Induction of habits in rats by a forced-choice procedure in T-maze and the effect of pre-test free exploration

A forced-choice procedure in T-maze designed for the induction of habits was used to induce strong habits in rats. The response choices of rats in 20 free-choice trials were compared after the rats had been subjected to 1 or 200 forced-choice trials to one side of the T-maze. After 200 forced-choice trials the rats showed a significant (p < .001) propensity for the habitual arm of the maze in the subsequent free-choice trials. The habit was at least as pronounced when analysed over the last 10 free-choice trials as when it was analysed over the first 10 free-choice trials. When the rats were given the opportunity to explore the entire maze immediately before the free-choice challenge after 200 forced-choice trials, this resulted in a large variation in the choice pattern of the individual rats, and a subgroup of rats choose the newly opened maze arm in 95-100% of the 20 free-choice trials.     

Anxiolytic effects of short- and long-term administration of cacao mass on rat elevated T-maze test

We demonstrated the effects of short- and long-term administration of cacao mass on anxiety in the elevated T-maze test, which is an animal model of anxiety. In the first study, we administered cacao mass (100 mg/100 g body weight) per os and immediately performed the elevated T-maze test. Short-term cacao mass significantly abolished delayed avoidance latency compared with the control but did not change escape latency. This result suggested that cacao mass administration reduced conditional fear-relating behavior. Short-term cacao mass administration did not affect the concentration of brain monoamines, emotion-related neurotransmitters such as norepinephrine, serotonin and dopamine, in the rat brain. In the next study, we fed a cacao mass-containing diet to rats for 2 weeks and performed the elevated T-maze test. Contrary to short-term administration, chronic consumption of cacao mass tended to increase avoidance latency and did not change escape latency. Brain serotonin concentration and its turnover were enhanced by chronic consumption of cacao mass. These results suggested that chronic consumption of cacao did not affect fear-related behavior but was involved in brain monoamine metabolism. In conclusion, we suggest that short-term cacao mass consumption showed an anxiolytic effect but chronic consumption did not.     

Inhibitory avoidance memory retention in the elevated T-maze is impaired after perivascular manipulation of the common carotid arteries

Perivascular manipulation promoted by the positioning of a silicone collar around the common carotid arteries causes local inflammation and has been suggested as an animal model of atherosclerosis. This manipulation induces biochemical and morphological changes that are similar to those observed in the early stage of atherosclerosis in humans. Based on evidences showing that atherosclerosis is associated with cognitive deficits in humans, we presently investigated the temporal consequences of the bilateral positioning of silicone collars around the common carotid arteries (n = 15) on inhibitory avoidance memory retention in male Wistar rats tested in the elevated T-maze. The effects of this procedure were compared to those observed in sham-operated animals (n = 15) and to those observed in animals submitted to permanent bilateral occlusion of the common carotid arteries (n = 16). Additionally we studied the effects of the pretreatment with the non-selective anti-inflammatory drug indomethacin (n = 13) or the selective COX-2 inhibitor celecoxib (n = 12) and compared the effects to those of the pretreatment with vehicle (n = 11). The results showed that the silicone collar implants induced deficits in memory retention when animals were tested 2 and 4, but not 15 or 30, days after surgery. Permanent bilateral occlusion of the common carotid arteries impaired avoidance retention up to 30 days after surgery. Pretreatment with indomethacin (2 mg/kg/day) or celecoxib (5 mg/kg/day) post surgery and up to 3 days thereafter did not prevent memory deficits caused by silicone collar implants. Our data suggest that the prostanoids that participate in the inflammatory process triggered by the placement of the silicone collar do not seem responsible for the deficit in memory retention observed during the first days after collar placement.     

Influence of the estrous cycle on the behavior of rats in the elevated T-maze

The elevated T-maze is an animal anxiety model which can discriminate between anxiety-like and fear-like behaviors. The estrous cycle is an important variable of the response in animal anxiety tests and is known to affect other models. The aim of the present study was to investigate the influence of the estrous cycle on behavior displayed in the elevated T-maze test. Seventeen male and 60 female rats were submitted to one session in this test, with the females being screened for the estrous cycle and divided into groups according to the various phases. The elevated T-maze had three arms of equal dimensions ( 50 cm x 10 cm), one enclosed by 40-cm high walls and perpendicular to the others, the apparatus being elevated 50 cm above the floor. Each rat was placed in the end of the enclosed arm and the latency for it to leave this arm was recorded. These measurements were repeated three times separated by 30-s intervals (passive avoidance). After trial 3, each rat was placed at the distal end of the right open arm and the latency to exit this arm was recorded. Whenever latencies were greater than 300 s the trial was finished. The results demonstrated females in diestrus exhibited anxiety-like behaviors while females in metaestrus behaved in a similar way as the males. There were no differences between groups in fear-like behaviors. The results also indicate the elevated T-maze to be a sensitive test to measure anxiety.     

Teratogenic effects of diazepam in the hamster.

Pregnant hamsters were treated with different doses of oral and intravenous diazepam during the period of organogenesis. Teratogenic effects of diazepam were observed following oral treatment on days 8 and 10 and following intravenous treatment on day 11 of gestation. Types of malformations included cleft palate, exencephaly, limb anomalies, and hemorrhage. A dose-effect relationship was not observed. Comparison with reported literature seems to indicate that diazepam may be a mild teratogen in some species.     

Solid-phase synthesis of neurokinin A antagonists. Comparison of the Boc and Fmoc methods.

During the preparation of the NK-2 selective tachykinin antagonist MEN 10208 (Thr-Asp-Tyr-D-Trp-Val-D-Trp-D-Trp-Arg-NH2) and its analogs by the solid-phase method employing the Boc strategy routinely used in our laboratory, we encountered difficulties in the coupling of hydrophobic amino acids D-Trp and Val. To study the coupling problems several syntheses of MEN 10208 and analogs were carried out with different activation strategies. These syntheses yielded considerable amounts of deletion sequences even though a negative Kaiser test was obtained after each coupling. Inaccessibility of the free amino group of the growing peptide due to steric hindrance of the hydrophobic residues during coupling, and for the ninhydrin complex during the Kaiser test, may account, at least in part, for the unsatisfactory synthetics results and for the false-negative ninhydrin tests. Repetition of each synthesis with the Fmoc strategy on a newly developed DOD resin for peptide amides using the DCC/HOBt chemistry gave superior results in terms of the yield and purity of the crude peptides. Therefore, the Fmoc strategy appears to offer advantages over the Boc method for the preparation of these peptides containing hydrophobic amino acids.     

Comparison of effects of decapitation and anesthesia on metabolic and hormonal parameters in Sprague-Dawley rats.

The modes of euthanasia by either anesthesia or by decapitation were compared by assessing several metabolic and hormonal parameters from plasma and hormone receptors from liver plasma membranes. Two different anesthetics were used. Compared to decapitation, euthanasia by anesthesia significantly increased plasma glucose and triglyceride levels but not plasma cholesterol. Plasma insulin was also significantly increased by anesthetics. No significant differences were observed in plasma glucagon levels or insulin and glucagon receptors from liver plasma membranes between rats euthanized by decapitation and anesthesia. Glucagon receptors were however, affected by dietary carbohydrates. It is concluded that in studies involving measurements of metabolic and hormonal parameters, the use of anesthesia is to be avoided for euthanasia and that decapitation should be the method of choice.     

In silico exploration of the effects of host genotype and nutrition on the genetic parameters of lambs challenged with gastrointestinal parasites

An in silico mathematical model was used to explore the effect of, and the interaction between, (i) nutrition, (ii) genotype for growth and (iii) genotype for resistance, on the estimates of genetic parameters for resistance and performance in a population of lambs trickle-challenged daily with 3,000 L3s of Teladorsagia circumcincta. A previously published model for nematode infections in sheep was developed to include heritable variation in sheep growth traits, as well as in immunologically controlled traits such as establishment of incoming larvae, mortality of the adult worms and fecundity of the adult female worms. The simulated population comprised 10,000 lambs, these being the offspring of 250 sires mated to 5,000 dams. The model assumed the lambs to be parasitologically naïve at weaning (2 months of age), at which point the trickle challenge commenced and the model was updated daily until slaughter (at 6 months of age). Dietary treatments included a good and a poor quality feed, offered ad libitum. Two genotypes for growth were assumed: (i) fast and (ii) slow growing. Three genotypes for resistance were used: (i) benchmark, (ii) susceptible and (iii) resistant, differing in their ability to cope with nematode infections. Genetic parameters for output traits, including growth rate, food intake, worm burden and faecal egg count were estimated using a linear mixed model, fitting sire as a random effect to capture genetic effects. Heritabilities and correlations were found to change over time. In general, the heritabilities of immunity traits increased over time, whereas genetic correlations between production and immunity traits became weaker. Diet had a significant effect on the means and the estimated correlations of output traits, while genotypes for growth and resistance had smaller effects. These results suggest that discrepancies between published genetic parameters for nematode resistance may be a function of environmental factors rather than differences in host genotype.     

Memorization of contextual and CS conditioned fear response (freezing) in a one-trial acquisition paradigm.

In fear-conditioned Wistar rats freezing was induced by the delivery of a series of footshocks paired to tones (CS) in a specific conditioning chamber (context). CS and contextual fear were acquired in the same single conditioning session without preexposition to the conditioning chamber (day 1). Different groups of animals were conditioned employing three increasing US (footshock) intensities (0.25, 0.5, 0.75 mA). During the retention sessions context and CS conditioned freezing (fear response) were measured using a paradigm that fulfilled the following conditions: i) CS freezing retention was measured in a context different from the conditioning one; ii) CS and context freezing were measured at increased delays after the training session (days 3 and 4, 14 and 15, 28 and 29). The results show that there are significant differences between CS and context freezing retention, which are clearly related to delay after the initial session and to US intensity. In particular: 1) conditioned freezing to a discrete tone is better retained than conditioned freezing to context (irrespective of US intensity); 2) context freezing is directly related to US intensity much more than to tone freezing; 3) context freezing is easier to extinguish than tone freezing. The results are discussed in relation to previous ones and to their relevance to freezing genesis neural correlates.     

Combined effects of elevated temperatures and reduced leaf litter quality on the life-history parameters of a saprophagous macroarthropod

Because soil macroinvertebrates strongly modify decomposition processes, it is important to know how their abundance will respond to global change. We investigated in laboratory microcosms, the effects of elevated temperatures and reduced leaf litter quality on the life‐history traits of a saprophagous macroarthropod (development time, growth, survival and reproduction). Millipedes (Polydesmus angustus) from an Atlantic temperate forest were reared throughout their life cycle (≥16 months) under two temperature regimes differing on average by 3.3 °C; in a factorial design, they were fed either on Atlantic leaf litter or on Mediterranean leaf litter with a higher C : N ratio; humidity was consistently high. The components of the population growth rate (r) were affected positively by the temperature rise and negatively by the switch from Atlantic to Mediterranean leaf litter. When both treatments were combined, litter effects offset temperature effects. These results show that the short‐term response of saprophagous macroarthropods to warming is positive but depends on the availability of high‐quality litter, which is difficult to predict in the global change context. In a parallel experiment, conspecific millipedes from a Mediterranean population, which have evolved for a long time in a warmer climate and on poor‐quality litter, were reared at elevated temperatures on Mediterranean leaf litter. All components of r were higher than in the Atlantic population under the same conditions. This suggests that in the longer term, macroarthropods can overcome detrimental trophic interactions. Based on our study and the literature, we conclude that for decades the positive effects of warming on saprophagous macrofauna should exceed the negative effects of changes in litter quality. The abundance of those organisms in temperate forests could increase, which is confirmed by latitudinal patterns in Europe. Studies aimed at predicting the impacts of global change on decomposition will need to consider interactions with soil macroinvertebrates.     

Effect of kassinin, neurokinin A and neurokinin B on drinking behaviour in the pigeon

Intracerebroventricular (i.c.v.) injection of kassinin produced a prompt and copious drinking response at doses of 10-1000 ng/pigeon, in the absence of other behavioural alterations or of changes in core temperature. Neurokinin A and B evoked drinking, but they were respectively 10 and 100 times less potent than kassinin. Intraperitoneal injection of kassinin elicited drinking, but at doses about 1000 X larger than the i.c.v. ones. The angiotensin antagonist [Sar1, Leu8]angiotensin II did not reduce drinking induced by i.c.v. kassinin, suggesting that its effect is not due to interaction with the central renin-angiotensin system. Moreover, the effect is apparently independent of the mechanisms controlling hypovolaemic and hyperosmotic thirst since exact additivity was found in the dipsogenic response when i.c.v. kassinin was administered in the presence of a hypovolaemic (subcutaneous (s.c.), polyethylene glycol) or hyperosmotic (s.c. hypertonic NaCl) dipsogenic stimulus. The present findings show that kassinin, neurokinin A and B share with the tachykinins already tested (eledoisin, physalaemin, substance P) a common dipsogenic action in pigeons. However, marked differences exist in their dipsogenic potency. This order of potency, eledoisin = kassinin = physalaemin greater than neurokinin A = substance P greater than neurokinin B, is not consistent with the tachykinin receptor subtypes so far proposed.     

Effects of alcoholic beverage treatment on spatial learning and fear memory in mice

Although chronic ethanol treatment is known to impair learning and memory, humans commonly consume a range of alcoholic beverages. However, the specific effects of some alcoholic beverages on behavioral performance are largely unknown. The present study compared the effects of a range of alcoholic beverages (plain ethanol solution, red wine, sake and whiskey; with a matched alcohol concentration of 10%) on learning and memory. 6-week-old C57BL6J mice were orally administered alcohol for 7 weeks. The results revealed that red wine treatment exhibited a trend toward improvement of spatial memory and advanced extinction of fear memory. Additionally, red wine treatment significantly increased mRNA levels of brain-derived neurotrophic factor (BDNF) and N-methyl-D-aspartate (NMDA) receptors in mice hippocampus. These results support previous reports that red wine has beneficial effects.     

Absence of the SP/SP receptor circuitry in the substance P-precursor knockout mice or SP receptor, neurokinin (NK)1 knockout mice leads to an inhibited cytokine response in granulomas associated with murine Taenia crassiceps infection

Neurocysticercosis, caused by the cestode Taenia solium, is the most common parasitic infection of the human central nervous system that leads to seizures. Taenia crassiceps cysticercosis in mice is an experimental model for Taenia solium cysticercosis. Similar to the human infection, live parasites cause little or no granulomatous inflammation. Dying parasites initiate a granulomatous reaction. The neuropeptide, substance P (SP), stimulates T-helper (TH) 1 cytokine production. In the current studies, we determined whether absence of SP/SP receptor circuitry in the SP-precursor, preprotachykinin, knockout or SP-receptor, neurokinin (NK) 1, knockout mice affected granuloma cytokine production. We hence compared the levels of Th1 cytokines interleukin (IL)-2 and interferon (IFN)-gamma, and levels of Th2/immunoregulatory cytokines IL-4 and IL-10, by enzyme-linked immunosorbent assay in T. crassiceps-induced granulomas derived from infected C57BL/6 wild type (WT) versus SP-precursor knockout and NK1 knockout mice. We found that mean levels of IL-2, IFN-gamma, IL-4, and IL-10 in infected WT-derived granulomas were significantly higher than those of granulomas derived from infected SP-precursor knockout or the NK1 receptor (NKIR)knockout mice. Levels of Th2/immunoregulatory cytokines, IL-4 and IL-10 were higher in early stage granulomas (histologically-staged on basis of evidence of parasite remnants) versus late stage granulomas (no parasite-remnants) of both knockouts, whereas the reverse was noted in WT-derived granulomas. These study established that the absence of an SP/SP receptor circuitry in the SP precursor knockout mice or NK1 receptor knockout mice led to an inhibited cytokine response.     

Nutritional parameters modify muricidal behavior of male Wistar rats: preventive effects of amino acids and 4' Cl diazepam.

In male Wistar rats fed a diet enriched in polyunsaturated fatty acids and starch (PUFA+S), the percentage of muricidal (Mu) rats increased to 82% within 60 days. Mu rats had higher serum triglyceride levels and lower cholesterol levels than non-Mu rats. Water intake decreased in all rats on the PUFA+S diet concurrently with the increase in the proportion of Mu rats; protracted water restriction in rats fed standard diet also increased the percentage of Mu rats. In the offspring of two Wistar females fed the PUFA+S diet, the proportion of young Mu rats was 67%. When the PUFA+S diet was replaced with standard diet, the induced Mu behavior was not reversed. PK11195 (6 mg/kg i.p.), clonazepam (0.2 mg/kg i.p.), and flumazenil (15 mg/kg i.p.) were ineffective in reversing the induced Mu behavior, whereas 4'-chlorodiazepam (5 mg/kg i.p.) or muscimol (0.5 mg/kg i.p.) caused reversals of 63% or 50%, respectively. A 5-hydroxytryptophan overload (60 mg/kg i.p.) also reversed Mu behavior by 71%. All reversal effects were temporary. Pretreatment with yeast for 7 days before the PUFA+S diet was given prevented induction for more than 90 days on the PUFA+S diet, while similar pretreatment 4'Cl-diazepam resulted in 71% prevention of induction. The results are analyzed in terms of the involvement of endozepin, vasopressin, and serotonin receptors, and of possible genetic parameters.     

Comparison of the effects of 3,5,3'-triiodothyroacetic Acid and triiodothyronine on goiter prevention and involution and on hepatic and skeletal parameters in rats.

3,5,3'-triiodothyroacetic acid (TRIAC) has been used to suppress pituitary TSH secretion with reported attenuation of extrapituitary effects. We investigated whether equivalent doses of T (3) and TRIAC preventing the induction of goiter by methimazole (MMI) had a different or similar impact on peripheral tissues, such as liver and bone. In particular, we compared the effects of both compounds on the activity of the hepatic thyroid hormone-responsive enzymes, malic enzyme and L-glicerol-3-P dehydrogenase; bone mineral density and biochemical parameters of bone turnover, such as bone alkaline phosphatase (b-ALP) and the carboxy-terminal telopeptide region of type I collagen (beta-CTX); and the activity of thyroid ornithine decarboxylase (ODC). We also compared the effects of T (3) and TRIAC on the involution of MMI-induced goiter. Our results showed that TRIAC was more effective than T (3) to reduce MMI-induced goiter in a short-term goiter involution assay. TRIAC increased hepatic enzymes activity and beta-CTX levels, a parameter of bone resorption, more than T (3). However, bone mineral density was not altered by either treatment. Both compounds even reduced ODC activity at doses that were not effective at the pituitary level. These results demonstrate increased TRIAC hepatic and antigoitrogenic activity compared to T (3). TRIAC induces an imbalance in bone remodeling without affecting bone mineral density. Further studies are required to clarify this point.     

Effect of GABA-negative preparations on the anxiolytic and sedative effects of diazepam

Experiments on rats have shown that bicuculline (2 mg/kg) and picrotoxin (2 mg/kg) abolish the anxiolytic action of diazepam (2.5 mg/kg). Bicuculline (2 and 4 mg/kg) decreases while picrotoxin transforms the sedative effect of diazepam to the anxiolytic one. Picrotoxin (2 mg/kg) reduces the sedative action of gamma-acetylenic GABA (100 mg/kg) but does not favour the manifestation of its anxiolytic effect. It is suggested that the GABA-ergic mechanisms play an important role in the sedative effect of diazepam.     

Comparison of the effects of the subjective complexity and verbal creativity on EEG spectral power parameters

The EEG recording was made when the subjects performed tasks that involved overcoming the stereotype (creative) and retrieving information from memory (noncreative) with the usual and complicated presentation of the initial material (incomplete proverbs and sayings without concluding words). The subjective complexity of the task performance under different conditions was assessed. The EEG power from 19 EEG derivations was compared in the β₂ and γ frequency bands. The creative task performance was associated with a marked increase in the EEG power; significantly more complicated noncreative tasks were not accompanied by marked changes in the EEG power in these bands.     

Bi-directional effect of cholecystokinin receptor-2 overexpression on stress-triggered fear memory and anxiety in the mouse

Fear, an emotional response of animals to environmental stress/threats, plays an important role in initiating and driving adaptive response, by which the homeostasis in the body is maintained. Overwhelming/uncontrollable fear, however, represents a core symptom of anxiety disorders, and may disturb the homeostasis. Because to recall or imagine certain cue(s) of stress/threats is a compulsory inducer for the expression of anxiety, it is generally believed that the pathogenesis of anxiety is associated with higher attention (acquisition) selectively to stress or mal-enhanced fear memory, despite that the actual relationship between fear memory and anxiety is not yet really established. In this study, inducible forebrain-specific cholecystokinin receptor-2 transgenic (IF-CCKR-2 tg) mice, different stress paradigms, batteries of behavioral tests, and biochemical assays were used to evaluate how different CCKergic activities drive fear behavior and hormonal reaction in response to stresses with different intensities. We found that in IF-CCKR-2 tg mice, contextual fear was impaired following 1 trial of footshock, while overall fear behavior was enhanced following 36 trials of footshock, compared to their littermate controls. In contrast to a standard Yerkes-Dodson (inverted-U shaped) stress-fear relationship in control mice, a linearized stress-fear curve was observed in CCKR-2 tg mice following gradient stresses. Moreover, compared to 1 trial, 36 trials of footshock in these transgenic mice enhanced anxiety-like behavior in other behavioral tests, impaired spatial and recognition memories, and prolonged the activation of adrenocorticotropic hormone (ACTH) and glucocorticoids (CORT) following new acute stress. Taken together, these results indicate that stress may trigger two distinctive neurobehavioral systems, depending on both of the intensity of stress and the CCKergic tone in the brain. A "threshold theory" for this two-behavior system has been suggested.     

An isobolographic analysis of the hypnotic effects of combinations of dexmedetomidine with fentanyl or diazepam in rats.

The effects of the combinations of dexmedetomidine-fentanyl and dexmedetomidine-diazepam on the righting reflex were studied in rats. The doses that block the righting reflex for the agents given alone and for their combinations were determined with a probit procedure and compared with an isobolographic analysis. The interactions between dexmedetomidine and fentanyl or diazepam were found to be synergistic. In the dexmedetomidine-diazepam combination studies, less than one-fourth of the single drug dose (for each of two agents) was needed to produce the required effect. These data confirm synergistic interactions between dexmedetomidine and fentanyl or diazepam in producing hypnotic-anesthetic action.