Effect of gamma irradiation on the behavioral properties of crotoxin

Crotoxin has been detoxified with gamma radiation in order to improve crotalic antiserum production. Nevertheless, present knowledge of the biological characteristics of irradiated crotoxin is insufficient to propose it as an immunizing agent. Crotoxin is known to increase the emotional state of rats and to decrease their exploratory behavior (Moreira EG, Nascimento N, Rosa GJM, Rogero JR and Vassilieff VS (1996) Brazilian Journal of Medical and Biological Research, 29: 629-632). Therefore, we decided 1) to evaluate the effects of crotoxin in the social interaction test, which has been widely used for the evaluation of anxiogenic drugs, and 2) to determine if irradiated crotoxin induces behavioral alterations similar to those of crotoxin in the social interaction, open-field and hole-board tests. Male Wistar rats (180-220 g) were used. Crotoxin (100, 250, and 500 µg/kg) was injected intraperitoneally 2 h before the social interaction test. Similarly, irradiated crotoxin (2000 Gy gamma radiation from a 60Co source) was administered at the doses of 100, 250, and 500 µg/kg for the hole-board test, and at the doses of 1000 and 2500 µg/kg for the open-field and social interaction tests. ANOVA complemented with the Dunnett test was used for statistical analysis (P<0.05). Crotoxin decreased the social interaction time (s) at the doses of 100, 250 and 500 µg/kg (means ± SEM) from 51.6 ± 4.4 to 32.6 ± 3.7, 28.0 ± 3.6 and 31.6 ± 4.4, respectively. Irradiated crotoxin did not induce behavioral alterations. These results indicate that 1) crotoxin may be an anxiogenic compound, and 2) in contrast to crotoxin, irradiated crotoxin was unable to induce behavioral alterations, which makes it a promising compound for the production of crotalic antiserum

Effects of entorhinal cortex lesions on memory in different tasks

Lesions of the entorhinal cortex produce retrograde memory impairment in both animals and humans. Here we report the effects of bilateral entorhinal cortex lesions caused by the stereotaxic infusion of N-methyl-D-aspartate (NMDA) in rats at two different moments, before or after the training session, on memory of different tasks: two-way shuttle avoidance, inhibitory avoidance and habituation to an open field. Pre- or post-training entorhinal cortex lesions caused an impairment of performance in the shuttle avoidance task, which agrees with the previously described role of this area in the processing of memories acquired in successive sessions. In the inhibitory avoidance task, only the post-training lesions had an effect (amnesia). No effect was observed on the open field task. The findings suggest that the role of the entorhinal cortex in memory processing is task-dependent, perhaps related to the complexity of each task

Developmental and behavioral effects of postnatal amitraz exposure in rats

The effects of postnatal amitraz exposure on physical and behavioral parameters were studied in Wistar rats, whose lactating dams received the pesticide (10 mg/kg) orally on days 1, 4, 7, 10, 13, 16 and 19 of lactation; control dams received distilled water (1 ml/kg) on the same days. A total of 18 different litters (9 of them control and 9 experimental) born after a 21-day gestation were used. The results showed that the median effective time (ET50) for fur development, eye opening, testis descent and onset of the startle response were increased in rats postnatally exposed to amitraz (2.7, 15.1, 21.6 and 15.3 days, respectively) compared to those of the control pups (1.8, 14.0, 19.9 and 12.9 days, respectively). The ages of incisor eruption, total unfolding of the external ears, vaginal and ear opening and the time taken to perform the grasping hindlimb reflex were not affected by amitraz exposure. Pups from dams treated with amitraz during lactation took more time (in seconds) to perform the surface righting reflex on postnatal days (PND) 3 (25.0 ± 2.0), 4 (12.3 ± 1.2) and 5 (8.7 ± 0.9) in relation to controls (10.6 ± 1.2; 4.5 ± 0.6 and 3.4 ± 0.4, respectively); the climbing response was not changed by amitraz. Postnatal amitraz exposure increased spontaneous motor activity of male and female pups in the open-field on PND 16 (140 ± 11) and 17 (124 ± 12), and 16 (104 ± 9), 17 (137 ± 9) and 18 (106 ± 8), respectively. Data on spontaneous motor activity of the control male and female pups were 59 ± 11 and 69 ± 10 for days 16 and 17 and 49 ± 9, 48 ± 7 and 56 ± 7 for days 16, 17 and 18, respectively. Some qualitative differences were also observed in spontaneous motor behavior; thus, raising the head, shoulder and pelvis matured one or two days later in the amitraz-treated offspring. Postnatal amitraz exposure did not change locomotion and rearing frequencies or immobility time in the open-field on PND 30, 60 and 90. The present findings indicate that postnatal exposure to amitraz caused transient developmental and behavioral changes in the exposed offspring and suggest that further investigation of the potential health risk of amitraz exposure to developing human and animal offsprings may be warranted.

Signaled two-way avoidance learning using electrical stimulation of the inferior colliculus as negative reinforcement: effects of visual and auditory cues as warning stimuli

The inferior colliculus is a primary relay for the processing of auditory information in the brainstem. The inferior colliculus is also part of the so-called brain aversion system as animals learn to switch off the electrical stimulation of this structure. The purpose of the present study was to determine whether associative learning occurs between aversion induced by electrical stimulation of the inferior colliculus and visual and auditory warning stimuli. Rats implanted with electrodes into the central nucleus of the inferior colliculus were placed inside an open-field and thresholds for the escape response to electrical stimulation of the inferior colliculus were determined. The rats were then placed inside a shuttle-box and submitted to a two-way avoidance paradigm. Electrical stimulation of the inferior colliculus at the escape threshold (98.12 ± 6.15 (A, peak-to-peak) was used as negative reinforcement and light or tone as the warning stimulus. Each session consisted of 50 trials and was divided into two segments of 25 trials in order to determine the learning rate of the animals during the sessions. The rats learned to avoid the inferior colliculus stimulation when light was used as the warning stimulus (13.25 ± 0.60 s and 8.63 ± 0.93 s for latencies and 12.5 ± 2.04 and 19.62 ± 1.65 for frequencies in the first and second halves of the sessions, respectively, P<0.01 in both cases). No significant changes in latencies (14.75 ± 1.63 and 12.75 ± 1.44 s) or frequencies of responses (8.75 ± 1.20 and 11.25 ± 1.13) were seen when tone was used as the warning stimulus (P>0.05 in both cases). Taken together, the present results suggest that rats learn to avoid the inferior colliculus stimulation when light is used as the warning stimulus. However, this learning process does not occur when the neutral stimulus used is an acoustic one. Electrical stimulation of the inferior colliculus may disturb the signal transmission of the stimulus to be conditioned from the inferior colliculus to higher brain structures such as amygdala

Effect of repeated restraint stress on memory in different tasks

The present study investigated the effect of repeated stress applied to female rats on memory evaluated by three behavioral tasks: two-way shuttle avoidance, inhibitory avoidance and habituation to an open field. Repeated stress had different effects on rat behavior when different tasks were considered. In the two-way active avoidance test the stressed animals presented memory of the task, but their memory scores were impaired when compared to all other groups. In the habituation to the open field, only the control group showed a significant difference in the number of rearings between training and testing sessions, which is interpreted as an adequate memory of the task. In the handled and chronically stressed animals, on the other hand, no memory was observed, suggesting that even a very mild repeated stress would be enough to alter habituation to this task. The performance in the inhibitory avoidance task presented no significant differences between groups. The findings suggest that repeated restraint stress might induce cognitive impairments that are dependent on the task and on stress intensity.

Defense reaction induced by a metabotropic glutamate receptor agonist microinjected into the dorsal periaqueductal gray of rats

The behavioral effects of trans-(±)-1-amino-1,3-cyclopentanedicarboxylic acid (t-ACPD), a metabotropic glutamate receptor (mGluR) agonist, or 0.9% (w/v) saline, injected into the dorsal periaqueductal gray (DPAG), was investigated. Male Wistar rats showed defense reactions characterized by jumps toward the top edges of the cages (saline = 0 vs t-ACPD = 6.0, medians P<0.05) and gallops (saline = 0 vs t-ACPD = 10.0, medians P<0.05) during the 60-s period after the beginning of the injection. In another experiment animals were placed inside an open arena for 5 min immediately after injection. Their behavior was recorded by a video camera and a computer program analyzed the videotapes. Eleven of fifteen rats injected with t-ACPD showed a short-lasting (about 1 min) flight reaction. No saline-treated animal showed this reaction (P<0.0005, chi-square test). The drug induced an increase in turning behavior (P = 0.002, MANOVA) and a decrease in the number of rearings (P<0.001, MANOVA) and grooming episodes (P<0.001, MANOVA). These results suggest that mGluRs play a role in the control of defense reactions in the DPAG.

Antidepressant-like effect of lead in adult mice

It has been reported that lead can cause behavioral impairment by inhibiting the N-methyl-D-aspartate (NMDA) receptor complex. MK-801, a noncompetitive NMDA receptor antagonist, exhibits an antidepressant-like action in the forced swimming test. The purpose of the present study was to determine whether subacute lead exposure in adult male Swiss mice weighing 30-35 g causes an antidepressant-like action in a forced swimming test. Mice were injected intraperitoneally (ip) with 10 mg/kg lead acetate or saline daily for 7 consecutive days. Twenty-four hours after the last treatment, the saline and lead-treated mice received an injection of MK-801 (0.01 mg/kg, ip) or saline and were tested in forced swimming and in open-field tests. Immobility time was similarly reduced in the saline-MK-801, Pb-saline and Pb-MK-801 groups compared to the saline-saline group (mean ± SEM; 197.3 ± 18.5, 193.5 ± 15.8, 191.3 ± 12.3 and 264.0 ± 14.4 s, respectively; N = 9). These data indicate that lead may exert its effect on the forced swimming test by directly or indirectly inhibiting the NMDA receptor complex. Lead treatment caused no deficit in memory of habituation and did not affect locomotor activity in an open-field (N = 14). However, mice that received MK-801 after lead exhibited a deficit in habituation (22% reduction in rearing responses between session 3 and 1; N = 14) as compared to control (41% reduction in rearing responses; N = 15), further suggesting that lead may have affected the NMDA receptor activity. Forced-swim immobility in a basin in two daily consecutive sessions was also significantly decreased by lead exposure (mean ± SEM; day 1 = 10.6 ± 3.2, day 2 = 19.6 ± 3.6; N = 16) as compared to control (day 1 = 18.4 ± 3.8, day 2 = 34.0 ± 3.7; N = 17), whereas the number of crossings was not affected by lead treatment, further indicating a specific antidepressant-like action of lead.

Acute buspirone abolishes the expression of behavioral dopaminergic supersensitivity in mice

Previous studies have shown that rats withdrawn from long-term treatment with dopamine receptor blockers exhibit dopaminergic supersensitivity, which can be behaviorally evaluated by enhanced general activity observed in an open-field. Recently, it has been reported that co-treatment with the non-benzodiazepine anxiolytic buspirone attenuates the development of haloperidol-induced dopaminergic supersensitivity measured by open-field behavior of rats. The aims of the present study were: 1) to determine, as previously reported for rats, if mice withdrawn from long-term neuroleptic treatment would also develop dopaminergic supersensitivity using open-field behavior as an experimental paradigm, and 2) to examine if acute buspirone administration would attenuate the expression of this behavioral dopaminergic supersensitivity. Withdrawal from long-term haloperidol treatment (2.5 mg/kg, once daily, for 20 days) induced a significant (30%) increase in ambulation frequency (i.e., number of squares crossed in 5-min observation sessions) but did not modify rearing frequency or immobility duration in 3-month-old EPM-M1 male mice observed in the open-field apparatus. Acute intraperitoneal injection of buspirone (3.0 and 10 but not 1.0 mg/kg, 12-13 animals per group) 30 min before open-field exposure abolished the increase in locomotion frequency induced by haloperidol withdrawal. These data suggest that the open-field behavior of mice can be used to detect dopaminergic supersensitivity, whose expression is abolished by acute buspirone administration.

Effects of elevated calcium on motor and exploratory activities of rats

The effects of serum and brain calcium concentration on rat behavior were tested by maintaining animals on either distilled water (N = 60) or water containing 1% calcium gluconate (N = 60) for 3 days. Animals that were maintained on high calcium drinking water presented increased serum calcium levels (control = 10.12 ± 0.46 vs calcium treated = 11.62 ± 0.51 µg/dl). Increase of brain calcium levels was not statistically significant. In the behavioral experiments each rat was used for only one test. Rats that were maintained on high calcium drinking water showed increased open-field behavior of ambulation (20.68%) and rearing (64.57%). On the hole-board, calcium-supplemented animals showed increased head-dip (67%) and head-dipping (126%), suggesting increased ambulatory and exploratory behavior. The time of social interaction was normal in animals maintained on drinking water containing added calcium. Rats supplemented with calcium and submitted to elevated plus-maze tests showed a normal status of anxiety and elevated locomotor activity. We conclude that elevated levels of calcium enhance motor and exploratory behavior of rats without inducing other behavioral alterations. These data suggest the need for a more detailed analysis of several current proposals for the use of calcium therapy in humans, for example in altered blood pressure states, bone mineral metabolism disorders in the elderly, hypocalcemic states, and athletic activities.

Dose-related effects of propericiazine in rats

We evaluated the effects of the neuroleptic agent propericiazine on animal models of anxiety and memory. Adult male Wistar rats (250 to 350 g) received intraperitoneal injections of propericiazine (0.05, 0.075 and 0.1 mg/kg), diazepam (1 mg/kg), saline, or diazepam vehicle (20% propylene glycol and 80% saline) 30 min prior to the experimental procedure. Animals (10-15 for each task) were tested for step-down inhibitory avoidance (0.3-mA footshock) and habituation to an open-field for memory assessment, and submitted to the elevated plus-maze to evaluate the effects of propericiazine in a model of anxiety. Animals treated with 0.075 mg/kg propericiazine showed a reduction in anxiety measures (P<0.05) similar to that observed in those treated with diazepam. Propericiazine at the doses of 0.05 and 0.1 mg/kg had no significant anxiolytic effects (P>0.05) in the elevated plus-maze model of anxiety. Memory was not affected by propericiazine in any of the tests, but was impaired by diazepam. The results indicate a dose-related, inverse U-shaped effect of propericiazine in an anxiety model, but not on memory tasks, perhaps reflecting involvement of the dopaminergic system in the mechanisms of anxiety.

Behavioral characteristics of the offspring of adolescent rats

The aim of this study was to test the hypothesis that, during adulthood, the offspring of adolescent rats differ in emotionality, learning and memory from the offspring of adult rats. The behavior of the offspring of adolescent (age, 50-55 days) and adult rats (age, 90-95 days) was tested in the open field, activity cage, and passive and active avoidance apparatus. The latencies during training and testing in the passive avoidance apparatus of the offspring of adolescent parents were shorter than the latencies of control offspring (P<0.001 on both training and testing days). Offspring of adolescent parents showed shorter latency time in acquisition trials during active avoidance testing compared to control offspring (P<0.001). They also showed a higher number of active avoidance responses in the last four blocks of acquisition (P<0.001) and first two blocks of extinction trials (P<0.05 and P<0.001, respectively). The offspring of adolescent parents showed higher latency on the first day of testing in the open field (P<0.01) and a lower latency on the third day of testing (P<0.01). They also showed higher activity during all three days of testing (1st and 2nd day: P<0.01; 3rd day: P<0.05). The spontaneous activity of the offspring of adolescent parents in the activity cage was higher in the last three intervals of testing (P<0.001). In summary, the offspring of adolescent parents were less anxious and tended to be more active. The results of two learning and memory tests were opposite, but could be explained by a higher exploratory drive of the offspring of adolescent parents. This was probably due to chronic malnutrition stress and the disturbed mother-infant relationship in the litters of adolescent mothers.

Comparison of voluntary ethanol intake by two pairs of rat lines used as genetic models of anxiety

The relationship between anxiety-related behaviors and voluntary ethanol intake was examined in two pairs of rat lines by the oral ethanol self-administration procedure. Floripa high (H) and low (L) rats selectively bred for contrasting anxiety responses in the open-field test, and two inbred strains, spontaneously hypertensive rats (SHR) and Lewis rats which are known to differ significantly when submitted to several behavioral tests of anxiety/emotionality, were used (9-10 animals/line/sex). No differences in the choice of ethanol solutions (2%, days 1-4, and 4%, days 5-8, respectively) in a 2-bottle paradigm were detected between Floripa H and L rats (1.94 ± 0.37 vs 1.61 ± 0.37 g/kg for ethanol intake on day 8 by the Floripa H and L rat lines, respectively). Contrary to expectations, the less anxious SHR rats consumed significantly more ethanol than Lewis rats (respective intake of 2.30 ± 0.45 and 0.72 ± 0.33 g/kg on day 8) which are known to be both addiction-prone and highly anxious. Regardless of strain, female rats consumed more ethanol than males (approximately 46%). The results showed no relationship between high anxiety and voluntary intake of ethanol for Floripa H and L rats. A negative association between these two variables, however, was found for SHR and Lewis rat strains. Data from the literature regarding the association between anxiety and alcohol intake in animal models are not conclusive, but the present results indicate that factors other than increased inborn anxiety probably lead to the individual differences in ethanol drinking behavior.

Antinociceptive effect of novel pyrazolines in mice

The antinociceptive effect of six novel synthetic pyrazolines (3-ethoxymethyl-5-ethoxycarbonyl-1H-pyrazole (Pz 1) and its corresponding 1-substituted methyl (Pz 2) and phenyl (Pz 3) analogues, and 3-(1-ethoxyethyl)-5-ethoxycarbonyl-1H-pyrazole (Pz 4) and its corresponding 1-substituted methyl (Pz 5) and phenyl (Pz 6) analogues) was evaluated by the tail immersion test in adult male albino mice. The animals (N = 11-12 in each group) received vehicle (5% Tween 80, 10 ml/kg, sc) or 1.5 mmol/kg of each of the pyrazolines (Pz 1-Pz 6), sc. Fifteen, thirty and sixty minutes after drug administration, the mice were subjected to the tail immersion test. Thirty minutes after drug administration Pz 2 and Pz 3 increased tail withdrawal latency (vehicle = 3.4 ± 0.2; Pz 2 = 5.2 ± 0.4; Pz 3 = 5.9 ± 0.4 s; mean ± SEM), whereas the other pyrazolines did not present antinociceptive activity. Dose-effect curves (0.15 to 1.5 mmol/kg) were constructed for the bioactive pyrazolines. Pz 2 (1.5 mmol/kg, sc) impaired motor coordination in the rotarod and increased immobility in the open-field test. Pz 3 did not alter rotarod performance and spontaneous locomotion, but increased immobility in the open field at the dose of 1.5 mmol/kg. The involvement of opioid mechanisms in the pyrazoline-induced antinociception was investigated by pretreating the animals with naloxone (2.75 µmol/kg, sc). Naloxone prevented Pz 3- but not Pz 2-induced antinociception. Moreover, naloxone pretreatment did not alter Pz 3-induced immobility. We conclude that Pz 3-induced antinociception involves opioid mechanisms but this is not the case for Pz 2.

Genotoxic, neurotoxic and neuroprotective activities of apomorphine and its oxidized derivative 8-oxo-apomorphine

Apomorphine is a dopamine receptor agonist proposed to be a neuroprotective agent in the treatment of patients with Parkinson's disease. Both in vivo and in vitro studies have shown that apomorphine displays both antioxidant and pro-oxidant actions, and might have either neuroprotective or neurotoxic effects on the central nervous system. Some of the neurotoxic effects of apomorphine are mediated by its oxidation derivatives. In the present review, we discuss recent studies from our laboratory in which the molecular, cellular and neurobehavioral effects of apomorphine and its oxidized derivative, 8-oxo-apomorphine-semiquinone (8-OASQ), were evaluated in different experimental models, i.e., in vitro genotoxicity in Salmonella/microsome assay and WP2 Mutoxitest, sensitivity assay in Saccharomyces cerevisiae, neurobehavioral procedures (inhibition avoidance task, open field behavior, and habituation) in rats, stereotyped behavior in mice, and Comet assay and oxidative stress analyses in mouse brain. Our results show that apomorphine and 8-OASQ induce differential mutagenic, neurochemical and neurobehavioral effects. 8-OASQ displays cytotoxic effects and oxidative and frameshift mutagenic activities, while apomorphine shows antimutagenic and antioxidant effects in vitro. 8-OASQ induces a significant increase of DNA damage in mouse brain tissue. Both apomorphine and 8-OASQ impair memory for aversive training in rats, although the two drugs showed a different dose-response pattern. 8-OASQ fails to induce stereotyped behaviors in mice. The implications of these findings are discussed in the light of evidence from studies by other groups. We propose that the neuroprotective and neurotoxic effects of dopamine agonists might be mediated, in part, by their oxidized metabolites.

Effect of Erythrina velutina and Erythrina mulungu in rats submitted to animal models of anxiety and depression

Erythrina velutina (EV) and Erythrina mulungu (EM), popularly used in Brazil as tranquilizing agents, were studied. The effects of acute and chronic oral treatment with a water:alcohol extract of EV (7:3, plant grounded stem bark; acute = 100, 200, 400 mg/kg; chronic = 50, 100, 200 mg/kg) were evaluated in rats (N = 11-12) submitted to the elevated T-maze (for avoidance and escape measurements) model of anxiety. This model was selected for its presumed capacity to elicit specific subtypes of anxiety disorders recognized in clinical practice: avoidance has been related to generalized anxiety and escape to panic. Additionally, animals were treated with the same doses of EV and EM (water:alcohol 7:3, inflorescence extract) and submitted to the forced swim test for the evaluation of antidepressant activity (N = 7-10). Both treatment regimens with EV impaired elevated T-maze avoidance latencies, without altering escape, in a way similar to the reference drug diazepam (avoidance 1, mean ± SEM, acute study: 131.1 ± 45.5 (control), 9.0 ± 3.3 (diazepam), 12.7 ± 2.9 (200 mg/kg), 28.8 ± 15.3 (400 mg/kg); chronic study: 131.7 ± 46.9 (control), 35.8 ± 29.7 (diazepam), 24.4 ± 10.4 (50 mg/kg), 29.7 ± 11.5 (200 mg/kg)). Neither EV nor EM altered measurements performed in the forced swim test, in contrast to the reference drug imipramine that significantly decreased immobility time after chronic treatment. These results were not due to motor alterations since no significant effects were detected in an open field. These observations suggest that EV exerts anxiolytic-like effects on a specific subset of defensive behaviors which have been associated with generalized anxiety disorder.