Saline Control Rats Research Articles

Benefits of Sebastiania hispida (Euphorbiaceae) extract and photobiomodulation therapy as potentially adjunctive strategies to be explored against snake envenoming.

The purpose of this study was to assess the topic use of Sebastiania hispida extract and low-level gallium-arsenide laser irradiation (GaAs, 904nm) to reduce the local myonecrosis and edema of Bothrops moojeni snake venom-injected gastrocnemius. Wistar rats receiving intramuscular venom injection (VBm) were compared with saline control (S) and envenomed rats receiving local exposure to plant extract (VExt) or laser irradiation (VL). The phytochemistry and thin-layer chromatography of S. hispida extract indicated the presence of phenolic compounds like gallic acid and flavonoids including quercetin. Gastrocnemius of VExt and VL groups had a significant reduction of edema and creatine kinase (CK) activities and a greater Myogenin (MyoG) expression compared to VBm group, with the plant extract efficacy better than laser exposure. Reduction of edema and serum CK activities reflects a lessening of muscle damage, whereas the increase of MyoG indicates myoblast differentiation and acceleration of muscle repair. The S. hispida richness in phenolic compounds and flavonoids, such as the light modulatory ability to triggering a multitude of cell signalings likely underlie the positive outcomes. Our findings suggest both treatments as potential auxiliary tools to be explored in clinical trials in combination with anti-venom therapy after Bothropic snakebites.

Prevention of Renal Medullary Vascular Congestion is Associated with Reduced Mortality and Preservation of Capillary Density Following Renal Ischemia

The aggregation of red blood cells in the microvascular of the renal medulla is a hallmark of ischemic acute kidney injury (AKI). Our lab has recently shown that pretreatment with low dose lipopolysaccharide (LPS) prior to ischemia-reperfusion (IR) prevents this vascular congestion. It remains unknown, however, whether prevention of medullary vascular congestion improves recovery from IR. The goal of the current study was to test the hypothesis that ‘prevention of vascular congestion with low dose LPS improves functional recovery of the kidney and attenuates rarefaction of the renal medulla following IR’. Male WKY rats (8 weeks) were implanted with radiotelemetry devices (DSI, St. Paul, MN) and allowed to recover for two weeks prior to beginning blood pressure measurement. Rats were then randomized to receive injections (i.p.) of either low dose LPS (1000µg/kg) (n=4) to prevent vascular congestion or saline control (n=4) once daily for 3 days. 1 day following the 3rd injection, all rats underwent a 45 minute, warm bilateral IR and were allowed to recover for 7 days. Blood pressure was measured for the duration of the study. Plasma was collected by tail vein pre-IR and at sacrifice for measurement of creatinine (Cr) and blood urea nitrogen (BUN). Vascular rarefaction was assessed at sacrifice in histological kidney sections via %Threshold Area of CD34+ vascular endothelium (Metamorph). Following recovery of IR, there was a 100% and 50% survival rate in LPS treated and saline control rats, respectively. Saline control rats (with vascular congestion following IR) had a decrease in CD34+ vascular density in the inner stripe of the outer medulla (OM) (Unpaired t-test, p=0.05) and the inner medulla (Unpaired t-test, p=0.009), but there were no differences in CD34+ staining in the cortex (Unpaired t-test, p=0.159) or outer stripe of the OM (Unpaired t-test, p=0.095) (Table 1). There were no differences in 24 hour mean arterial pressure between groups over the course of the study (two-way RM ANOVA, pINTERACTION=0.689). At sacrifice, plasma Cr (pINTERACTION=0.249) and BUN (pINTERACTION=0.345) were not different between groups and had recovered to near pre-IR values. Prevention of vascular congestion in LPS treated rats increased the 7 day survival to 100% following a 45 minute bilateral IR in contrast to control rats which, as expected, had a 50% survival rate. Importantly, this prevention of congestion was associated with preserved capillary density (CD43+) in the renal medulla. These data indicate an important role of vascular congestion to independently drive increased mortality and capillary rarefaction following IR. As renal vascular rarefaction has been implicated as an important factor in the progression to chronic kidney disease following AKI, prevention of vascular congestion may be an important target to prevent this deleterious sequelae.

Dysregulated expression of the alternatively spliced variant mRNAs of the mu opioid receptor gene, OPRM1, in the medial prefrontal cortex of male human heroin abusers and heroin self-administering male rats.

Heroin, a mu agonist, acts through the mu opioid receptor. The mu opioid receptor gene, OPRM1, undergoes extensive alternative splicing, creating an array of splice variants that are conserved from rodent to humans. Increasing evidence suggests that these OPRM1 splice variants are pharmacologically important in mediating various actions of mu opioids, including analgesia, tolerance, physical dependence, rewarding behavior, as well as addiction. In the present study, we examine expression of the OPRM1 splice variant mRNAs in the medial prefrontal cortex (mPFC), one of the major brain regions involved in decision-making and drug-seeking behaviors, of male human heroin abusers and male rats that developed stable heroin-seeking behavior using an intravenous heroin self-administration (SA) model. The results show similar expression profiles among multiple OPRM1 splice variants in both human control subjects and saline control rats, illustrating conservation of OPRM1 alternative splicing from rodent to humans. Moreover, the expressions of several OPRM1 splice variant mRNAs were dysregulated in the postmortem mPFCs from heroin abusers compared to the control subjects. Similar patterns were observed in the rat heroin SA model. These findings suggest potential roles of the OPRM1 splice variants in heroin addiction that could be mechanistically explored using the rat heroin SA model.

Withdrawal From Cocaine Self-administration Alters the Regulation of Protein Translation in the Nucleus Accumbens

BackgroundCue-induced cocaine craving incubates during abstinence from cocaine self-administration. Expression of incubation ultimately depends on elevation of homomeric GluA1 alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid receptors in the nucleus accumbens (NAc). This adaptation requires ongoing protein translation for its maintenance. Aberrant translation is implicated in central nervous system diseases, but nothing is known about glutamatergic regulation of translation in the drug-naïve NAc or after incubation. MethodsNAc tissue was obtained from drug-naïve rats and from rats after 1 or >40 days of abstinence from extended-access cocaine or saline self-administration. Newly translated proteins were labeled using 35S-Met/Cys or puromycin. We compared basal overall translation and its regulation by metabotropic glutamate receptor 1 (mGlu1), mGlu5, and N-methyl-D-aspartate receptors (NMDARs) in drug-naïve, saline control, and cocaine rats, and we compared GluA1 and GluA2 translation by immunoprecipitating puromycin-labeled proteins. ResultsIn all groups, overall translation was unaltered by mGlu1 blockade (LY367385) but increased by mGlu5 blockade (MTEP). NMDAR blockade (AVP) increased overall translation in drug-naïve and saline control rats but not in cocaine/late withdrawal rats. Cocaine/late withdrawal rats exhibited greater translation of GluA1 (but not GluA2), which was not further affected by NMDAR blockade. ConclusionsOur results suggest that increased GluA1 translation contributes to the elevated homomeric GluA1 alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid receptor levels in the NAc that mediate incubation. Additional contributions to incubation-related plasticity may result from loss of the braking influence on translation normally exerted by NMDARs. Apart from elucidating incubation-related adaptations, we found a suppressive effect of mGlu5 on NAc translation regardless of drug exposure, which is opposite to results obtained in the hippocampus and points to heterogeneity of translational regulation between brain regions.

PAN-811 prevents chemotherapy-induced cognitive impairment and preserves neurogenesis in the hippocampus of adult rats.

Chemotherapy-induced cognitive impairment (CICI) occurs in a substantial proportion of treated cancer patients, with no drug currently available for its therapy. This study investigated whether PAN-811, a ribonucleotide reductase inhibitor, can reduce cognitive impairment and related suppression of neurogenesis following chemotherapy in an animal model. Young adult rats in Chemo and Chemo+PAN-811 groups received 3 intraperitoneal (i.p.) injections of methotrexate (MTX) and 5-fluorouracil (5-FU), and those in Saline and Saline+PAN-811 groups received equal volumes of physiological saline at 10-day intervals. PAN-811 in saline was delivered through i.p. injection, 10 min following each saline (Saline+PAN-811 group) or MTX/5-FU (Chemo+PAN-811 group) treatment, while equal volumes of saline were delivered to Saline and Chemo groups. Over Days 31–66, rats were administered tests of spatial memory, nonmatching-to-sample rule learning, and discrimination learning, which are sensitive to dysfunction in hippocampus, frontal lobe and striatum, respectively. On Day 97, neurogenesis was immnunohistochemically evaluated by counting doublecortin-positive (DCX+) cells in the dentate gyrus (DG). The results demonstrated that the Chemo group was impaired on the three cognitive tasks, but co-administration of PAN-811 significantly reduced all MTX/5-FU-induced cognitive impairments. In addition, MTX/5-FU reduced DCX+ cells to 67% of that in Saline control rats, an effect that was completely blocked by PAN-811 co-administration. Overall, we present the first evidence that PAN-811 protects cognitive functions and preserves neurogenesis from deleterious effects of MTX/5-FU. The current findings provide a basis for rapid clinical translation to determine the effect of PAN-811 on CICI in human.

Alterations in brain activation in response to prolonged morphine withdrawal-induced behavioral inflexibility in rats.

The inability to stop a repetitive maladaptive behavior is a main problem in addictive disorders. Neuroadaptations that are associated with behavioral inflexibility may be involved in compulsive drug use. The aim of the present study was to investigate the pattern of behavioral inflexibility during morphine withdrawal and map brain activation that is linked to alterations in flexibility. We first analyzed the effects of chronic morphine exposure on reversal learning after 2-week (short-term) and 6-week (prolonged) morphine withdrawal. We then compared the level of neuronal activation using cFos immunohistochemistry in 15 brain areas between rats that underwent morphine withdrawal and saline-control rats after a test of reversal learning. Only prolonged morphine withdrawal impaired reversal learning. Rats that exhibited impairments in reversal learning presented a significant decrease in cFos expression in the orbitofrontal cortex (OFC), including the medial, lateral, and ventral OFC. cFos expression significantly increased in the dorsomedial striatum and major subregions of the medial prefrontal cortex (mPFC) in the morphine group. Rats that underwent prolonged morphine withdrawal exhibited no significant changes in cFos expression in the dorsolateral striatum, nucleus accumbens, amygdala, paraventricular thalamic nucleus, or motor cortex. The rats that underwent short-term withdrawal did not present any changes in cFos expression in any of these brain regions. Altogether, these data suggest that alterations in the function of the frontal cortex and its striatal connections during the late morphine withdrawal phase may underlie the disruption of inhibitory control in opioid dependence.

Comparative analysis of MBD-seq and MeDIP-seq and estimation of gene expression changes in a rodent model of schizophrenia

We conducted a comparative study of multiplexed affinity enrichment sequence methodologies (MBD-seq and MeDIP-seq) in a rodent model of schizophrenia, induced by in utero methylazoxymethanol acetate (MAM) exposure. We also examined related gene expression changes using a pooled sample approach. MBD-seq and MeDIP-seq identified 769 and 1771 differentially methylated regions (DMRs) between F2 offspring of MAM-exposed rats and saline control rats, respectively. The assays showed good concordance, with ~56% of MBD-seq-detected DMRs being identified by or proximal to MeDIP-seq DMRs. There was no significant overlap between DMRs and differentially expressed genes, suggesting that DNA methylation regulatory effects may act upon more distal genes, or are too subtle to detect using our approach. Methylation and gene expression gene ontology enrichment analyses identified biological processes important to schizophrenia pathophysiology, including neuron differentiation, prepulse inhibition, amphetamine response, and glutamatergic synaptic transmission regulation, reinforcing the utility of the MAM rodent model for schizophrenia research.

The area postrema (AP) and the parabrachial nucleus (PBN) are important sites for salmon calcitonin (sCT) to decrease evoked phasic dopamine release in the nucleus accumbens (NAc)

The pancreatic hormone amylin and its agonist salmon calcitonin (sCT) act via the area postrema (AP) and the lateral parabrachial nucleus (PBN) to reduce food intake. Investigations of amylin and sCT signaling in the ventral tegmental area (VTA) and nucleus accumbens (NAc) suggest that the eating inhibitory effect of amylin is, in part, mediated through the mesolimbic ‘reward’ pathway. Indeed, administration of the sCT directly to the VTA decreased phasic dopamine release (DA) in the NAc. However, it is not known if peripheral amylin modulates the mesolimbic system directly or whether this occurs via the AP and PBN. To determine whether and how peripheral amylin or sCT affect mesolimbic reward circuitry we utilized fast scan cyclic voltammetry under anesthesia to measure phasic DA release in the NAc evoked by electrical stimulation of the VTA in intact, AP lesioned and bilaterally PBN lesioned rats.Amylin (50μg/kg i.p.) did not change phasic DA responses compared to saline control rats. However, sCT (50μg/kg i.p.) decreased evoked DA release to VTA-stimulation over 1h compared to saline treated control rats. Further investigations determined that AP and bilateral PBN lesions abolished the ability of sCT to suppress evoked phasic DA responses to VTA-stimulation. These findings implicate the AP and the PBN as important sites for peripheral sCT to decrease evoked DA release in the NAc and suggest that these nuclei may influence hedonic and motivational processes to modulate food intake.

Establishment of a schizophrenic animal model through chronic administration of MK-801 in infancy and social isolation in childhood

BackgroundAlthough an increasing amount of evidence supports a “two-hit” hypothesis for the neurodevelopmental model of schizophrenia, there has been no development in animal models to test this hypothesis. MethodsAn animal model was established by chronic administration of 0.1, 0.3, and 0.5mg/kg MK-801 in P7-P21 rats followed by four weeks of social isolation in childhood and then five days of social housing. Animal behaviors were measured by the open field (OF) test, the novel object recognition (NOR) test, the prepulse inhibition (PPI) test, and the elevated plus maze (EPM) test. ResultsWe found a significant decrease in the NOR index in adolescent rats compared to saline control rats when administering 0.5mg/kg of MK-801 (P=0.02). We found that social isolation had no significant effect on NOR index, though social isolation significantly increased the total distance traveled and significantly decreased the resting time in adolescent rats in the OF test (P<0.001 and P=0.003, respectively). In contrast, we observed that MK-801 administration showed no significant effects on either total distance traveled or resting time. Both MK-801 administration and social isolation had no significant effect on the percent of PPI and startle amplitudes in adolescent rats. Social isolation significantly reduced the open arm entries in adolescent rats in the EPM test (P=0.023), but it did not reduce the ratio to enter the open arms and the stay time in open arm. Administration of MK-801 showed no significant effect on the indexes of entering the open arms in the EPM test on adolescent rats. ConclusionMK-801 intervention in infancy is associated with the damage of long-term visual memory, whereas social isolation in childhood is associated with the increased spontaneous activity and anxiety levels. Administration of MK-801 in infancy and social isolation in childhood are two independent factors on the neurodevelopmental defects.

Transgenerational effects of adolescent nicotine exposure in rats: Evidence for cognitive deficits in adult female offspring

This study investigated whether adolescent nicotine exposure in one generation of rats would impair the cognitive capacity of a subsequent generation. Male and female rats in the parental F0 generation were given twice-daily i.p. injections of either 1.0mg/kg nicotine or an equivalent volume of saline for 35days during adolescence on postnatal days 25–59 (P25–59). After reaching adulthood, male and female nicotine-exposed rats were paired for breeding as were male and female saline control rats. Only female offspring were used in this experiment. Half of the offspring of F0 nicotine-exposed breeders and half of the offspring of F0 saline control rats received twice-daily i.p. injections of 1.0mg/kg nicotine during adolescence on P25–59. The remainder of the rats received twice-daily saline injections for the same period. To evaluate transgenerational effects of nicotine exposure on complex cognitive learning abilities, F1 generation rats were trained to perform a highly structured serial pattern in a serial multiple choice (SMC) task. Beginning on P95, rats in the F1 generation were given either 4days of massed training (20patterns/day) followed by spaced training (10 patterns/day) or only spaced training. Transgenerational effects of adolescent nicotine exposure were observed as greater difficulty in learning a “violation element” of the pattern, which indicated that rats were impaired in the ability to encode and remember multiple sequential elements as compound or configural cues. The results indicated that for rats that received massed training, F1 generation rats with adolescent nicotine exposure whose F0 generation parents also experienced adolescent nicotine exposure showed poorer learning of the violation element than rats that experienced adolescent nicotine exposure only in the F1 generation. Thus, adolescent nicotine exposure in one generation of rats produced a cognitive impairment in the next generation.

Sex differences in hypothalamic-mediated tonic norepinephrine release for thermal hyperalgesia in rats

Neuropathic pain is treated using serotonin norepinephrine reuptake inhibitors with mixed results. Pain facilitation mediated by α1-adrenoceptors may be involved, but whether norepinephrine (NE) is tonically released is unclear. The aim of this study was to determine whether NE is tonically released from A7 cells following chronic constriction injury (CCI), and if the lateral hypothalamus (LH) plays a role in this release in male and female rats with nociceptive and neuropathic pain types. Neuropathic groups received left CCI while nociceptive groups remained naïve to injury. Fourteen days later, rats were given intrathecal infusion of either the α1-adrenoceptor antagonist WB4101, the α2-adrenoceptor antagonist yohimbine (74μg), or normal saline for control. Paw withdrawal latency (PWL) from a thermal stimulus was measured. The generalized estimated equation method was used for statistical analysis. Nociceptive rats given WB4101 had a PWL significantly longer than saline control (7.89±0.63 vs. 5.87±0.52s), while the PWL of neuropathic rats given WB4101 was 13.20±0.52s compared to 6.78±0.52s for the saline control rats. Yohimbine had no significant effect. Microinjection of cobalt chloride (CoCl) in the A7 catecholamine cell group to prevent synaptic transmission blocked the effect of WB4101 in all groups, supporting the notion that spinally descending A7 cells tonically release NE that contributes to α1-mediated nociceptive facilitation. Microinjection of CoCl into the left LH blocked the effect of WB4101 in nociceptive and neuropathic male rats, but had no effect in female rats of either pain type, suggesting differential innervation. These findings indicate that tonic release of NE acts at pronociceptive α1-adrenoceptors, that this effect is greater in rats with nerve damage, and that, while NE comes primarily from the A7 cell group, LH innervation of the A7 cell group is different between the sexes.

Reversal of MicroRNA Dysregulation in an Animal Model of Pulmonary Hypertension.

BackgroundAnimals models have played an important role in enhancing our understanding of the pathogenesis of pulmonary arterial hypertension (PAH). Dysregulation of the profile of microRNAs (miRNAs) has been demonstrated in human tissues from PAH patients and in animal models. In this study, we measured miRNA levels in the monocrotaline (MCT) rat model of PAH and examined whether blocking a specific dysregulated miRNA not previously reported in this model, attenuated PAH. We also evaluated changes in miRNA expression in lung specimens from MCT PAH rats overexpressing human prostacyclin synthase, which has been shown to attenuate MCT PAH.MethodsExpression levels of a panel of miRNAs were measured in MCT-PAH rats as compared to naïve (saline) control rats. Subsequently, MCT PAH rats were injected with a specific inhibitor (antagomiR) for miR-223 (A223) or a nonspecific control oligonucleotide (A-control) 4 days after MCT administration, then weekly. Three weeks later, RV systolic pressure and RV mass were measured. Total RNA, isolated from the lungs, microdissected pulmonary arteries, and right ventricle, was reverse transcribed and real-time quantitative PCR was performed. MiRNA levels were also measured in RNA isolated from paraffin sections of MCT-PAH rats overexpressing prostacyclin synthase.ResultsMiRs 17, 21, and 223 were consistently upregulated, whereas miRs 126, 145, 150, 204, 424, and 503 were downregulated in MCT PAH as compared to vehicle control. A223 significantly reduced levels of miR-223 in PA and lungs of MCT PAH rats as compared to levels measured in A-control or control MCT PAH rats, but A223 did not attenuate MCT PAH. Right ventricular mass and right ventricular systolic pressure in rats treated with A223 were not different from values in A-control or MCT PAH rats. In contrast, analysis of total RNA from lung specimens of MCT PAH rats overexpressing human prostacyclin synthase (hPGIS) demonstrated reversal of MCT-induced upregulation of miRs 17, 21, and 223 and an increase in levels of miR-424 and miR-503. Reduction in bone morphogenetic receptor 2 (BMPR2) messenger (m)RNA expression was not altered by A223, whereas human prostacyclin synthase overexpression restored BMPR2 mRNA to levels in MCT PAH to levels measured in naive controls.ConclusionsInhibition of miR-223 did not attenuate MCT PAH, whereas human prostacyclin synthase overexpression restored miRNA levels in MCT PAH to levels detected in naïve rats. These data may establish a paradigm linking attenuation of PAH to restoration of BMPR2 signaling.

Zoledronic acid suppresses callus remodeling but enhances callus strength in an osteoporotic rat model of fracture healing

Mini-abstractIn this study, we demonstrated that the use of zoledronic acid does not impair fracture healing, but results in superior callus size and resistance at the fracture site, which could be the consequence of a lower rate of bone turnover due to its anti-catabolic effect. ObjectiveTo investigate the effect of inhibition of bone remodeling by the bisphosphonate, zoledronic acid, on callus properties in an osteoporotic rat model of fracture healing. MethodsOvariectomized (OVX) rats were randomly divided into four treatment groups (n=24 per group): saline control (CNT); and three systemic zoledronic acid-injected groups (0.1mg/kg), administered 1day (ZOLD1), 1week (ZOLW1), and 2weeks (ZOLW2) after fracture. Rats were killed at either 6 or 12weeks postoperatively. Postmortem analyses included radiography, microcomputed tomography, histology, histomorphometry, biomechanical tests, and nanoindentation tests. ResultsTreatment with zoledronic acid led to a significant increase in trabecular bone volume within the callus, as well as in callus resistance, compared to those in the saline control rats; delayed administration (ZOLW2) reduced intrinsic material properties, including ultimate stress and elastic modulus, and microarchitecture parameters, including bone volume/total volume (BV/TV), trabecular thickness (Tb.Th), and connectivity density (Conn.D), compared with ZOLD1 at 12weeks after surgery. OVX had a negative effect on the progression of endochondral ossification at 6weeks. Zoledronic acid administration at an early stage following fracture may bind to early callus, and thus not affect subsequent callus formation and endochondral ossification, while delayed administration (ZOLW2) mildly suppresses bony callus remodeling. ConclusionThe superior results obtained with zoledronic acid (ZOLD1, ZOLW1, and ZOLW2) compared to CNT in terms of callus size and resistance could be the consequence of a lower rate of bone turnover at the fracture site due to the anti-catabolic effect of zoledronic acid. Mild suppression of callus remodeling by delayed administration did not impair the initial phase of the fracture healing process.

Social interaction reward decreases p38 activation in the nucleus accumbens shell of rats

We have previously shown that animals acquired robust conditioned place preference (CPP) to either social interaction alone or cocaine alone. Recently it has been reported that drugs of abuse abnormally activated p38, a member of mitogen-activated protein kinase family, in the nucleus accumbens. In this study, we aimed to investigate the expression of the activated form of p38 (pp38) in the nucleus accumbens shell and core of rats expressing either cocaine CPP or social interaction CPP 1 h, 2 h and 24 h after the CPP test. We hypothesized that cocaine CPP will increase pp38 in the nucleus accumbens shell/core as compared to social interaction CPP. Surprisingly, we found that 24 h after social interaction CPP, pp38 neuronal levels were decreased in the nucleus accumbens shell to the level of naïve rats. Control saline rats that received saline in both compartments of the CPP apparatus and cocaine CPP rats showed similar enhanced p38 activation as compared to naïve and social interaction CPP rats. We also found that the percentage of neurons expressing dopaminergic receptor D2R and pp38 was also decreased in the shell of the nucleus accumbens of social interaction CPP rats as compared to controls. Given the emerging role of p38 in stress/anxiety behaviors, these results suggest that (1) social interaction reward has anti-stress effects; (2) cocaine conditioning per se does not affect p38 activation and that (3) marginal stress is sufficient to induce p38 activation in the shell of the nucleus accumbens.

Intra-ventral tegmental area HIV-1 Tat1-86 attenuates nicotine-mediated locomotor sensitization and alters mesocorticolimbic ERK and CREB signaling in rats.

Cigarette smoking prevalence in the HIV-positive individuals is profoundly higher than that in the HIV-negative individuals. We have demonstrated that HIV-1 transgenic rats exhibit attenuated nicotine-mediated locomotor activity, altered cAMP response element binding protein (CREB) and extracellular regulated kinase (ERK1/2) signaling in the mesocorticolimbic regions. This study investigated the role of HIV-1 transactivator of transcription (Tat) protein in the alterations of nicotine-mediated behavior and the signaling pathway observed in the HIV-1 transgenic rats. Rats received bilateral microinjection of recombinant Tat1–86 (25 μg/side) or vehicle directed at ventral tegmental area (VTA) followed by locomotor testing in response to 13 daily intravenous injections of nicotine (0.05 mg/kg, freebase, once/day) or saline. Further, we examined the phosphorylated levels of CREB (pCREB) and ERK1/2 (pERK1/2) in the prefrontal cortex (PFC), nucleus accumbens (NAc) and VTA. Tat diminished baseline activity in saline control rats, and attenuated nicotine-induced behavioral sensitization. Following repeated saline injection, the basal levels of pERK1 in the NAc and VTA and pERK2 in VTA were lower in the vehicle control group, relative to the Tat group. After repeated nicotine injection, pERK1 in NAc and VTA and pERK2 in VTA were increased in the vehicle group, but not in the Tat group. Moreover, repeated nicotine injections decreased pCREB in the PFC and VTA in the Tat group but not in the vehicle group. Thus, these findings indicate that the direct injection of Tat at the VTA may mediate CREB and ERK activity in response to nicotine-induced locomotor activity.

Interaction of prenatal stress and morphine alters prolactin and seizure in rat pups

Prenatal exposure to stress and morphine has complicated effects on epileptic seizure. In the present study, effect of prenatal forced-swim stress and morphine co-administration on pentylenetetrazol (PTZ) induced epileptic behaviors and prolactin blood level (PBL) was investigated in rat offspring. Pregnant Wistar rats were divided to four groups of control–saline, control–morphine, stressed–saline and stressed–morphine. In the stressed group, pregnant rats were placed in 25°C water on gestation days 17, 18 and 19 (GD17, GD18 and GD19) for 30min. In the morphine/saline group, pregnant rats received morphine (10, 12 and 15mg/kg, IP, on GD17, GD18 and GD19, respectively) or saline (1ml, IP). In the morphine/saline-stressed group, the rats received morphine or saline and then exposed to stress. On postnatal days 6 and 15 (P6 and P15), blood samples were obtained and PBL was determined. At P15 and P25, the rest of the pups was injected with PTZ to induce seizure. Then, epileptic behaviors of each rat were observed individually. Latency of first convulsion decreased in control–morphine and stressed–saline groups while increased in stressed–morphine rats compared to control–saline group on P15 (P=0.04). Number of tonic–clonic seizures significantly increased in control–morphine and stressed–saline rats compared to control–saline group at P15 (P=0.02). PBL increased in stressed–saline, control–morphine and stress–morphine groups compared to control–saline rats. It can be concluded that prenatal exposure of rats to forced-swim stress and morphine changed their susceptibility to PTZ-induced seizure and PBL during infancy and prepubertal period. Co-administration of morphine attenuated effect of stress on epileptic behaviors.

The Effect of the Medicinal Mushrooms Agaricus brasiliensis and Ganoderma lucidum (Higher Basidiomycetes) on the Erythron System in Normal and Streptozotocin-Induced Diabetic Rats.

Diabetes mellitus (DM) is accompanied by the development of hypoxia, which disturbs the physicochemical properties of the erythrocyte membrane and further leads to the occurrence of anemia and a reduction of the lifespan. In response, the body activates compensatory reactions directed at a renewal of the red blood cell pool and an increase in tissue oxygenation. In this study the influence of Agaricus brasiliensis and Ganoderma lucidum medicinal mushroom mycelia on the erythron system of control and streptozotocin (STZ)-induced diabetic rats were investigated. Wistar outbred white male rats were intraperitoneally injected with saline (control rats) or STZ (50 mg/kg, DM rats) and orally treated with placebo or submerged culture mycelium powder (1 g/kg/day) for 2 weeks. Peripheral blood erythrocytes were collected. Hypoglycemic effects of A. brasiliensis and G. lucidum occurred in the diabetic rats, as evidenced by decreased blood glucose and glycosylated hemoglobin concentrations. In STZ-diabetic animals treated with submerged culture mycelium powder, an increase in the number of erythrocytes in the bloodstream (an antianemic effect), erythrocyte resistance to acid hemolysis, and the normalization of fetal hemoglobin concentrations, along with the intensification of erythropoiesis were observed. In conclusion, our results suggest that in diabetic animals A. brasiliensis and G. lucidum have therapeutic effects that manifest in hypoglycemic and antianemic action.

Decreased Expression of Aquaporin-1 in Lung Tissue of Silicotic Rats.

Aquaporin-1 (AQP-1), found in the early 1990s, a water channel protein in the cell membranes of mammals, has been reported to play an important role in water balance of the respiratory system. However, there are a few studies about the role of AQP in occupational pulmonary disease such as silicosis. This study is to explore the information of aquaporin-1 (AQP-1) in the pathogenesis of silicosis by examining AQP expression, distribution, and location in the lung tissue of a silicotic rat model. Male Wistar SPF rats were divided randomly into the following 8 groups (n = 8 per group): (1) saline control group: instillation of 1 mL sterile physiological saline; (2) silica groups (ld, 7d, 14d, 28d, 42d, 56d): instillation of a suspension of 50 mg silica dust in a total volume of 1 mL sterile physiological saline; (3) the normal control group without treatment. Immunohistochemistry, immunofluorescence, and western blot were used to detect distribution and expression of AQP-1 in the lung tissue of rats exposed to silica. The expression of AQP-1 between normal and the saline control rats showed no significant difference, but was decreased in the silicotic model rats' lung. The expression of AQP-1 decreased in silicotic rats, which suggests that AQP-1 may play an important role in the formation of silicosis.

Social learning reverses conditioned aversion to nicotine via oxytocin receptor in the infralimbic cortex in rats (839.4)

The initial aversive response to cigarettes in most smokers is thought to be ameliorated by the presence of peers who also smoke. We previously reported that social learning (i.e. detecting an odor cue associated with nicotine from peer rats) reversed the development of conditioned taste aversion (CTA) between self‐administered nicotine and a contingent olfactogustatory cue in adolescent rats (PMID:21796102). Herein, we focused on the role of oxytocin in infralimbic cortex (IL) as a mechanism for the effect of social learning. Bilateral lesions of IL before (F1,13=18.1, p=0.001) but not after (F1,16=0.481, p&gt;0.05) self‐administration reduced the effect of social learning on tests of CTA. We also found that carbon disulfide contained in rats’ breath was the signal mediating social learning and its effect also required intact IL. We then infused a specific oxytocin receptor antagonist, desGly‐NH2‐d(CH2)5[D‐Tyr]OVT, (0.028 ~ 2.8 μg/ml, a gift from Dr. Mannings) bilaterally into the IL, which dose‐dependently reduced the effect of carbon disulfide on nicotine CTA (F4,30=3.9, p=0.01). The highest dose of the antagonist had no effect on i.v. saline control rats. These results suggested that the release of oxytocin in the IL is a critical signal that reversed the conditioned aversion between nicotine and its olfactogustatory cue, which in turn enabled voluntary nicotine intake in adolescent rats.Key words: nicotine; aversion; social learning; infralimbic cortex; oxytocin

Anti-Platelet Therapy with Clopidogrel Prevents Endothelial Dysfunction and Vascular Remodeling in Aortas from Hypertensive Rats

The aim was to investigate the beneficial effects of clopidogrel in thoracic aorta function and structure and to characterize if P2Y12 receptors contribute to these effects. Male Sprague Dawley rats were infused with angiotensin II [(Ang II) 60 ng.min−1, 14 days] or saline (control rats) and were simultaneously treated with clopidogrel (10 mg.kg−1.day−1) or vehicle. After 14 days, systolic blood pressure (mmHg) was similar in Ang II-hypertensive rats treated with clopidogrel or vehicle (199±9 vs. 190±11, respectively). Systolic blood pressure in control rats was not altered by clopidogrel treatment (128±1 vs. vehicle, 134±2). Endothelium-dependent relaxation induced by 2-MeS-ADP was decreased in aortas from vehicle-treated Ang II-hypertensive rats, compared to vehicle-treated control rats. This response was elicited via activation of P2Y1 and P2Y12 receptors. In the presence of L-NAME and indomethacin, 2-MeS-ADP induced contraction and this response was augmented in vehicle-treated Ang II-hypertensive rats, compared to vehicle-treated control rats. The contraction to 2-MeS-ADP was evoked by P2Y13 and P2Y12 receptor activation. Clopidogrel-treatment did not normalize relaxation or contractile responses induced by 2-MeS-ADP in aortas from Ang II-hypertensive rats. P2Y1 and P2Y12 protein expression was increased, whereas P2Y13 receptor expression was reduced in aorta from vehicle-treated Ang II-hypertensive rats. Endothelium-dependent relaxation upon acetylcholine-stimulation was reduced in vehicle-treated Ang II-hypertensive rats, and clopidogrel treatment was effective in improving endothelial function. Clopidogrel also prevented vascular remodeling, evidenced by augmented media thickness in aortas from Ang II-hypertensive rats. Clopidogrel has beneficial effects on the aortic endothelium of Ang II-hypertensive rats, but its effects do not seem to be directly related to the presence of P2Y12 receptors in this vessel.