Pharmacological activation of mediodorsal thalamic GABA-A receptors modulates morphine/cetirizine-induced changes in the prefrontal cortical GFAP expression in a rat model of neuropathic pain.

The present study investigated the involvement of mediodorsal thalamic (MD) GABA-A receptors in cetirizine/morphine-induced anti-allodynia using a rat model of neuropathic pain. To assess the importance of the prefrontal cortex (PFC) for chronic pain processing, its expression level changes of glial fibrillary acidic protein (GFAP) were measured following drug treatments. Each animal was subjected to chronic constriction of the sciatic nerve surgery simultaneously with the MD cannulation under stereotaxic surgery. The results showed that the administration of morphine (3-5 mg/kg) or cetirizine (1-3 mg/kg) produced significant analgesia in neuropathic rats. Systemic administration of cetirizine (2.5 and 3 mg/kg) potentiated the analgesic response to a low and intolerance dose of morphine (3 mg/kg). Intra-MD microinjection of muscimol, a selective GABA-A receptor agonist (0.005-0.01 µg/rat), increased the cetirizine/morphine-induced anti-allodynia, while muscimol by itself did not affect neuropathic pain. The neuropathic pain was associated with the increased PFC expression level of GFAP, suggesting the impact of chronic pain on PFC glial management. Interestingly, the anti-allodynia was associated with a decrease in the PFC expression level of GFAP under the drugs' co-administration. Thus, cetirizine has a significant potentiating effect on morphine response in neuropathic pain via interacting with the MD GABA-A receptors. It seems that neuropathic pain affects the prefrontal cortex GFAP signaling pathway. In clinical studies, these findings can be considered to create a combination therapy with low doses of GABA-A receptor agonist plus cetirizine and morphine to manage neuropathic pain.

Ameliorating effect offluoxetine on tamoxifen-induced memory loss: The role of corticolimbic NMDA receptors and CREB/BDNF/cFos signaling pathways in rats.

Tamoxifen-induced cognitive dysfunction may lead to fluoxetine consumption in patients with breast cancer. Since the brain mechanisms are unclear in tamoxifen/fluoxetine therapy, the blockade effect of hippocampal/amygdala/prefrontal cortical NMDA receptors was examined in fluoxetine/tamoxifen-induced memory retrieval. We also assessed the corticolimbic signaling pathways in memory retrieval under the drug treatment in adult male Wistar rats. Using the Western blot technique, the expression levels of the cAMP response element-binding protein (CREB), brain-derived neurotrophic factor (BDNF), and cFos were evaluated in the corticolimbic regions. The results showed that pre-test administration of fluoxetine (3 and 5 mg/kg, i.p.) improved tamoxifen-induced memory impairment in the passive avoidance learning task. Pre-test bilateral microinjection of D-AP5, a selective NMDA receptor antagonist, into the dorsal hippocampal CA1 regions and the central amygdala (CeA), but not the medial prefrontal cortex (mPFC), inhibited the improving effect of fluoxetine on tamoxifen response. It is important to note that the microinjection of D-AP5 into the different sites by itself did not affect memory retrieval. Memory retrieval increased the signaling pathway of pCREB/CREB/BDNF/cFos in the corticolimbic regions. Tamoxifen-induced memory impairment decreased the hippocampal/PFC BDNF level and the amygdala level of pCREB/CREB/cFos. The improving effect of fluoxetine on tamoxifen significantly increased the hippocampal/PFC expression levels of BDNF, the PFC/amygdala expression levels of cFos, and the ratio of pCREB/CREB in all targeted areas. Thus, NMDA receptors' activity in the different corticolimbic regions mediates fluoxetine/tamoxifen memory retrieval. The corticolimbic synaptic plasticity changes likely accompany the improving effect of fluoxetine on tamoxifen response.

Post-Weaning Treatment with Probiotic Inhibited Stress-Induced Amnesia in Adulthood Rats: The Mediation of GABAergic System and BDNF/c-Fos Signaling Pathways.

The current study aimed to examine the effect of post-weaning treatment with probiotics on memory formation under stress during the adult period in male Wistar rats. Considering GABA is a potential mediator between probiotics and the host, the present study also investigated the involvement of the GABAergic system in the probiotic response. The hippocampal and prefrontal cortical (PFC) expression levels of BDNF and c-Fos were also assessed to show whether the treatments affect the memory-related signaling pathway. Three weeks after birth, the post-weaning rats were fed with probiotic water (PW) or tap water (TW) for 2, 3, 4, or 5 weeks. Exposure to acute stress impaired memory formation in a passive avoidance learning task. Feeding the post-weaning animals with probiotic strains (3, 4, or 5 weeks) inhibited stress-induced amnesia of the adult period. Post-training intracerebroventricular (ICV) microinjection of muscimol improved stress-induced amnesia in the animals fed with TW. ICV microinjection of muscimol inhibited probiotic treatment's significant effect on the stress response in the memory task. The expression levels of BDNF and c-Fos in the PFC and the hippocampus were significantly decreased in the stress animal group. The levels of BDNF and c-Fos were increased in the PW/stress animal group. The muscimol response was compounded with the decreased levels of BDNF and c-Fos in the PFC and the hippocampus. Thus, the GABA-A receptor mechanism may mediate the inhibitory effect of this probiotic mixture on stress-induced amnesia, which may be associated with the PFC and hippocampal BDNF/c-Fos signaling changes.

Ventral hippocampal NMDA receptors mediate the effects of nicotine on stress-induced anxiety/exploratory behaviors in rats.

Brief exposure to stress increases the tendency to drug abuse, especially cigarette smoking. It seems that nicotine abuse alleviates some psychological and physiological stress symptoms. The present study investigated the effect of nicotine administration on stress-induced anxiety-like behavior in adult male Wistar rats. Also, the possible role of the ventral hippocampal (VH) glutamatergic NMDA receptors was examined in the stress-induced anxiety-like behavior under nicotine administration. The anxiogenic-like effects of forced swimming stress (10 min) were shown by decreases in the head-dipping behavior, rearing, and locomotor activity in a hole-board task. Interestingly, the administration of the different doses of nicotine (0.075 and 0.1 mg/kg, i.p.) inhibited stress-induced anxiogenic-like behaviors. Bilateral microinjection of NMDA (0.1 µg/rat) into the VH potentiated the response of an ineffective dose of nicotine (0.05 mg/kg, i.p.) on stress-induced anxiety-like behavior. The microinjection of D-AP5 (1.5 and 2 µg/rat) into the VH inhibited the response of an effective dose of nicotine (0.1 mg/kg, i.p.) on anxiety-like behavior induced by acute stress. Intra-VH microinjection of D-AP5 reversed the potentiating effect of NMDA on nicotine response. Intra-VH microinjection of NMDA or D-AP5 by itself did not affect stress-induced anxiety-like behavior. Taken together, we can conclude that nicotine inhibited stress-induced anxiogenic-like behaviors, possibly via the ventral hippocampal NMDA receptors mechanism.

miR-21: a promising biomarker for the early detection of colon cancer.

PURPOSE: The aim of this study was to compare the expression of miR-21 gene in stages II-IV of formalin-fixed paraffin-embedded (FFPE) tissue in patients with colon cancer and introduce miR-21 as a potential molecular marker for detection of colon cancer in the early stages. INTRODUCTION: Currently, identification of key molecules involved in the pathogenesis of cancer is one of the areas under consideration. miRNAs, are small RNAs which have been identified in many cancers. In this study, we investigated the expression of miR-21 in three pathologic stages in patients with colon cancer in the north of Iran. PATIENTS AND METHODS: A total of 40 FFPE samples were obtained from patients with stages II, III, and IV from hospitals in Mazandaran and Golestan provinces. After extraction of RNA, treatment with DNase I and cDNA synthesis was performed and miR-21 expression was assessed by qPCR. Then, the data were analyzed using statistical software R (3.4.3). RESULTS: The expression of miR-21 in stage II was significantly different from stage IV. However, no significant difference was observed between the other stages. In stage II, the level of miR-21 expression was higher in men than women. Moreover, in the second pathological stage, miR-21 expression was reduced in patients with adjacent lymphoid tissue engagement. In addition, the expression of miR-21 in grade I was significantly higher than grade II. CONCLUSION: The results of this study suggest that miR-21 can be a diagnostic marker for early stages of colon cancer, especially in men. It can also be considered as a good candidate for targeted treatment of colon cancer in the early stages of the disease. Furthermore, for the first time, we suggested that miR-21 can be a good molecular marker for classification of the stages of colon cancer.

Corticolimbic analysis of microRNAs and protein expressions in scopolamine-induced memory loss under stress.

The aim of the present study was to assess thealterations of corticolimbic microRNAs and protein expressions in the effect of scopolamine with or without stress on passive-avoidance memory in male Wistar rats. The expressions of miR-1, miR-10 and miR-26 and also the levels of p-CREB, CREB, C-FOS and BDNF in the prefrontal cortex (PFC), the hippocampus and the amygdala were evaluated using RT-qPCR and Western blotting techniques. The data showed that the administration of a muscarinic receptor antagonist, scopolamine or the exposure to 30 min stress significantly induced memory loss. Interestingly, the injection of an ineffective dose of scopolamine (0.5 mg/kg) alongside with exposure to an ineffective time of stress (10 min) impaired memory formation, suggesting a potentiative effect of stress on scopolamine response. Our results showed that memory formation was associated with the down-regulated expression of miR-1, miR-10 and miR-26 in the PFC and the hippocampus, but not the amygdala. The relative expression increase of miR-1 and miR-10 in the PFC and the hippocampus was shown in memory loss induced by scopolamine administration or 30-min stress. The PFC level of miR-10 and also hippocampal level of miR-1 and miR-10 were significantly up-regulated, while amygdala miR-1 and miR-26 were down-regulated in scopolamine-induced memory loss under stress. Memory formation increased BDNF, C-FOS and p-CREB/CREB in the PFC, the hippocampus and the amygdala. In contrast, the PFC, hippocampal and amygdala protein expressions were significantly decreased in memory loss induced by scopolamine administration (2 mg/kg), stress exposure (for 30 min) or scopolamine (0.5 mg/kg) plus stress (10 min). One of the most significant findings to emerge from this study is that the stress exposure potentiated the amnesic effect of scopolamine may via affecting the expressions of miRs and proteins in the PFC, the hippocampus and the amygdala. It is possible to hypothesis that corticolimbic signaling pathways play a critical role in relationship between stress and Alzheimer's disease.

Interactive effects of morphine and nicotine on memory function depend on the central amygdala cannabinoid CB1 receptor function in rats.

The present study investigated the possible involvement of the central amygdala (CeA) cannabinoid receptors type-1 (CB1Rs) in the interactive effects of morphine and nicotine on memory formation in a passive avoidance learning task. Our results showed that systemic administration of morphine (3 and 6mg/kg, s.c.) immediately after training phase impaired memory consolidation and induced amnesia. Administration of nicotine (0.3 and 0.6mg/kg, s.c.) before testing phase significantly restored morphine-induced amnesia, suggesting a cross state-dependent learning between morphine and nicotine. The results showed that while the administration of the lower dose of nicotine (0.1mg/kg, s.c.) per se did not induce a significant effect on morphine-induced amnesia, intra-CeA injection of arachidonylcyclopropylamide (ACPA), a cannabinoid CB1 receptor agonist (3 and 4ng/rat), significantly potentiated the nicotine response. Furthermore, the blockade of the CeA cannabinoid CB1 receptors by the injection of AM251 (0.75 and 1ng/rat) reversed the potentiative effect of nicotine (0.6mg/kg, s.c.) on morphine-induced amnesia. It should be considered that bilateral injection of the same doses of ACPA or AM251 (0.5-1ng/rat) into the CeA by itself had no effect on morphine response in a passive avoidance learning task. Confirmed by the cubic interpolation planes, the dose-response data revealed a cross-state-dependent learning between morphine and nicotine which may be mediated by the CeA endocannabinoid system via CB1 receptors.

Hippocampal nicotinic receptors have a modulatory role for ethanol and MDMA interaction in memory retrieval.

The aim of the current study was to examine the effect of dorsal hippocampal nicotinic acetylcholine receptors (nAChRs) activation on the functional interaction between ethanol and 3,4-methylenedioxy-N-methylamphetamine (MDMA or ecstasy) in memory retrieval. The dorsal hippocampal CA1 regions of adult male NMRI mice were bilaterally cannulated and memory retrieval was measured in a step-down type passive avoidance apparatus. Post-training or pre-test systemic administration of ethanol (1g/kg, i.p.) induced amnesia. Pre-test administration of ethanol reversed pre-training ethanol-induced amnesia, suggesting ethanol state-dependent learning. Pre-test intra-CA1 microinjection of different doses of MDMA (0.25-1µg/mouse) with an ineffective dose of ethanol (0.25g/kg, i.p.) also induced amnesia. Interestingly, pre-test intra-CA1 microinjection of MDMA (0.25-1µg/mouse) potentiated ethanol state-dependent learning. On the other hand, the activation of the dorsal hippocampal nAChRs by pre-test microinjection of nicotine (0.1-1µg/mouse, intra-CA1) improved amnesia induced by the co-administration of MDMD and ethanol. It is important to note that intra-CA1 microinjection of the same doses of MDMA or nicotine could not affect memory formation by itself. Pre-test intra-CA1 microinjection of nicotine (0.3-0.9µg/mouse) could not reverse amnesia induced by pre-training administration of ethanol while this treatment enhanced MDMA response on ethanol state-dependent learning. Thus, it can be concluded that there may be functional interactions among ethanol, MDMA and nicotine via the dorsal hippocampal nicotinic acetylcholine receptor mechanism in memory retrieval and drug state-dependent learning.

Alterations in the hippocampal phosphorylated CREB expression in drug state-dependent learning.

The present study investigated the possible alterations of hippocampal CREB phosphorylation in drug state-dependent memory retrieval. One-trial step-down passive avoidance task was used to assess memory retrieval in adult male NMRI mice. Pre-training administration of ethanol (1g/kg, i.p.) induced amnesia. Pre-test administration of ethanol (1g/kg, i.p) or nicotine (0.7 mg/kg, s.c.) reversed ethanol-induced amnesia, indicating ethanol- or ethanol-nicotine induced state-dependent learning (STD). Using Western blot analysis, it was found that the p-CREB/CREB ratio in the hippocampus increased in the mice that showed successful memory retrieval as compared with untrained mice. In contrast, pre-training administration of ethanol (1g/kg, i.p.) decreased the hippocampal p-CREB/CREB ratio in comparison with the control group. The hippocampal p-CREB/CREB ratio enhanced in ethanol- and ethanol-nicotine induced STD. Moreover, memory impairment induced by pre-training administration of WIN (1 mg/kg, i.p.) improved in the animals that received pre-test administration of WIN (1 mg/kg, i.p.), ethanol (0.5 g/kg, i.p.) or nicotine (0.7 mg/kg, s.c.), suggesting a cross STD between the drugs. The p-CREB/CREB ratio in the hippocampus decreased in the of WIN-induced amnesia and STD groups in comparison with the control group. In addition, cross state-dependent learning between WIN and ethanol or nicotine was associated with the increase of the hippocampal p-CREB/CREB ratio. It can be concluded that phosphorylation of CREB in the hippocampus is a critical event underlying the interaction of co-administration of drugs on memory retrieval in passive avoidance learning.

Dorsal hippocampal cannabinoid CB1 receptors mediate the interactive effects of nicotine and ethanol on passive avoidance learning in mice.

The present study evaluated the involvement of the dorsal hippocampal cannabinoid CB1 receptors in the combined effect of ethanol and nicotine on passive avoidance learning in adult male mice. The results indicated that pre-training administration of ethanol (1 g/kg, i.p.) impaired memory retrieval. Pre-test administration of ethanol (0.5 and 1 g/kg, i.p.) or nicotine (0.5 and 0.7 mg/kg, s.c.) significantly reversed ethanol-induced amnesia, suggesting a functional interaction between ethanol and nicotine. Pre-test microinjection of a selective CB1 receptor agonist, ACPA (3 and 5 ng/mouse), plus an ineffective dose of ethanol (0.25 g/kg) or nicotine (0.3 mg/kg) improved memory retrieval, while ACPA by itself could not reverse ethanol-induced amnesia. Pre-test intra-CA1 microinjection of a selective CB1 receptor antagonist, AM251 (0.5-2 ng/mouse), did not lead to a significant change in ethanol-induced amnesia. However, pre-test intra-CA1 microinjection of AM251 prevented the ethanol (1 g/kg) or nicotine (0.7 mg/kg) response on ethanol-induced amnesia. In order to support the involvement of the dorsal hippocampal CB1 receptors in nicotine response, the scheduled mixed treatments of AM251 (0.1-1 ng/mouse), ACPA (5 ng/mouse) and nicotine (0.3 mg/kg) were used. The results indicated that AM251 reversed the response of ACPA to the interactive effects of nicotine and ethanol in passive avoidance learning. Furthermore, pre-test intra-CA1 microinjection of the same doses of ACPA or AM251 had no effect on memory retrieval. These findings show that the cannabinoid CB1 receptors of dorsal hippocampus are important in the combined effect of ethanol and nicotine on passive avoidance learning.

Morphine-induced anxiolytic-like effect in morphine-sensitized mice: involvement of ventral hippocampal nicotinic acetylcholine receptors.

In the present study, the effects of repeated intra-ventral hippocampal (intra-VH) microinjections of nicotinic acetylcholine receptor agonist or antagonist on morphine-induced anxiolytic-like behavior were investigated in morphine-sensitized mice using elevated plus-maze. Intraperitoneal (i.p.) administration of different doses of morphine (5, 7.5 and 10mg/kg) increased the percentage of open arm time (%OAT), open arm entries (%OAE), but not locomotor activity, indicating an anxiolytic-like response to morphine. The maximum response was obtained by 7.5mg/kg of the opioid. The anxiety-like behavior which was induced by a lower dose of morphine (5mg/kg) was significantly increased in mice that had previously received once daily injections of morphine (10 and 20mg/kg, i.p.) for 3 days. It should be considered that this treatment also increased locomotor activity in morphine-sensitized mice. Furthermore, the response to an ineffective dose of morphine (5mg/kg, i.p.) in the EPM was significantly increased in the animals that had previously received nicotine for 3 days (0.1, 0.3, 0.5 and 0.7 µg/mouse; intra-VH), 5 min prior to the injections of morphine (5mg/kg/day × 3 days; i.p.). On the other hand, the increase of morphine-induced anxiolytic-like effect in animals that had previously received the 3-day morphine (20mg/kg) was dose dependently suppressed by once daily injections of mecamylamine (0.5, 1 and 2 µg/mouse/day × 3 days; intra-VH). It is important to note that repeated intra-VH administrations of the same doses of nicotine or mecamylamine alone caused no significant change in morphine (5mg/kg)-induced anxiety-like parameters in the EPM. In conclusion, it seems that morphine sensitization affects the anxiety-like behavior in the EPM and the cholinergic system in the ventral hippocampus, via nicotinic receptors, may play an important role in this effect.

Ethanol state-dependent memory: involvement of dorsal hippocampal muscarinic and nicotinic receptors.

In the present study, the effects of bilateral injections of cholinergic agents into the hippocampal CA1 regions (intra-CA1) on ethanol state-dependent memory were examined in mice. A single-trial step-down passive avoidance task was used for the assessment of memory retention in adult male NMRI mice. Pre-training intraperitoneal injection (i.p.) of ethanol (0.25, 0.5 and 1g/kg) dose dependently induced impairment of memory retention. Pre-test administration of ethanol (0.5 and 1 g/kg, i.p.) induced state-dependent retrieval of the memory acquired under pre-training ethanol (1 g/kg, i.p.) influence. Pre-test intra-CA1 injection of physostigmine (2.5 and 5 microg/mouse, intra-CA1) or nicotine (0.3 and 0.5 microg/mouse, intra-CA1) improved pre-training ethanol (1g/kg)-induced retrieval impairment. Moreover, pre-test administration of physostigmine (2.5 and 5 microg/mouse, intra-CA1) or nicotine (0.3 and 0.5 microg/mouse, intra-CA1) with an ineffective dose of ethanol (0.25 g/kg) significantly restored the retrieval and induced ethanol state-dependent memory. Pre-test intra-CA1 injection of the muscarinic receptor antagonist, atropine (4 and 8 microg/mouse, intra-CA1) or the nicotinic receptor antagonist, mecamylamine (2 and 4 microg/mouse, intra-CA1) 5 min before the administration of ethanol (1g/kg, i.p.) dose dependently inhibited ethanol state-dependent memory. Pre-test intra-CA1 administration of physostigmine (0.5, 2.5 and 5 microg/mouse), atropine (2, 4 and 8 microg/mouse), nicotine (0.1, 0.3 and 0.5 microg/mouse) or mecamylamine (1, 2 and 4 microg/mouse) alone cannot affect memory retention. These findings implicate the involvement of a dorsal hippocampal cholinergic mechanism in ethanol state-dependent memory and also it can be concluded that there may be a cross-state dependency between ethanol and acetylcholine.