Acute foot-shock stress decreased seizure susceptibility against pentylenetetrazole-induced seizures in mice: Interaction between endogenous opioids and cannabinoids.

BACKGROUND: Stressful conditions affect the brain's neurotransmission and neural pathways that are involved in seizure susceptibility. Stress alters the intensity and/or frequency of seizures. Although evidence indicates that chronic stress exerts proconvulsant effects and acute stress has anticonvulsant properties, the underlying mechanisms which mediate these effects are not well understood. In the present study, we assessed the role of endogenous opioids, endocannabinoids, as well as functional interaction between opioid and cannabinoid systems in the anticonvulsant effects of acute foot-shock stress (FSS) against pentylenetetrazole (PTZ)-induced seizures in mice. METHODS: Prolonged intermittent FSS was chosen as an acute stress model. Seizure threshold was determined after 30 min of stress induction in male Naval Medical Research Institute (NMRI) mice (20-30 g). Opioid and cannabinoid receptor antagonists were administered before animal placement in the FSS apparatus. RESULTS: Acute FSS significantly decreased seizure susceptibility in animals. The administration of the cannabinoid receptor 1 (CB1) antagonist, AM251, completely blocked the anticonvulsant effect of acute FSS at the doses of 1 pg/kg-100 µg/kg but not at 1 fg/kg. Pretreatment with the nonspecific opioid receptor antagonist, naltrexone (NTX), significantly inhibited the anticonvulsant effects of acute FSS at 1 and 2 mg/kg but not at 0.3 mg/kg. However, coadministration of the subeffective doses of AM251 (1 fg/kg) and NTX (0.3 mg/kg) reversed the anticonvulsant effects of acute FSS. CONCLUSIONS: Opioid and cannabinoid systems are involved in the anticonvulsant effects of acute FSS, and these neurotransmission systems interact functionally in response to acute FSS.

Anticonvulsant effect of minocycline on pentylenetetrazole-induced seizure in mice: involvement of nitric oxide and N-methyl-d-aspartate receptor.

Anticonvulsant effects of minocycline have been explored recently. This study was designed to examine the anticonvulsant effect of acute administration of minocycline on pentylenetetrazole-induced seizures in mouse considering the possible role of the nitric oxide/N-methyl-d-aspartate (NMDA) pathway. We induced seizure using intravenous administration of pentylenetetrazole. Our results showed that acute administration of minocycline increased the seizure threshold. Furthermore, co-administration of subeffective doses of the nonselective nitric oxide synthase (NOS) inhibitor NG-l-arginine methyl ester (10 mg/kg) and the neuronal NOS inhibitor 7-nitroindazole (40 mg/kg) enhanced the anticonvulsant effect of subeffective doses of minocycline (40 mg/kg). We found that inducible NOS inhibitor aminoguanidine (100 mg/kg) had no effect on the antiseizure effect of minocycline. Moreover, l-arginine (60 mg/kg), as a NOS substrate, reduced the anticonvulsant effect of minocycline. We also demonstrated that pretreatment with the NMDA receptor antagonists ketamine (0.5 mg/kg) and MK-801 (0.05 mg/kg) increased the anticonvulsant effect of subeffective doses of minocycline. Results showed that minocycline significantly decreased the hippocampal nitrite level. Furthermore, co-administration of a neuronal NOS inhibitor like NMDA receptor antagonists augmented the effect of minocycline on the hippocampal nitrite level. In conclusion, we revealed that anticonvulsant effect of minocycline might be, at least in part, due to a decline in constitutive hippocampal nitric oxide activity as well as inhibition of NMDA receptors.

Protective effects of gabapentin against the seizure susceptibility and comorbid behavioral abnormalities in the early socially isolated mice.

Adolescence is a pivotal period of brain development during lifespan, which is sensitive to stress exposure. Early social isolation stress (SIS) is known to provoke a variety of psychiatric comorbidities as well as seizure risk. Psychiatric comorbidities present challenging dilemmas for treatment and management in people with seizure disorders. In this study, we aimed to investigate whether gabapentin (GBP) as an anti-epileptic drug is able to alleviate the seizure activity as well as comorbid behavioral abnormalities in socially isolated mice. Results showed that early SIS induced proconvulsant effects along with depressive, aggressive and anxiety-like behaviors. Whereas the administration of both acute and chronic GBP at sub-effective doses produced no alterations in the behavioral profile of socially conditioned counterparts the same treatments effectively reversed the seizure susceptibility to pentylenetetrazole and behavioral deficits in isolated mice. Results of the study indicate that 1) Early SIS could be considered as an animal model of psychosocial stress to investigate the psychiatric comorbidities in seizure disorders, 2) Chronic administration of low dose GBP prevented the shaping of behavioral abnormalities in adulthood, 3) Chronic administration of low dose GBP produced no negative behavioral effects in socially conditioned mice suggesting the safety of the drug, 4) Gabapentin at low doses may be considered as an agent for management of epilepsy in individuals with psychiatric comorbidities.

Involvement of ATP-sensitive potassium channels and the opioid system in the anticonvulsive effect of zolpidem in mice.

Zolpidem is a hypnotic medication that mainly exerts its function through activating γ-aminobutyric acid (GABA)A receptors. There is some evidence that zolpidem may have anticonvulsive effects. However, the mechanisms underlying this effect have not been elucidated yet. In the present study, we used the pentylentetrazole (PTZ)-induced generalized seizure model in mice to investigate whether zolpidem can affect seizure threshold. We also further evaluated the roles of ATP-sensitive potassium (KATP) channels as well as µ-opioid receptors in the effects of zolpidem on seizure threshold. Our data showed that zolpidem in a dose-dependent manner increased the PTZ-induced seizure threshold. The noneffective (i.e., did not significantly alter the PTZ-induced seizure threshold by itself) doses of KATP channel blocker (glibenclamide) and nonselective opioid receptor antagonist (naloxone) were able to inhibit the anticonvulsive effect of zolpidem. Additionally, noneffective doses of either KATP channel opener (cromakalim) or nonselective µ-opioid receptor agonist (morphine) in combination with a noneffective dose of zolpidem exerted a significant anticonvulsive effect on PTZ-induced seizures in mice. A combination of noneffective doses of naloxone and glibenclamide, which separately did not affect zolpidem effect on seizure threshold, inhibited the anticonvulsive effects of zolpidem. These results suggest a role for KATP channels and the opioid system, alone or in combination, in the anticonvulsive effects of zolpidem.

Atorvastatin attenuates the antinociceptive tolerance of morphine via nitric oxide dependent pathway in male mice.

The development of morphine-induced antinociceptive tolerance limits its therapeutic efficacy in pain management. Atorvastatin, or competitive inhibitor of 3-hydroxy-methyl-glutaryl coenzyme A (HMG-CoA) reductase, is mainstay agent in hypercholesterolemia treatment. Beyond the cholesterol-lowering activity, exploration of neuroprotective properties of this statin indicates its potential benefit in central nervous disorders. The aim of the present study was to assess the effects of atorvastatin in development and expression of morphine-induced analgesic tolerance in male mice and probable involvement of nitric oxide. Chronic and acute treatment with atorvastatin 10 and 20mg/kg, respectively, could alleviate morphine tolerance in development and expression phases. Chronic co-administration of nitric oxide synthase (NOS) inhibitors including L-NAME (non selective NOS inhibitor; 2mg/kg), aminoguanidine (selective inducible NOS inhibitor; 50mg/kg) and 7-NI (selective neuronal NOS inhibitor; 15mg/kg) with atorvastatin blocked the protective effect of atorvastatin in tolerance reversal. Moreover, reversing the atorvastatin effect was also observed in acute simultaneous treatment of L-NAME (5mg/kg) and aminoguanidine (100mg/kg) with atorvastatin. Co-treatment of guanylyl cyclase inhibitor, ODQ (chronic dose: 10mg/kg and acute dose: 20mg/kg) was associated with prevention of atorvastatin anti-tolerance properties. Our results revealed that the atorvastatin modulating role in morphine antinociceptive tolerance is mediated at least in part via nitric oxide in animal pain models of hot plate and tail flick.

Lithium attenuates the proconvulsant effect of adolescent social isolation stress via involvement of the nitrergic system.

In this study, we tested whether acute administration of lithium mitigates the deleterious effect of adolescent social isolation stress (SIS) on seizure susceptibility. In comparison with socially conditioned (SC) mice, isolated conditioned (IC) mice exhibited an increase in seizure susceptibility to pentylenetetrazole. Acute administration of lithium (10mg/kg) reversed the proconvulsant effect of SIS in IC mice, but this effect was not observed in SC mice. Coadministration of subthreshold doses of lithium (3mg/kg) with nitric oxide synthase (NOS) inhibitors reversed the effect of SIS on seizure susceptibility and decreased hippocampal nitrite levels in IC animals. In addition, a subthreshold dose of a nitric oxide precursor reduced the protective effect of lithium on seizure susceptibility and increased nitrite levels in the hippocampus of IC mice. These results suggest that lithium exerts a protective influence against the proconvulsant effect of adolescent SIS via a nitrergic system that includes activation of neuronal NOS in the hippocampus.

NMDA receptor antagonists attenuate the proconvulsant effect of juvenile social isolation in male mice.

Experiencing psychosocial stress in early life, such as social isolation stress (SIS), is known to have negative enduring effects on the development of the brain and behavior. In addition to anxiety and depressive-like behaviors, we previously showed that juvenile SIS increases susceptibility to pentylenetetrazole (PTZ)-induced seizures in mice through enhancing the nitrergic system activity in the hippocampus. In this study, we investigated the possible involvement of N-methyl-D-aspartate (NMDA) receptors in proconvulsant effects of juvenile SIS. Applying 4 weeks of SIS to juvenile male mice at postnatal day 21-23, we observed an increased susceptibility to PTZ as well as anxiety and depressive-like behaviors in adult mice. Intraperitoneal (i.p.) administration of NMDA receptor antagonists, MK-801 (0.05 mg/kg) and ketamine (0.5mg/kg), reversed the proconvulsant effects of SIS in Isolated (and not social) housed animals. Co-administration of non-effective doses of nitric oxide synthase (NOS) inhibitors, 7NI (25mg/kg) and L-NAME (10mg/kg), with NMDA receptor antagonists, MK-801 (0.01 mg/kg) and ketamine (0.1mg/kg) attenuated the proconvulsant effects of juvenile SIS only in isolated housed mice. Also, using real time RT-PCR, we showed that hippocampal upregulation of NR2B subunit of NMDA receptor may play a critical role in proconvulsant effects of juvenile SIS by dysregulation of NMDA/NO pathway. In conclusion, results of present study revealed that experiencing SIS during adolescence predisposes the co-occurrence of seizure disorders with psychiatric comorbidities and also, alteration of NMDA receptor structure and function in hippocampus plays a role in proconvulsant effects of juvenile SIS through enhancing the NMDA/NO pathway.

Possible involvement of nitrergic and opioidergic systems in the modulatory effect of acute chloroquine treatment on pentylenetetrazol induced convulsions in mice.

Chloroquine has long been used for the treatment of malaria and rheumatological disorders. Accumulating evidence suggests potential use of chloroquine as a neuroprotective agent. Several studies have reported that endogenous opioids and nitric oxide (NO) system mediate the chloroquine effects. In the present study, the involvements of endogenous opioids and NO in the modulatory effects of chloroquine on pentylenetetrazol-induced seizures were assessed in mice. Chloroquine 5mg/kg significantly increased the seizure threshold, but this effect was reversed with naltrexone 1mg/kg. Acute co-administration of l-NAME (non-selective NO synthase (NOS) inhibitor, 5mg/kg) or 7-NI (selective neuronal NOS inhibitor, 40 mg/kg) with the effective dose of chloroquine completely inhibited its anticonvulsant effects. Acute single injection of a sub-effective dose of l-arginine (NO precursor, 60 mg/kg) with a sub-effective dose of chloroquine 2.5mg/kg increased the seizure threshold but administration of L-arginine 60 mg/kg with chloroquine 10mg/kg decreased the seizure threshold. Moreover, the combination of the lower doses of naltrexone (0.1mg/kg) and 7-NI (15 mg/kg) showed additive effects in blocking the chloroquine-induced anticonvulsant properties. Chloroquine 5mg/kg enhanced the hippocampal nitrite levels. Chloroquine at the dose of 20mg/kg decreased the seizure threshold. This effect was inhibited through L-NAME (5mg/kg), 7-NI (40 mg/kg) and naltrexone (1mg/kg) administration with this dose of chloroquine. In conclusion, NO signaling probably through neuronal NOS, but not inducible NOS could be involved in the opioid-dependent anticonvulsant effects of chloroquine in this model of seizures in mice. It seems that nitric oxide and opioid systems are involved in modulatory effect of chloroquine on seizures induced by pentylenetetrazol.

Morphine modulates the effects of histamine H1 and H3 receptors on seizure susceptibility in pentylenetetrazole-induced seizure model of mice.

Histamine regulates release of neurotransmitters such as dopamine, serotonin, gamma-aminobutyric acid (GABA), glutamate and also is involved in several functions in central nervous system (CNS). It has been shown that histamine participates in disorders like seizure. It has been well documented that morphine dose-dependently induces anti or proconvulsant effects. In the current study, we firstly showed that morphine (1mg/kg) exerts anticonvulsant effects which significantly reversed by naltrexone administration. Secondly, we determined seizure threshold for H1 and H3 receptors agonists and antagonists in mouse model of pentylenetetrazole (PTZ)-induced clonic seizures. Our results showed that activation of H1 receptors by 2-(2-Pyridyl)-ethylamine exerts anticonvulsant properties while inhibition of H1 receptors by pyrilamine maleate induced proconvulsant effects. Furthermore, we showed that immepip dihydrobromide, a H3 receptor agonist, increased seizure susceptibility to PTZ whereas thioperamide, a H3 receptor antagonist increased seizure threshold. We also revealed that pretreatment with morphine potently reversed the effects of histaminergic system on seizure threshold suggesting the involvement of opioid system in alteration of seizure threshold by histaminergic drugs.

Experiencing neonatal maternal separation increased the seizure threshold in adult male mice: Involvement of the opioid system.

Experiencing early-life stress has been considered as a potent risk factor for the development of many of brain disorders, including seizures. Intervening mechanisms through which neonatal maternal separation (MS) alters the seizure susceptibility in adulthood have not been well studied. In the current study, by applying 180 min of MS stress (PND 2-14), we determined the seizure susceptibility and considered the role of the opioid system. Maternal separation increased the seizure threshold, and administration of anticonvulsant/proconvulsant doses of morphine (1 and 30 mg/kg, respectively) reversed the impact of MS. Using tail flick and hot plate tests, we exposed animals to 30 min Restraint stress (RS) and found that MS decreased the pain threshold, suggesting the hyporesponsiveness of the opioid system. These results supported the abnormal seizure activity observed in the MS mice and suggested that abnormalities in the opioid system following MS alter seizure susceptibility in later life.

Co-occurrence of anxiety and depressive-like behaviors following adolescent social isolation in male mice; possible role of nitrergic system.

Approximately more than 50% of patients with depression have the co-occurrence of anxiety, which complicates the treatment of disease. Recently, social isolation stress (SIS) paradigm has been suggested as an animal model to investigate the underlying mechanism involved in depression-anxiety co-occurrence. In this study, applying six weeks of SIS to adolescent mice, we tested whether nitrergic system plays a role in co-occurrence of depression and anxiety. In this study, comparisons between socially and isolated conditioned (SC and IC) animals showed that SIS induces behaviors relevant to depression and anxiety in IC mice and in addition, nitrergic system is involved in mediating the negative outcomes of SIS. Administration of subeffective doses of aminoguanidine (a specific inducible nitric oxide synthase inhibitor or iNOS, 50mg/kg) and L-NAME (non-specific inhibitor of NOS, 10mg/kg) significantly reversed the negative effects of SIS on behavioral profile as well as nitrite levels in the cortex of IC mice, Although administration of subeffective dose of 7-nitroindazole (a specific neuronal NOS inhibitor, 25mg/kg) decreased the nitrite levels in the hippocampus, but had no effect on depressant and anxiogenic effects of SIS. Results of this study confirmed that SIS is an appropriate animal model to investigate the potential mechanisms in depression-anxiety co-occurrence. We also showed that nitrergic system has contributed to co-occurrence of depression and anxiety in IC mice as an underlying mechanism.

Proconvulsant effect of post-weaning social isolation stress may be associated with dysregulation of opioid system in the male mice.

Opioid system has been reported to be involved in the consequences of post-weaning social isolation stress (SIS) such as hypoalgesia and social behaviors. Also, previous studies have shown that SIS increases mu opioid receptor expression in the regions of the brain associated with epileptogenesis such as basolateral amygdala and cortex. Interestingly, experiencing SIS increases seizure risk in the adulthood. Regarding the SIS-induced alterations in the opioid system, we hypothesize that increase in opioidergic system activity (mostly by mu receptor) may be associated with increase in vulnerability to seizures. In non-stressed mice, morphine at low doses (1 mg/kg) has an anticonvulsant effect on seizure threshold while higher doses (60 mg/kg) are proconvulsant. To support the hypothesis, we showed that administration of anticonvulsant dose of morphine (1 mg/kg) to socially isolated male mice not only was not able to reverse the negative effect of SIS on seizure susceptibility to pentylenetetrazole but also enhanced it. These results support our hypothesis that proconvulsant effect of post-weaning social isolation stress may be associated with dysregulation of opioid system in the adult male mice.

Involvement of the nitrergic system in the proconvulsant effect of social isolation stress in male mice.

Social isolation stress (SIS) in adolescence is accompanied by neurobehavioral disturbances and pathophysiological changes in certain regions of the CNS such as the hippocampus. In this study, we tested whether SIS impacts seizure susceptibility in postnatal male mice due to a role of hippocampal nitric oxide (NO). To do this, we used the pentylenetetrazole (PTZ) model of clonic seizures, open-field test, hole-board test, forced swimming test, and plasma corticosterone assay. We aimed to evaluate if 4 weeks of SIS is capable of decreasing seizure threshold along with altering affective and neuroendocrine responses in isolated conditioned (IC) animals in comparison with socially conditioned (SC) animals. In addition, we applied subeffective doses of NO precursor L-arginine (25, 50, and 100mg/kg) and NOS inhibitors 7-NI (15 and 40 mg/kg), aminoguanidine (50 and 100mg/kg), and L-NAME (10 and 15 mg/kg) to both IC and SC groups prior to the determination of seizure threshold. Injection of a single dose of all mentioned drugs did not induce changes in seizure threshold of SC mice. On the other hand, L-NAME and 7-NI, but not aminoguanidine, modulated the proconvulsant effect of SIS, while L-arginine augmented the latter effect. We also measured the hippocampal nitrite levels after the administration of the aforementioned drugs. Social isolation stress increased the nitrite levels in comparison with those in SC mice, whereas 7-NI and L-NAME, unlike aminoguanidine, mitigated the effect of SIS. Additionally, L-arginine boosted the effects of SIS on nitrite production. In summary, we showed that SIS enhanced seizure susceptibility in the PTZ model of clonic seizures through the activation of the nitrergic system in the hippocampus. Also, we proved that nNOS, but not iNOS, accounts for these changes following SIS.