline and peripheral fibrocartilage, and a dorso-superior ligamentous compartment. Several scoring modules to systematically assess SIJ magnetic resonance imaging (MRI) in SpA have been developed. Nearly all of them are based on the cartilaginous joint compartment alone. However, there are only limited data about the frequency of inflammatory lesions in the ligamentous compartment and their potential diagnostic utility in axial SpA. We therefore aimed to evaluate the ligamentous compartment on sacroiliac joint MRI for lesion distribution and potential incremental value towards diagnosis of SpA over and above the traditional assessment of the cartilaginous compartment alone.MethodsTwo independent cohorts of 69 and 88 consecutive back pain patients ≤50 years were referred for suspected SpA (cohort A) or acute anterior uveitis plus back pain (cohort B). Patients were classified according to rheumatologist expert opinion based on clinical, radiographic and laboratory examination as having nonradiographic axial SpA (nr-axSpA; n = 51), ankylosing spondylitis (n = 34), or nonspecific back pain (NSBP; n = 72). Five blinded readers assessed SIJ MRI globally for presence/absence of SpA. Bone marrow edema (BME) and fat metaplasia were recorded in the cartilaginous and ligamentous compartment. The incremental value of evaluating the ligamentous additionally to the cartilaginous compartment alone for diagnosis of SpA was graded qualitatively. We determined the lesion distribution between the two compartments, and the impact of the ligamentous compartment evaluation on diagnostic utility.ResultsMRI bone marrow lesions solely in the ligamentous compartment in the absence of lesions in the cartilaginous compartment were reported in just 0–2.0/0–4.0 % (BME/fat metaplasia) of all subjects. Additional assessment of the ligamentous compartment was regarded as essential for diagnosis in 0 and 0.6 %, and as contributory in 28.0 and 7.7 % of nr-axSpA patients in cohorts A and B, respectively. Concomitant BME in both compartments was evident in 11.6–42.0 % of nr-axSpA and 2.1–2.4 % of NSBP patients.ConclusionAssessing the ligamentous compartment on SIJ MRI provided no incremental value for diagnosis of axial SpA. However, concomitant BME in both compartments may help discriminate nr-axSpA from NSBP.     

The Effects of Environmental Control on Cognition in Rats (Rattus norvegicus)

The aim of this study was to allow nonhuman animals to control their environment using operant conditioning procedures and to assess the effect of control on cognitive tasks. The study tested 4 predictions: (a) rats (Rattus norvegicus) will control a light stimulus; (b) animals will exhibit preferences for particular stimulus strengths; (c) animals who exert control over environmental stimuli will show improved performance on cognitive tasks compared with animals who lack control; and (d) at the end of the operant phase, experimental subjects will have lower corticosterone levels than animals who lack control. Experimental subjects did show control over a light stimulus and performed significantly better over time in a discrimination task compared with subjects who could not control their environment. There was no difference in corticosterone levels between control and experimental subjects. The results will both contribute to our understanding of how control of environmental stimuli affects the welfare of animals in captive environments and aid in designing experimental conditions that will increase validity and reliability in research.     

Rat behavior in a light/dark chamber: place preference task

After a 6-week social isolation (from 22nd to 70th day from birth) male Wistar rats (a sibship, 10 animals), were tested for 20 min in a light/dark box in order to reveal behavioral features of choice of their spatial localization in unknown conditions. Socially deprived rats significantly differed from control animals in longer time of the room choice, higher number of entries of the light section, and higher number of rearings, which was probably explained by their higher anxiety and lack of experience to estimate unexpected stimuli and select a response. The mean level of behavioral activity during exploration of the box defined by the number of elementary operations per minute remained constant and was significantly higher in the socially deprived rats than in the control animals.     

The effects of environmental control on cognition in rats (Rattus norvegicus)

The aim of this study was to allow nonhuman animals to control their environment using operant conditioning procedures and to assess the effect of control on cognitive tasks. The study tested 4 predictions: (a) rats (Rattus norvegicus) will control a light stimulus; (b) animals will exhibit preferences for particular stimulus strengths; (c) animals who exert control over environmental stimuli will show improved performance on cognitive tasks compared with animals who lack control; and (d) at the end of the operant phase, experimental subjects will have lower corticosterone levels than animals who lack control. Experimental subjects did show control over a light stimulus and performed significantly better over time in a discrimination task compared with subjects who could not control their environment. There was no difference in corticosterone levels between control and experimental subjects. The results will both contribute to our understanding of how control of environmental stimuli affects the welfare of animals in captive environments and aid in designing experimental conditions that will increase validity and reliability in research.     

Laughing Rats Are Optimistic

Emotions can bias human decisions- for example depressed or anxious people tend to make pessimistic judgements while those in positive affective states are often more optimistic. Several studies have reported that affect contingent judgement biases can also be produced in animals. The animals, however, cannot self-report; therefore, the valence of their emotions, to date, could only be assumed. Here we present the results of an experiment where the affect-contingent judgement bias has been produced by objectively measured positive emotions. We trained rats in operant Skinner boxes to press one lever in response to one tone to receive a food reward and to press another lever in response to a different tone to avoid punishment by electric foot shock. After attaining a stable level of discrimination performance, the animals were subjected to either handling or playful, experimenter-administered manual stimulation – tickling. This procedure has been confirmed to induce a positive affective state in rats, and the 50-kHz ultrasonic vocalisations (rat laughter) emitted by animals in response to tickling have been postulated to index positive emotions akin to human joy. During the tickling and handling sessions, the numbers of emitted high-frequency 50-kHz calls were scored. Immediately after tickling or handling, the animals were tested for their responses to a tone of intermediate frequency, and the pattern of their responses to this ambiguous cue was taken as an indicator of the animals'' optimism. Our findings indicate that tickling induced positive emotions which are directly indexed in rats by laughter, can make animals more optimistic. We demonstrate for the first time a link between the directly measured positive affective state and decision making under uncertainty in an animal model. We also introduce innovative tandem-approach for studying emotional-cognitive interplay in animals, which may be of great value for understanding the emotional-cognitive changes associated with mood disorders.     

Aluminum affects glial system and behavior of rats

Aluminum (Al) has been associated with neuronal dysfunction. These neuronal changes may involve glial alterations. We intend to evaluate the consequence of Al on the glial system and the behavior of rats exposed chronically to 0.3% of aluminum chloride in drinking water during 4 months in adulthood (A) or since intra-uterine age (IU); animals from this latter group were sacrificed at four months of age. Our data show an intense glial fibrillary acidic protein (GFAP)-immunoreactivity with a high density of astrocytes in both treated groups compared with controls. However, in IU rats, astrocytes display prominent glial cell bodies and processes. A and IU rat groups perform a significantly reduced locomotor activity. However, using the dark/light box test, the IU rats prefer to spend more time in the enlightened compartment compared to other groups. Behavioral and glial changes caused by Al exposure bring support for the role of Al in brain dysfunction involving glial alterations.     

Proton Gradients in Intact Cyanobacteria

The internal pH values of two unicellular cyanobacterial strains were determined with electron spin resonance probes, over an external pH range of 6 to 9, in the light and in the dark. The slow growing, thylakoid-lacking Gloeobacter violaceus was found to have a low capacity for maintaining a constant internal pH. The distribution pattern of weak acid and amine nitroxide spin probes across the cell membranes of this organism, in the light and in the dark, was consistent with the assumption that it contains a single intracellular compartment. At an external pH of 7.0, intracellular pH was 6.8 in the dark and 7.2 in the light. The cells of Agmenellum quadruplicatum, a marine species, were found to contain two separate compartments; in the dark, the pH of the cytoplasmic and the intrathylakoid spaces were calculated to be 7.2 and 5.5, respectively. Upon illumination, the former increased and the latter decreased by about 0.5 pH units.     

Period-Dependent Oscillatory Free-Run in the Locomotor Activity Rhythm of the Field Mouse Mus booduga Under Skeleton Photoperiodic Regimes

The entrainment behaviour of the circadian rhythm of locomotor activity in the field mouse Mus booduga was studied in order to evaluate the role of the animals' free-running period (τ) and the duration of skeleton photoperiods in determining entrainment of animals with τ values beyond and close to the “limits of entrainment”. We predicted that animals with τ lesser than the lower “limit of entrainment” would entrain only to short skeleton photoperiods (≤ 6 h) and not to longer skeleton photoperiods. Experimental animals (n = 25) were entrained to light/dark (LD) 12:12 h schedule, and then subjected to various skeleton photoperiods in which the duration of one of the two intervals of darkness was successively reduced while holding the zeitgeber period (T) constant. Some animals (n = 9) entrained to long as well as short photoperiods, whereas others (n = 5) entrained only to extremely short skeleton photoperiods of 6 h or less. The mean τ of the animals entraining to all photoperiods (23.78 ± 0.22 h) was significantly greater than that of the animals that entrained only to very short skeleton photoperiods (22.43 ± 0.41 h) (t _{df 12} = 5.3, p < 0.001). We also selected a few animals (n = 11) with average τ value of 23.13 ± 0.38 h and studied them under several skeleton photoperiods. To our surprise the animals which were subjected to restricted dark intervals invariably underwent “phase-jump” assuming the longer dark interval as “subjective night”. We suggest that the observed variation in entrainment behaviour might be due to the variation seen among individual animals in τ and the shape of their PRC. These results support the view that the duration of the skeleton photoperiod and the τ of an individual animal interact to determine its entrainment, and underscore the relevance of inter-individual variation in circadian organisation to studies of circadian rhythms.     

Efficient Lymphoreticular Prion Propagation Requires PrPc in Stromal and Hematopoietic Cells

In most prion diseases, infectivity accumulates in lymphoreticular organs early after infection. Defects in hematopoietic compartments, such as impaired B-cell maturation, or in stromal compartments, such as abrogation of follicular dendritic cells, can delay or prevent lymphoreticular prion colonization. However, the nature of the compartment in which prion replication takes place is controversial, and it is unclear whether this compartment coincides with that expressing the normal prion protein (PrP(c)). Here we studied the distribution of infectivity in splenic fractions of wild-type and fetal liver chimeric mice carrying the gene that encodes PrP(c) (Prnp) solely on hematopoietic or on stromal cells. We fractionated spleens at various times after intraperitoneal challenge with prions and assayed infectivity by bioassay. Upon high-dose challenge, chimeras carrying PrP(c) on hematopoietic cells accumulated prions in stroma and in purified splenocytes. In contrast, after low-dose challenge ablation of Prnp in either compartment prevented splenic accumulation of infectivity, indicating that optimal prion replication requires PrP(c) expression by both stromal and hematopoietic compartments.     

Specific subunits of voltage-gated sodium channel underlying the subthreshold membrane potential oscillations: Potential therapeutic targets for neuropathic pain

The treatment of neuropathic pain remains a major challenge to pain clinicians. Certain nociceptive and non-nociceptive dorsal root ganglion (DRG) neurons may develop abnormal spontaneous activities following peripheral nerve injury, which is believed to be a major contributor to chronic pain. Subthreshold membrane potential oscillation (SMPO) observed in injured DRG neurons was reported to be involved in the generation of abnormal spontaneous activity. Tetrodotoxin-sensitive sodium (Na⁺) channels were testified to be involved in the generation of SMPO, but their specific subunits have not been clarified. We hypothesize that the subunits of voltage-gated sodium channel, Na_v1.3 and Na_v1.6, are involved in the generation of SMPO. An attempt to test this hypothesis may lead to a new therapeutic strategy for neuropathic pain.