Changes in the Behavior and Body Weight of Mature, Adult Male Wistar Han Rats after Reduced Social Grouping and Social Isolation.

Changes in housing density, including individual housing, are commonly necessary in animal research. Obtaining reproducibility and translational validity in biomedical research requires an understanding of how animals adapt to changes in housing density. Existing literature mainly addresses acclimatization after transportation. We used a within-subject design to examine changes in behavior and weight gain of 4-mo-old male Wistar Han rats after reduction of their social group (RSG; due to removal of one rat from a cage containing 3 rats) and social isolation (SI; the removed rat) for the subsequent 2 wk. Changes in weight gain and in exploratory and center-avoidance behavior in an inescapable open arena (OA) were measured before (D0) and on days 7 and 14 (D7 and D14, respectively) after social change. The motor response to d-amphetamine (1.5 mg/kg), which stimulates behavioral arousal in response to novelty, was assessed at D14. Within-subject design revealed that RSG rats in OA had less locomotion at D7 but not more center-avoidance behavior and had returned to the D0 activity level at D14; SI rats in OA had consistently less locomotion and more center-avoidance behavior. Rearing behavior during OA exposure did not change in either group. However, SI rats showed more center-avoidance behavior in OA, greater weight gain, and less amphetamine-induced rearing at D14 as compared with RSG rats. These data indicate that after RSG, mature adult male rats require 2 wk to return to their baseline level of OA-related behavior, while after SI they gain weight and acquire maladaptive exploratory and center-avoidance behavior. The finding that SI produces maladaptive behavioral and physiologic alterations in adult male rats deserves attention because these changes could have confounding effects on research findings.

The Influence of Unlimited Sucrose Intake on Body Weight and Behavior-Findings from a Mouse Model.

A potential relationship between unrestricted sucrose intake (USI), overweight, and emotional/behavioral control has not been well documented. We examined the influence of USI and having less sweetness than expected on body weight (BW), motor/exploratory, anxiety-like, and social dominant behavior in adult C57BL/6J male mice. Animals had free access to water (group 1) or 32% sucrose and water (sucrose groups 2-5) for 10 days. Then, group 2 remained with 32% sucrose while groups 3-5 were subjected to the downshift (24 h access to 4%, 8%, or 16% sucrose). All experimental groups were weighed and tested in the novel-open arena (NA), elevated plus maze (EPM), and tube tests to assess BW, motor/exploratory, anxiety-like, and social dominance behavior, respectively. USI did not influence animals' BW but produced hyperactivity and anxiolytic-like behavior, which was evident in EPM but not in NA; the outcomes of the downshift were comparable. USI did not influence successes/wins in the tube test but altered emotions that drive the winning, favoring a less anxious behavioral phenotype; this was not evident in the downshifted groups. Observed findings suggest that USI promotes sensation-seeking and motivates dominance, without changing BW, while blunted emotional base of social dominance might be an early mark of the downshift.

The Influence of Social Isolation on Social Orientation, Sociability, Social Novelty Preference, and Hippocampal Parvalbumin-Expressing Interneurons in Peripubertal Rats - Understanding the Importance of Meeting Social Needs in Adolescence.

The fulfillment of belonging needs underlies a variety of behaviors. In order to understand how social needs unmet during maturation shape everyday life, we examined social motivation and cognition in peripubertal rats, as a rodent model of adolescence, subjected to social isolation (SI) during early and early-to-mid adolescence. The behavioral correlates of social orientation (social space preference), sociability (preference for social over non-social novelty), and social novelty preference (SNP) were examined in group-housed (GH) and single-housed (SH) rats in a 3-chamber test. The response to social odors was examined to gain insights into the developmental role of social odors in motivated social behavior. Differentiation between appetitive (number of visits/approaches) and consummatory (exploratory time) aspects of motivated social behavior was done to determine which facet of social motivation characterizes maturation when social needs are met and which aspect dominates when social needs are unsatisfied. The SI-sensitive parvalbumin-expressing interneurons (PVI) in the hippocampus were examined using immunohistochemistry. The main findings are the following: (1) in GH rats, the preference for social space is not evident regardless of animals' age, while sociability becomes apparent in mid-adolescence strictly through consummatory behavior, along with complete SNP (appetitive, consummatory); (2) SH promotes staying in a social chamber/space regardless of animals' age and produces an appetitive preference for it only in early-adolescent animals; (3) SH promotes sociability (appetitive, consummatory) regardless of the animals' age and prevents the SNP; (4) the preference for a social odor is displayed in all the groups through consummatory behavior, while appetitive behavior is evident only in SH rats; (5) the response to social odors does not commensurate directly to the response to conspecifics; (6) SH does not influence PVI in the hippocampus, except in the case of early-adolescence when a transient decrease in the dentate gyrus is observed. These results accentuate the developmental complexity of social motivation and cognition, and the power of SI in adolescence to infringe social maturation at different functional levels, promoting appetitive behavior toward peers overall but harming the interest for social novelty. The findings emphasize the importance of the fulfillment of basic social needs in the navigation through the social world.

Altered hedonic, novelty-, stress- and D-amphetamine-induced response due to social isolation in peripuberty.

Reduction in direct social contact with peers during early adolescence is thought to be a risk factor for an increase in depressive symptoms, but there is still no clear evidence to suggest early behavioral manifestations and their association with the later outcome of social distancing during this period. To address this question, we used social isolation paradigm in peripubertal rats as the rodent model of adolescence. The litter was an experimental unit. On postnatal day 29, each litter gave group-housed and single-housed males, which were reared and tested one week and two weeks thereafter. Psychomotor/emotional response to novelty in exploration-based tasks, behavioral and neuronal responses to the drug reward (D-amphetamine), motivation/hedonic behavior, physiological and response to physiological stress were examined. Social isolation in peripubertal rats manifested through: hyper-reactivity/agitation and the state anxiety/risk-taking at an early stage; reduced behavioral response to D-amphetamine and altered neural processing of this stimulus, at a later stage; consummatory hypohedonia that deepened over time without changing the motivation to eat; unchanged body weight gain and resting blood corticosterone, cortisol and glucose levels over time; altered blood biochemistry (silenced corticosterone and increased glucose) due to overnight fasting only at an early stage. Our results highlight that the outcome of reduced direct social contact with peers during peripuberty is dynamic, with the cluster of atypical early symptoms that evolve into the syndrome that is delicate for assessment through routinely measurable behavior and biomarkers of stress, but with progressive consummatory hypohedonia and unaffected motivation to eat as stable marks.

A New Look at an Old Drug: Cumulative Effects of Low Ribavirin Doses in Amphetamine-Sensitized Rats.

BACKGROUND: Psychotic states related to psychostimulant misuse in patients with hepatitis C virus infection may complicate acceptance and reaction to antiviral treatment. This observation equally applies to the widely used ribavirin therapy. OBJECTIVE: We examined psychomotor and body weight gain responses to low ribavirin doses after cessation of intermittent amphetamine treatment in adult rats to assess its role in neurobehavioral outcome during psychostimulant withdrawal. METHOD: The model of amphetamine-induced (1.5 mg/kg/day, i.p., 7 consecutive days) motor sensitization and affected body weight gain was established in adult male Wistar rats. Then, additional cohort of amphetaminesensitized rats was subjected to saline (0.9% NaCl; 1 mL/kg/day; i.p.) or ribavirin (10, 20 and 30 mg/kg/day, i.p.) treatment for 7 consecutive days. Animals' motor activity in a novel environment was monitored after the 1st and the 7th saline/ribavirin injection. Body weight gain was calculated as appropriate. Determination and quantification of ribavirin in the brain tissue were performed also. RESULTS: The 1st application of ribavirin to amphetamine-sensitized rats affected/decreased their novelty-induced motor activity only at a dose of 30 mg/kg. After the 7th application, ribavirin 30 mg/kg/day still decreased, while 10 and 20 mg/kg/day increased novelty-induced motor activity. These behavioral effects coincided with the time required to reach maximum ribavirin concentration in the brain. Body weight gain during withdrawal was not influenced by any of the doses tested. CONCLUSION: Ribavirin displays central effects that in repeated treatment, depending on the applied dose, could significantly influence psychomotor response but not body weight gain during psychostimulant/amphetamine withdrawal.

Motivation, risk-taking and sensation seeking behavior in propofol anesthesia exposed peripubertal rats.

Adolescent neurodevelopment confer vulnerability to the actions of treatments that produce adaptations in neurocircuitry underlying motivation, impulsivity and reward. Considering wide usage of a sedative-hypnotic agent propofol in clinical practice, we examined whether propofol is a challenging treatment for peripubertal brain. Motivation/hedonic behavior (sucrose preference test), approach/avoidance behavior (elevated plus maze test) and response to dissociative drug phencyclidine (PCP) were studied in peripubertal rats (the rodent model of periadolescence) after propofol anesthesia exposure (PAE). Neurodegeneration (Fluoro-Jade staining) and the expression of proteins (Western blot) involved in excitatory synaptic transmission and activity-dependent synaptic stabilization in the medial prefrontal cortex (mPFC) and striatum (components of motivation/reward circuitry; process both appetitive and aversive events) were examined as well. In peripubertal rats PAE produced 1) transient brain-region specific changes in the expression of N-methyl-d-aspartate (NMDA) receptor subunits NR2A and NR2B, PSD-95 and N-cadherin, without neurotoxicity, 2) hyperlocomotor response to PCP, 3) no changes in preference for palatable 1% sucrose solution and a decrease in food eaten, 4) preference for 20% sucrose solution without changes in food eaten, 5) stretch-attended postures and open arms entries in the elevated plus maze test. Overall, these novel findings show that PAE leaves transient synaptic trace recognized as early form of synaptic plasticity related to passive drug exposure in the brain systems implicated in motivation/reward, increases drug-responsiveness, favors risk-taking and preference of novel/intense stimuli repairing otherwise present motivational deficiency. These findings accentuate multifaceted response to propofol in peripuberty and the importance of environmental stability for the most favorable neurobehavioral recovery.

The influence of propofol anesthesia exposure on nonaversive memory retrieval and expression of molecules involved in memory process in the dorsal hippocampus in peripubertal rats.

BACKGROUND: The effects of anesthetic drugs on postoperative cognitive function in children are not well defined and have not been experimentally addressed. AIMS: The present study aimed to examine the influence of propofol anesthesia exposure on nonaversive hippocampus-dependent learning and biochemical changes involved in memory process in the dorsal hippocampus, in peripubertal rats as the rodent model of periadolescence. METHODS: The intersession spatial habituation and the novel object recognition tasks were used to assess spatial and nonspatial, nonaversive hippocampus-dependent learning. The exposure to anesthesia was performed after comparably long acquisition phases in both tasks. Behavioral testing lasted for 2 consecutive days (24-hour retention period). Changes in the expression of molecules involved in memory retrieval/reconsolidation were examined in the dorsal hippocampus by Western blot and immunohistochemistry, at the time of behavioral testing. RESULTS: Exposure to propofol anesthesia resulted in inappropriate assessment of spatial novelty at the beginning of the test session and affected continuation of acquisition in the spatial habituation test. The treatment did not affect recognition of the novel object at the beginning of the test session but it attenuated overall preference to novelty, reflecting retrieval of a weak memory. The expression of phosphorylated extracellular signal-regulated kinase 2 (involved in memory retrieval) was decreased while the level of phosphorylated Ca2+ /calmodulin-dependent protein kinase IIα and early growth response protein 1 (involved in memory reconsolidation) was increased in the dorsal hippocampus. The level of Finkel-Biskis-Jinkins murine osteosarcoma viral oncogene homolog B (neuronal activity indicator) was increased in the dorsal dentate gyrus. Enhanced exploratory activity was still evident in the propofol anesthesia exposure (PAE) group 48 hour after the treatment in both tasks. CONCLUSION: In peripubertal rats, propofol anesthesia exposure affects memory retrieval and acquisition of new learning in the spatial and nonspatial, nonaversive learning tasks 24 hour after the treatment, along with the expression of molecules that participate in memory retrieval/reconsolidation in the dorsal hippocampus. These results may have clinical implications, favoring control of basic cognitive functions in older children after the propofol exposure.

Brain molecular changes and behavioral alterations induced by propofol anesthesia exposure in peripubertal rats.

BACKGROUND: Propofol is commonly used in modern anesthesiology. Some findings suggest that it is highly addictive. AIM: In this study it was examined whether propofol anesthesia exposure was able to induce behavioral alterations and brain molecular changes already described in addictive drug usage in peripubertal rats, during the onset of mid/periadolescence as a developmental period with increasing vulnerability to drug addiction. METHODS: The expression of D1 dopamine receptor, a dopamine, and cAMP-regulated phosphoprotein with a Mr 32 000; Ca2+ /calmodulin-dependent protein kinase IIα; and Finkel-Biskis-Jinkins murine osteosarcoma viral oncogene homolog-B was examined in peripubertal rats 4, 24, and 48 hour after propofol anesthesia exposure by Western blot and immunohistochemistry. Brain regions of interest were the medial prefrontal cortex, the striatum, and the thalamus. Anxiety and behavioral cross-sensitization to d-amphetamine were examined as well. RESULTS: Significant increase in the expression of dopamine and cAMP-regulated phosphoprotein with a Mr 32 000 phosphorylated at threonine 34, a postsynaptic marker of dopaminergic neurotransmission, and Finkel-Biskis-Jinkins murine osteosarcoma viral oncogene homolog-B, a marker of neuronal activity, was detected in the thalamus of experimental animals 4-24 hour after the treatment, with the accent on the paraventricular thalamic nucleus. Significant increase in the expression of Ca2+ /calmodulin-dependent protein kinase IIα phosphorylated at threonine 286, a sensor of synaptic activity, was observed in the prefrontal cortex and the striatum 24 hour after propofol anesthesia exposure. It was accompanied by a significant decrease in Finkel-Biskis-Jinkins murine osteosarcoma viral oncogene homolog-B expression in the striatum. Decreased behavioral inhibition in aversive environment and increased motor response to d-amphetamine in a context-independent manner were observed as well. CONCLUSION: In peripubertal rats, propofol anesthesia exposure induces transient molecular and behavioral response that share similarities with those reported previously for addictive drugs. In the absence of additional pharmacological manipulation, all detected effects receded within 48 hour after the treatment.

Resistance to DNA Damaging Agents Produced Invasive Phenotype of Rat Glioma Cells-Characterization of a New in Vivo Model.

Chemoresistance and invasion properties are severe limitations to efficient glioma therapy. Therefore, development of glioma in vivo models that more accurately resemble the situation observed in patients emerges. Previously, we established RC6 rat glioma cell line resistant to DNA damaging agents including antiglioma approved therapies such as 3-bis(2-chloroethyl)-1-nitrosourea (BCNU) and temozolomide (TMZ). Herein, we evaluated the invasiveness of RC6 cells in vitro and in a new orthotopic animal model. For comparison, we used C6 cells from which RC6 cells originated. Differences in cell growth properties were assessed by real-time cell analyzer. Cells' invasive potential in vitro was studied in fluorescently labeled gelatin and by formation of multicellular spheroids in hydrogel. For animal studies, fluorescently labeled cells were inoculated into adult male Wistar rat brains. Consecutive coronal and sagittal brain sections were analyzed 10 and 25 days post-inoculation, while rats' behavior was recorded during three days in the open field test starting from 25th day post-inoculation. We demonstrated that development of chemoresistance induced invasive phenotype of RC6 cells with significant behavioral impediments implying usefulness of orthotopic RC6 glioma allograft in preclinical studies for the examination of new approaches to counteract both chemoresistance and invasion of glioma cells.

The Fas Ligand/Fas Death Receptor Pathways Contribute to Propofol-Induced Apoptosis and Neuroinflammation in the Brain of Neonatal Rats.

A number of experimental studies have reported that exposure to common, clinically used anesthetics induce extensive neuroapoptosis and cognitive impairment when applied to young rodents, up to 2 weeks old, in phase of rapid synaptogenesis. Propofol is the most used general anesthetic in clinical practice whose mechanisms of neurotoxicity on the developing brain remains to be examined in depth. This study investigated effects of different exposures to propofol anesthesia on Fas receptor and Fas ligand expressions, which mediate proapoptotic and proinflammation signaling in the brain. Propofol (20 mg/kg) was administered to 7-day-old rats in multiple doses sufficient to maintain 2-, 4- and 6-h duration of anesthesia. Animals were sacrificed at 0, 4, 16 and 24 h after termination of anesthesia. It was found that propofol anesthesia induced Fas/FasL and downstream caspase-8 expression more prominently in the thalamus than in the cortex. Opposite, Bcl-2 and caspase-9, markers of intrinsic pathway activation, were shown to be more influenced by propofol treatment in the cortex. Further, we have established upregulation of caspase-1 and IL-1ß cytokine transcription as well as subsequent activation of microglia that is potentially associated with brain inflammation. Behavioral analyses revealed that P35 and P60 animals, neonatally exposed to propofol, had significantly higher motor activity during three consecutive days of testing in the open field, though formation of the intersession habituation was not prevented. This data, together with our previous results, contributes to elucidation of complex mechanisms of propofol toxicity in developing brain.

Early physical and motor development of mouse offspring exposed to valproic acid throughout intrauterine development.

Clinical research has identified developmental delay and physical malformations in children prenatally exposed to the antiepileptic drug (AED) valproic acid (VPA). However, the early signs of neurodevelopmental deficits, their evolution during postnatal development and growth, and the dose effects of VPA are not well understood. The present study aimed to examine the influence of maternal exposure to a wide dose range (50, 100, 200 and 400mg/kg/day) of VPA during breeding and gestation on early physical and neuromotor development in mice offspring. Body weight gain, eye opening, the surface righting reflex (SRR) and tail suspension test (TST) were examined in the offspring at postnatal days 5, 10 and 15. We observed that: (1) all tested doses of VPA reduced the body weight of the offspring and the timing of eye opening; (2) offspring exposed to VPA displayed immature forms of righting and required more time to complete the SRR; (3) latency for the first immobilization in the TST is shorter in offspring exposed to higher doses of VPA; however, mice in all groups exposed to VPA exhibited atypical changes in this parameter during the examined period of maturation; (4) irregularities in swinging and curling activities were observed in animals exposed to higher doses of VPA. This study points to delayed somatic development and postponed maturation of the motor system in all of the offspring prenatally exposed to VPA, with stronger effects observed at higher doses. The results implicate that the strategy of continuous monitoring of general health and achievements in motor milestones during the early postnatal development in prenatally VPA-exposed offspring, irrespectively of the dose applied, could help to recognize early developmental irregularities.

Long-term dietary restriction differentially affects the expression of BDNF and its receptors in the cortex and hippocampus of middle-aged and aged male rats.

Dietary restriction (DR) exerts significant beneficial effects in terms of aging and age-related diseases in many organisms including humans. The present study aimed to examine the influence of long-term DR on the BDNF system at the transcriptional and translational levels in the cortex and hippocampus of middle-aged (12-month-old) and aged (24-month-old) male Wistar rats. The obtained results revealed that the DR upregulated the expression of exon-specific BDNF transcripts in both regions, followed by elevated levels of mBDNF only in the cortex in middle-aged animals. In aged animals, DR modulated BDNF protein levels by increasing proBDNF and by declining mBDNF levels. Additionally, elevated levels of the full-length TrkB accompanied by a decreased level of the less-glycosylated TrkB protein were observed in middle-aged rats following DR, while in aged rats, DR amplified only the expression of the less-glycosylated form of TrkB. The levels of phosphorylated TrkB(Y816) were stable during aging regardless of feeding. Reduced levels of p75(NTR) were detected in both regions of middle-aged DR-fed animals, while a significant increase was measured in the cortex of aged DR-fed rats. These findings shed additional light on DR as a modulator of BDNF system revealing its disparate effects in middle-aged and aged animals. Given the importance of the proBDNF/BDNF circuit-level expression in different brain functions and various aspects of behavior, it is necessary to further elucidate the optimal duration of the applied dietary regimen with regard to the animal age in order to achieve its most favorable effects.