Chronic Intermittent Administration Research Articles

Hyperhomocysteinemia Increases Cortical Excitability and Aggravates Mechanical Hyperalgesia and Anxiety in a Nitroglycerine-Induced Migraine Model in Rats.

Homocysteine is a sulfur-containing endogenous amino acid leading to neurotoxic effects at high concentrations. Population studies suggest an association between plasma homocysteine levels and the risk of migraine headaches. The aim of this study was to analyze the sensitivity of rats with prenatal hyperhomocysteinemia (hHCY) in respect of the development of behavioral correlates of headache and spreading cortical depolarization (CSD) in a migraine model induced by the administration of the nitric oxide (NO) donor nitroglycerin. Animals with hHCY were characterized by migraine-related symptoms such as mechanical hyperalgesia, high-level anxiety, photophobia, as well as an enhanced level of neuronal activity in the somatosensory cortex along with a lower threshold of CSD generation. Likewise, acute or chronic intermittent administration of nitroglycerin also induced the development of mechanical allodynia, photophobia and anxiety in control groups. However, these symptoms were more pronounced in rats with hHCY. Unlike hHCY, nitroglycerin administration did not affect the threshold of CSD generation, but like hHCY, increased the background neuronal activity in layers 2/3 and 4 of the cerebral cortex. The latter was more pronounced in animals with hHCY. Thus, the migraine profile associated with hHCY can be further exaggerated in conditions with enhanced levels of migraine triggering the gaseous transmitter NO. Our data are consistent with the view that high levels of plasma homocysteine can act as a risk factor for the development of migraine.

Reduction of Postoperative Cognitive Deficits in Aged Mice by Chronic Intermittent Propofol

Postoperative cognitive dysfunction (POCD) can increase morbidity and mortality after surgery, especially in the elderly. Current evidence from both clinical/epidemiological studies and animal studies suggest that POCD is likely due to factors related to the surgery itself and at least not primarily due to the effects of general anesthetic agents. In contrast, propofol, the most commonly used intravenous general anesthetic agent which is acting mainly via GABAA receptors, has been shown to have neuroprotective effects by attenuating caspase activation, apoptosis, and mitochondrial dysfunction. We therefore set out to assess the effects of chronic intermittent propofol on postoperative cognitive deficits in aged mice. Abdominal surgery (exploratory laparotomy) under isoflurane anesthesia was performed in 21‐24 months‐old mice. Animals received either chronic intermittent propofol (75 mg/kg i.p.) or vehicle (IntralipidR) every 5th day throughout the experiment. The laparotomy was performed on day 17. Two days later, the mice were employed in the open field test. There was no difference in postoperative locomotor activity among groups (Surgery+Vehicle, Surgery+Propofol, No surgery+Vehicle, No surgery+Propofol). Then, cognitive functions were assessed using the Y maze, novel object recognition, contextual fear conditioning and Morris water maze tests. We found that laparotomy resulted in impaired learning and memory as determined in the behavioral test battery compared to no surgery controls. However, perioperative chronic intermittent propofol strongly attenuated the surgery‐induced memory impairments. Moreover, propofol per se improved the (likely age‐related) cognitive dysfunction in the no surgery controls. Our findings suggest that perioperative intermittent propofol administration might prevent or attenuate surgery‐induced cognitive dysfunction, and thus potentially reduce the liability for undesired long‐term outcomes. Thus, chronic intermittent administration of propofol may represent a novel therapeutic strategy with translational potential.

Acute and chronic ethanol administration: A potential model of behavioral sensitization using wheel-running in male CD1 mice

RationaleWe define behavioral sensitization as an augmented response to subsequent dosing after chronic intermittent administration of a drug. However, the biphasic effects of ethanol (EtOH), first stimulatory followed by depressive, make animal models of behavioral sensitization rare. ObjectivesThis study aimed to determine a dose of EtOH that did not depress wheel-running (WR) in CD1 mice and then to develop a model of EtOH-induced behavioral sensitization. MethodsFor the first part of this study, male CD1 mice (n = 24, 6/group) were administered either phosphate buffer saline (PBS), 0.5 g/kg, 1 g/kg, or 2 g/kg EtOH at a volume of 3 ml/kg, intraperitoneally (IP). Mice were divided into equal groups and received the weight-based dose once daily on Days 1, 2, 3, 4, and 5. All mice received a challenge dose of 0.5 g/kg on Day 10. In both parts of the study, mice were habituated to the running wheel for 5 min prior to dosing and wheel running was measured for 10 min after each dose. ResultsThe acute dose-response of EtOH effects on wheel running determined a significant difference between doses in wheel running (p < 0.05), with a post-hoc analysis establishing that 0.5 g/kg EtOH resulted in significantly more WR compared to 2 g/kg EtOH (p < 0.05). The chronic study demonstrated a significant main effect of Day (1 vs. 5 vs. Challenge, p < 0.001) and an interaction between Day and Treatment, with post-hoc analysis determining the effect to be between PBS and EtOH WR on Day 5 (p < 0.05). In addition, Bonferroni post-hoc analysis determined no differences between Days in the PBS condition, but a significant difference in the EtOH condition between Day 1 and Day 5 (p < 0.001) and that difference from Day 1 persisted when comparing to the Challenge Day (p < 0.01). ConclusionAfter chronic, intermittent, low dose administration of EtOH, male mice showed an increase in activity as measured by wheel running. Therefore, we laid the groundwork for a potentially useful rodent model for EtOH-induced behavioral sensitization.

Dose-Dependent Degeneration of Leydig Cells Following Kisspeptin-10 Administration: An Ultrastructural Study.

The discovery of kisspeptin signaling as a key regulator of gonadotropin- releasing hormone (GnRH) secretion from the hypothalamus enhanced our understanding of the neuroendocrine regulation of mammalian reproduction. Effects of central and peripheral administration of kisspeptin on plasma gonadotropins, testosterone, and spermatogenesis are studied in detail. The present study was conducted to check the ultrastructure of Leydig cells in prepubertal male rats in response to the administration of a range of kisspeptin doses. We administered a range of kisspeptin-10 doses (1 μg, 1 ηg, and 10 ρg) intraperitoneally to prepubertal male Sprague-Dawley rats (PND 35) twice daily after every 12 hours. Control rats were injected with physiological saline in parallel. At the end of the treatment, plasma concentrations of testosterone were measured by competitive binding radioimmunoassay, and small pieces of rat testicular tissue were processed for electron microscopy to examine the ultrastructure of Leydig cells. Plasma testosterone concentration was reduced significantly at 1ηg (P<0.05) and 1μg (P<0.01) doses as compared to control. Distinct ultrastructural changes categorized as dilatation of cytoplasmic organelles, irregularly shaped nuclei with nuclear membrane invaginations, reduced nuclear sizes, degeneration, and vacuolation were observed in the kisspeptin-10 treated Leydig cells as compared to control. Quantification of the data showed reduced Leydig cell indices and hyperplasia of the interstitial cells. It is concluded that chronic intermittent administration of kisspeptin-10 has a dose-dependent degenerative effect on the plasma testosterone levels and Leydig cells ultrastructure in prepubertal male rats.

Partially hydrolyzed guar gum attenuates non-alcoholic fatty liver disease in mice through the gut-liver axis.

BACKGROUNDThe gut-liver axis has attracted much interest in the context of chronic liver disease pathogenesis. Prebiotics such as dietary fibers were shown to attenuate non-alcoholic fatty liver disease (NAFLD) by modulating gut microbiota. Partially hydrolyzed guar gum (PHGG), a water-soluble dietary fiber, has been reported to alleviate the symptoms of various intestinal diseases and metabolic syndromes. However, its effects on NAFLD remain to be fully elucidated.AIMTo determine whether treatment with PHGG attenuates NAFLD development in mice through the gut-liver axis.METHODSSeven-week-old male C57BL/6J mice with increased intestinal permeability were fed a control or atherogenic (Ath) diet (a mouse model of NAFLD) for 8 wk, with or without 5% PHGG. Increased intestinal permeability was induced through chronic intermittent administration of low-dose dextran sulfate sodium. Body weight, liver weight, macroscopic findings in the liver, blood biochemistry [aspartate aminotransferase (AST) and alanine aminotransferase (ALT), total cholesterol, triglyceride, free fatty acids, and glucose levels], liver histology, myeloperoxidase activity in liver tissue, mRNA expression in the liver and intestine, serum endotoxin levels in the portal vein, intestinal permeability, and microbiota and short-chain fatty acid (SCFA) profiles in the cecal samples were investigated.RESULTSMice with increased intestinal permeability subjected to the Ath diet showed significantly increased serum AST and ALT levels, liver fat accumulation, liver inflammatory (tumor necrosis factor-α and monocyte chemotactic protein-1) and fibrogenic (collagen 1a1 and α smooth muscle actin) marker levels, and liver myeloperoxidase activity, which were significantly attenuated by PHGG treatment. Furthermore, the Ath diet combined with increased intestinal permeability resulted in elevated portal endotoxin levels and activated toll-like receptor (TLR) 4 and TLR9 expression, confirming that intestinal permeability was significantly elevated, as observed by evaluating the lumen-to-blood clearance of fluorescein isothiocyanate-conjugated dextran. PHGG treatment did not affect fatty acid metabolism in the liver. However, it decreased lipopolysaccharide signaling through the gut-liver axis. In addition, it significantly increased the abundance of cecal Bacteroides and Clostridium subcluster XIVa. Treatment with PHGG markedly increased the levels of SCFAs, particularly, butyric acid, acetic acid, propionic acid, and formic acid, in the cecal samples.CONCLUSIONPHGG partially prevented NAFLD development in mice through the gut-liver axis by modulating microbiota and downstream SCFA profiles.

Emotional memory impairment produced by binge drinking in mice is counteracted by the anti-inflammatory indomethacin

The Binge Drinking (BD) pattern of alcohol consumption, prevalent in adolescents and young adults, has been associated with memory impairment. In addition, evidence shows that alcohol abuse causes neuroinflammation, which may contribute to the brain damage produced by alcohol and explain its cognitive consequences. In this study, we evaluated the effectiveness of the anti-inflammatory indomethacin in counteracting the memory impairment produced by alcohol (ethanol) in adolescent mice of both sexes. Animals were randomly assigned to one of four groups for each sex: SS (saline + saline), SA (saline + alcohol), SI (saline + indomethacin) and AI (alcohol + indomethacin). They were injected acutely (Experiment 1) or chronically intermittent (Experiment 2) with saline, ethanol (3 g/kg) and indomethacin (10 mg/kg). All subjects were evaluated in an inhibitory avoidance task 96 h after treatment. With acute administration, SA groups showed significantly lower Test latencies than SS groups, while AI groups had similar latencies to controls. The chronic-intermittent administration of alcohol, an animal model of BD, produced significant emotional memory impairment -blocking learning in males- which was counteracted by indomethacin, as the AI groups had similar latencies to the SS groups. No significant differences were observed in locomotor activity or analgesia. In conclusion, alcohol BD (one or several episodes) impairs emotional memory in mice. This impairment is not secondary to the effects of alcohol BD on locomotor activity or pain sensitivity, and it is counteracted by indomethacin. Therefore, the memory impairment produced by alcohol BD seems to be mediated, in part, by neuroinflammatory processes. These findings open a window for new treatments for alcohol use disorders.

The development of a mouse model of mTBI-induced post-traumatic migraine, and identification of the delta opioid receptor as a novel therapeutic target.

Post-traumatic headache is the most common and long-lasting impairment observed following mild traumatic brain injury, and frequently has migraine-like characteristics. The mechanisms underlying progression from mild traumatic brain injury to post-traumatic headache are not fully understood. The aim of this study was to develop a mouse model of post-traumatic headache and identify mechanisms and novel targets associated with this disorder. We combined the closed head weight-drop method and the nitroglycerin chronic migraine model. To induce mild traumatic brain injury, a weight was dropped onto intact crania of mildly anesthetized mice, and mechanical responses to chronic-intermittent administration of nitroglycerin, a human migraine trigger, were determined at multiple time points post-injury. Low dose nitroglycerin (0.1 mg/kg) evoked acute periorbital and hind paw allodynia in both mild traumatic brain injury and sham animals. However, only mild traumatic brain injury mice developed chronic hypersensitivity to low dose nitroglycerin. Migraine medications, sumatriptan and topiramate, inhibited post-traumatic headache-associated allodynia. In addition, the delta opioid receptor agonist, SNC80, also blocked post-traumatic headache-associated allodynia. Finally, we examined the expression of calcitonin gene-related peptide within this model and found that it was increased in trigeminal ganglia two weeks post-mild traumatic brain injury. Overall, we have established a mouse model of post-traumatic headache and identified the delta opioid receptor as a novel therapeutic target for this disorder.

Increased Asics Expression via the Camkii-CREB Pathway in a Novel Mouse Model of Trigeminal Pain

Background/Aims: Migraine is a disabling condition that severely impacts socioeconomic function and quality of life. The focus of this study was to develop a mouse model of trigeminal pain that mimics migraine. Methods: After undergoing dural cannulation surgery, mice were treated with repeated dural doses of an acidic solution to induce trigeminal pain. Results: The method elicited intermittent, head-directed wiping and scratching as well as the expression of both the c-FOS gene in the spinal trigeminal nucleus caudalis and calcitonin gene related peptide (CGRP) in the periaqueductal grey matter. Interestingly, the acid-induced trigeminal pain behaviour was inhibited by amiloride, an antagonist of acid-sensing ion channels (ASICs), but not by AMG-9810, an inhibitor of transient receptor potential cation channel V1(TRPV1). In addition, the relative mRNA and protein expression levels of ASIC1a and ASIC3 were increased in the acid-induced trigeminal nociceptive pathways. Furthermore, blocking CaMKII with KN-93 significantly reduced the acid-induced trigeminal pain behaviour and c-FOS gene expression. Conclusion: The data suggested that chronic intermittent administration of an acidic solution to mice resulted in trigeminal hypersensitivity and that dural acid-induced trigeminal pain behaviour in mice may mechanistically mimic migraine. The observations here identify an entirely novel treatment strategy for migraine.

Characterization of a novel model of chronic migraine

Chronic migraine is a disabling condition that affects hundreds of millions of individuals worldwide. The development of novel migraine treatments has been slow, in part as a result of a lack of predicative animal models. We have developed a new model of chronic migraine involving the use of nitroglycerin (NTG), a known migraine trigger in humans. Chronic intermittent administration of NTG to mice resulted in acute mechanical hyperalgesia with each exposure as well as a progressive and sustained basal hyperalgesia. This chronic basal hyperalgesia occurred in a dose-dependent fashion and persisted for days after cessation of NTG administration. NTG-evoked hyperalgesia was exacerbated by the phosphodiesterase 5 inhibitor sildenafil, also a human migraine trigger, consistent with nitric oxide as a primary mediator of this hyperalgesia. The acute but not the chronic basal hyperalgesia was significantly reduced by the acute migraine therapy sumatriptan, whereas both the acute and chronic hyperalgesia was significantly attenuated by the migraine preventive therapy topiramate. Chronic NTG-induced hyperalgesia is a mouse model that may be useful for the study of mechanisms underlying progression of migraine from an episodic to a chronic disorder, and for the identification and characterization of novel acute and preventive migraine therapies.

The effect of chronic kisspeptin administration on seminal fructose levels in male mice

The discovery that kisspeptin was critical for normal fertility in all mammalian species including humans, ushered in a new chapter in our understanding of the control of GnRH secretion. Kisspeptin, the product of the KISS1 gene, plays an essential role in the regulation of spermatogenesis acting primarily at the hypothalamic level of the gonadotropic axis. Among the many identified substances in human semen, fructose is becoming increasingly significant. Fructose is synthesized and secreted by the seminal vesicles. Its synthesis is regulated by androgens and it is correlated directly with the levels of testosterone. Dose dependent degeneration of seminal vesicle has been described following intraperitoneal kisspeptin treatment; however, effects of kisspeptin administration on the levels of seminal fructose remain elusive till date. The present study, therefore, addresses the effects of 12-day administration of kisspeptin on seminal fructose levels in male mice. Kisspeptin-10 was administered intraperitoneally at different dosage concentrations (1 μg, 1 ng, and 10 ρg) to adult male mice, twice daily for 12 days. Seminal fructose levels were studied photometrically after 12 days of treatment. At the end of the treatment, seminal fructose levels decreased significantly after all tested doses. Chronic intermittent kisspeptin-10 administration negatively regulates seminal fructose levels in adult male mice.

Suppression of Hypothalamic–Pituitary–Adrenal Axis by Acute Heroin Challenge in Rats During Acute and Chronic Withdrawal from Chronic Heroin Administration

It is known that heroin dependence and withdrawal are associated with changes in the hypothalamic-pituitary-adrenal (HPA) axis. The objective of these studies in rats was to systematically investigate the level of HPA activity and response to a heroin challenge at two time points during heroin withdrawal, and to characterize the expression of associated stress-related genes 30min after each heroin challenge. Rats received chronic (10-day) intermittent escalating-dose heroin administration (3×2.5mg/kg/day on day 1; 3×20mg/kg/day by day 10). Hormonal and neurochemical assessments were performed in acute (12h after last heroin injection) and chronic (10days after the last injection) withdrawal. Both plasma ACTH and corticosterone levels were elevated during acute withdrawal, and heroin challenge at 20mg/kg (the last dose of chronic escalation) at this time point attenuated this HPA hyperactivity. During chronic withdrawal, HPA hormonal levels returned to baseline, but heroin challenge at 5mg/kg decreased ACTH levels. In contrast, this dose of heroin challenge stimulated the HPA axis in heroin naïve rats. In the anterior pituitary, pro-opiomelanocortin (POMC) mRNA levels were increased during acute withdrawal and retuned to control levels after chronic withdrawal. In the medial hypothalamus, however, the POMC mRNA levels were decreased during acute withdrawal, and increased after chronic withdrawal. Our results suggest a long-lasting change in HPA abnormal responsivity during chronic heroin withdrawal.

Behavioural and neurochemical comparison of chronic intermittent cathinone, mephedrone and MDMA administration to the rat

The synthetic cathinone derivative, mephedrone, is a controlled substance across Europe. Its effects have been compared by users to 3,4-methylenedioxymethamphetamine (MDMA), but little data exist on its pharmacological properties. This study compared the behavioural and neurochemical effects of mephedrone with cathinone and MDMA in rats. Young-adult male Lister hooded rats received i.p. cathinone (1 or 4mg/kg), mephedrone (1, 4 or 10mg/kg) or MDMA (10mg/kg) on two consecutive days weekly for 3 weeks or as a single acute injection (for neurochemical analysis). Locomotor activity (LMA), novel object discrimination (NOD), conditioned emotional response (CER) and prepulse inhibition of the acoustic startle response (PPI) were measured following intermittent drug administration. Dopamine, 5-hydroxytryptamine (5-HT) and their major metabolites were measured in striatum, frontal cortex and hippocampus by high performance liquid chromatography 7 days after intermittent dosing and 2h after acute injection. Cathinone (1, 4mg/kg), mephedrone (10mg/kg) and MDMA (10mg/kg) induced hyperactivity following the first and sixth injections and sensitization to cathinone and mephedrone occurred with chronic dosing. All drugs impaired NOD and mephedrone (10mg/kg) reduced freezing in response to contextual re-exposure during the CER retention trial. Acute MDMA reduced hippocampal 5-HT and 5-HIAA but the only significant effect on dopamine, 5-HT and their metabolites following chronic dosing was altered hippocampal 3,4-dihydroxyphenylacetic acid (DOPAC), following mephedrone (4, 10mg/kg) and MDMA. At the doses examined, mephedrone, cathinone, and MDMA induced similar effects on behaviour and failed to induce neurotoxic damage when administered intermittently over 3 weeks.

Chronic cannabinoid exposure reduces phencyclidine-induced schizophrenia-like positive symptoms in adult rats

Chronic cannabis use can induce psychotic states that resemble schizophrenia. Yet, schizophrenic patients often smoke cannabis as a form of self-medication to counter the aversive symptoms of schizophrenia. We recently demonstrated an ameliorating effect of cannabinoid self-administration (SA) on negative and cognitive schizophrenia-like symptoms induced experimentally by the non-competitive N-methyl-D-aspartate receptor antagonist phencyclidine (PCP). Whether cannabinoid SA alleviates or exacerbates schizophrenia-like positive symptoms is still unclear. This follow-up study aimed to evaluate the effect of self-administered cannabinoid on PCP-induced schizotypic positive symptoms in adult rats. Male rats were trained to self-administer either the cannabinoid CB1 receptor agonist WIN 55,212-2 (WIN; 12.5μg/kg/infusion) or its vehicle (Veh) intravenously. The effects of acute and chronic intermittent intraperitoneal administration of PCP (2.5mg/kg) on motor parameters were then tested in Veh-SA and WIN-SA. Cannabinoid SA significantly attenuated the psychotomimetic effects of PCP exposure observed in control rats. Following acute PCP administration, WIN-SA animals displayed more frequent rearing and lower anxiety-like profile than Veh-SA rats. WIN-SA rats also exhibited lower behavioural sensitisation to chronic PCP treatment as demonstrated by reduced hyperlocomotion in response to an acute PCP challenge. In addition, parallel experiments performed in experimenter-administered rats that received WIN at comparable SA doses confirmed the ameliorating effects of cannabinoid exposure on PCP-induced schizotypic behaviours, indicating that motivational effects were not responsible for the ameliorative effects of cannabinoids. Our results indicate that cannabis may exert protective effects on positive schizotypic symptoms in adult animals such as hypermotility and anxiety state.

Cannabinoid‐induced delayed gastric emptying is selectively increased upon intermittent administration in the rat: role of CB1 receptors

Cannabinoids acutely administered depress central, cardiovascular and gastrointestinal functions. These effects might be modified upon repeated administration. Compared to the effects induced by daily administration, those induced by intermittent administration are less known. The effect of intermittent treatment with the CB1/CB2 cannabinoid agonist WIN55,212-2 (WIN) was studied in the rat. Male rats received saline, vehicle or WIN at 0.5 (low-WIN) or 5 (high-WIN) mg kg(-1) week(-1) for 4 weeks. WIN effects on the central nervous system (cannabinoid tetrad tests), cardiovascular function and gastrointestinal motor function were evaluated after the first and last doses, and, where appropriate, 1 week after the last dose. To determine the involvement of CB1 receptors in the chronic effect of WIN, the CB1 receptor antagonist/inverse agonist AM251 (1 mg kg(-1)) was used. High- (but not low-) WIN induced the four signs of the cannabinoid tetrad, and reduced gastrointestinal motility, but did not alter cardiovascular parameters. Upon chronic intermittent administration, tolerance did not clearly develop to WIN effects. Quite the opposite, depression of gastric emptying was intensified. No effect was long-lasting. Repeated administration of AM251 was more efficacious than single administration to block WIN chronic central effects, but the opposite occurred regarding lower intestinal motility. Upon intermittent administration, hypersensitization may develop to some effects (particularly delayed gastric emptying) induced by cannabinoid agonists. CB1 antagonists/inverse agonists may show different efficacy upon repeated or single administration to block cannabinoid-induced central and gastrointestinal effects. Thus, cannabinoid effects are dependent on the pattern of drug administration.

Fluctuating and constant valproate administration gives equivalent seizure control in rats with genetic and acquired epilepsy

Purpose Controlled-release formulations of Valproate (VPA) reduce side effects by minimizing peak plasma VPA concentrations in patients with epilepsy. However, the impact of this on anti-seizure efficacy has not been thoroughly explored. Here the pharmacokinetics and pharmacodynamics of chronic intermittent (consequently, peak VPA concentrations) and continuous VPA administration were directly compared in two rat models of epilepsy. Methods Genetic Absence Epilepsy Rats from Strasbourg (GAERS) received a single acute bolus of VPA (100 mg/kg intravenously) combined with electroencephalography (EEG) and/or blood sampling for 180 min post-injection. GAERS and epileptic rats post-kainic acid-induced status epilepticus were chronically infused intravenously (3–5 days, respectively) with (i) saline followed by in random order (ii) intermittent and (iii) continuous VPA (42 mg/kg/h), separated by two days of wash-out. Seizures were quantified using video-EEG monitoring and VPA levels measured in brain, cerebrospinal fluid and plasma. Results Following acute VPA administration seizure suppression in GAERS persisted after plasma VPA levels became very low. Chronic intermittent and continuous VPA significantly suppressed seizures in both models ( p < 0.01) with no difference between administration regimens. In GAERS, the pattern of seizure suppression during intermittent treatment was constant, in contrast to the fluctuating VPA plasma and brain levels. There was discordance between the temporal pattern of plasma, brain VPA levels and seizure suppression efficacy in GAERS. Conclusion Administration regimes that result in fluctuating VPA blood levels achieve equivalent sustained seizure suppression as those that maintain steady mid-range concentrations.

Subchronic and chronic PCP treatment produces temporally distinct deficits in attentional set shifting and prepulse inhibition in rats

We have previously demonstrated that subchronic (five daily administrations of 2.6 mg/kg PCP) and chronic intermittent administration of 2.6 mg/kg PCP to rats produces hypofrontality and other neurochemical changes akin to schizophrenia pathology (Cochran et al., Neuropsychopharmacology, 28:265-275, 2003). We sought to determine whether behavioral alterations related to discrete aspects of schizophrenia are also induced by these PCP treatment regimes. Following administration of vehicle or PCP according to the protocols described above, rats were assessed for attentional set shifting ability, prepulse inhibition (PPI), or social interaction and the locomotor response to a challenge dose of amphetamine. Ability to shift attentional set was impaired 72 h after the last PCP administration following the subchronic and chronic intermittent treatment regimes. PPI was disrupted after each acute administration of PCP in animals under the subchronic treatment regime. However, PPI deficits were not sustained 72 h after the last of five daily administrations. In subchronic and chronic PCP treated animals, no change was found in social interaction behavior, and there was little change in baseline or amphetamine-stimulated locomotor activity, employed as an indicator of dopaminergic hyperfunction. The temporally distinct behavioral effects of these PCP treatment regimes suggest that PPI deficits relate directly to acute NMDA receptor antagonism, whereas the more enduring set shifting deficits relate to the longer term consequences of NMDA receptor blockade. Therefore, these subchronic and chronic PCP treatment regimes produce hypofrontality (Cochran et al., Neuropsychopharmacology, 28:265-275, 2003) and associated prefrontal cortex-dependent deficits in behavioral flexibility which mirror core deficits in schizophrenia.

Deletion of the 5‐HT3A‐receptor subunit blunts the induction of cocaine sensitization

Serotonin (5-HT) receptors are classified into seven groups (5-HT(1-7)), comprising at least 14 structurally and pharmacologically distinct receptor subtypes. Pharmacological antagonism of ionotropic 5-HT(3) receptors has been shown to modulate both behavioral and neurochemical aspects of the induction of sensitization to cocaine. It is not known, however, if specific molecular subunits of the 5-HT(3) receptor influence the development of cocaine sensitization. To address this question, we studied the effects of acute and chronic intermittent cocaine administration in mice with a targeted deletion of the gene for the 5-HT(3A)-receptor subunit (5-HT(3A)-/-). 5-HT(3A) (-/-) mice showed blunted induction of cocaine-induced locomotor sensitization as compared with wild-type littermate controls. 5-HT(3A) (-/-) mice did not differ from wild-type littermate controls on measures of basal motor activity or response to acute cocaine treatment. Enhanced locomotor response to saline injection following cocaine sensitization was observed equally in 5-HT(3A) (-/-) and wild-type mice suggesting similar conditioned effects associated with chronic cocaine treatment. These data show a role for the 5-HT(3A)-receptor subunit in the induction of behavioral sensitization to cocaine and suggest that the 5-HT(3A) molecular subunit modulates neurobehavioral adaptations to cocaine, which may underlie aspects of addiction.

Tyrosine Kinase Phosphorylation of GABAA Receptor α1, β2 and γ2 Subunits Following Chronic Intermittent Ethanol (CIE) Exposure of Cultured Cortical Neurons of Mice

There is evidence that many of the GABA(A) receptor subunits contain consensus sequence for tyrosine kinase, and phosphorylation may play a key role in ethanol's regulation of GABA(A) receptors. Recently, we investigated the effect of chronic exposure of ethanol (CE) on tyrosine kinase phosphorylation and reported that there was an up-regulation in tyrosine kinase phosphorylation of the beta(2)- and gamma(2)- subunits and no effect on alpha(1)-subunit of the GABA(A) receptor in the cultured cortical neurons of mice. In the present study, we have further investigated the effect of chronic intermittent administration of ethanol (CIE) on tyrosine kinase phosphorylation of the GABA(A) receptor subunits (alpha(1), beta(2), and gamma(2)) in the mouse cultured cortical neurons by immunoprecipitation and Western blot techniques. We observed that there was an up-regulation in the tyrosine kinase phosphorylation of the GABA(A )receptor beta(2)- and gamma(2)-subunits following CIE exposure, and no effect on alpha(1)-subunit in the cultured cortical neurons of mice. These CIE changes, unlike CE, were not reverted back to the control level following ethanol withdrawal even after 7 days. Acute exposure of ethanol did not cause any change in the tyrosine kinase regulation of the GABA(A) receptor subunits. In conclusion, the CIE exposure, unlike chronic/acute ethanol exposure, regulates the tyrosine kinase phosphorylation of the selective population of GABA(A )receptors in a long lasting manner.

Chronic Intermittent Administration of HIF Prolyl Hydroxylase Inhibitor Significantly Induces Erythropoietin and Increases Total Hemoglobin in Normal Rhesus Macaques.

Introduction: Injectable recombinant erythropoietin (rHuEPO) is used in many clinical disorders of anemia. Prior attempts at EPO induction by gene transfer strategies were problematic rendering them less attractive. Hypoxia inducible factor (HIF) is important in mediating EPO expression and erythropoiesis. Under normoxic conditions, HIF-alpha protein is usually undetectable due to rapid inactivation by oxygen-dependent HIF prolyl hydroxylases (HIF-PH). In hypoxic conditions or with HIF-PH inhibitors, HIF-alpha protein levels are increased and maintained due to reduced activity of HIF-PH. Upon stabilization HIF-alpha is transported to the nucleus to increase the expression of EPO, aminolevulinic acid synthase (ALAS), iron-transport proteins, nitric oxide synthase, and others. FG-2216 is an orally active HIF-PH inhibitor that induces EPO expression and erythropoiesis in vitro and in vivo, and improves iron bioavaliability by increasing iron mobilization from intestinal and macrophage stores and by decreasing hepcidin expression. Here we report on the pharmacokinetics and pharmacodynamics of FG-2216 to induce EPO expression and erythropoiesis in non-human primates with chronic intermittent oral dosing.Methods: FG-2216 was obtained from FibroGen, Inc. FG-2216 was given to rhesus macaques (4-7 yrs old) by oral gavage. Drug and EPO levels were measured at 0,2,4,8,10,24,48, and 72 hours following a single dose of 40 or 60mg/kg. 40 or 60mg/kg of drug was then given twice a week to 5 male animals for 3–6 months. After 6–8 weeks of twice weekly dosing at 60mg/kg, weekly 15–20% weight per volume phlebotomy was introduced for 6–8 weeks to simulate chronic anemia while drug dosing was continued. Weekly weights, complete blood counts, and metabolic profiles were obtained to assess efficacy and safety. Two male animals received vehicle control.Results: FG-2216 is rapidly absorbed and has excellent oral bioavailability, with peak plasma drug concentration between 1–2 hours and a half-life of 7–8 hours. The pharmacokinetic curves were dose-proportional and predictable. After 6–8 weeks of 2 doses per week, all animals tolerated the drug without significant side effects, and animal weights were stable with no changes in serum electrolytes, hepatic, or renal parameters. Peak plasma EPO increased from a baseline of <20 to 132 and 1950 mIU/ml after 40 and 60 mg/kg doses, respectively. There were transient 2-3 fold increases in reticulocytes. All treated animals demonstrated an increase in total hemoglobin: 11.95 +/− 0.33 g/dL (baseline), 13.17 +/− 0.32 (40mg/kg), and 14.58 +/− 0.64 (60mg/kg). Absolute hemoglobin increases ranged from 0.66 to 1.8 (40mg/kg) and 0.97 to 4.04 (60mg/kg) g/dL. When phlebotomy was introduced with continued FG-2216 dosing, treated animals maintained their hemoglobin, 14.03 +/− 0.06 g/dL, while control animals had their hemoglobin reduced to 11.54 +/− 0.09g/dL.Conclusion: FG-2216 is an orally bioavailable HIF-PH inhibitor that is well tolerated and a potent inducer of EPO, with greater than 50-fold increases in EPO above baseline with a 60mg/kg dose. When administered orally twice a week, FG-2216 significantly increases total hemoglobin in normal rhesus macaques. Under the stress of weekly phlebotomy, FG-2216 maintained the hemoglobin of treated animals in the normal range, averaging 2.5 g/dL higher than animals receiving placebo. These results have broad clinical implications.

Vesicular monoamine transporter-2 and aromatic l-amino acid decarboxylase gene therapy prevents development of motor complications in parkinsonian rats after chronic intermittent l-3,4-dihydroxyphenylalanine administration

Motor complications after chronic l-3,4-dihydroxyphenylalanine ( l-DOPA) therapy occur partly because of the sensitization to dopaminergic agents resulting from pulsatile dopaminergic stimulation. The loss of presynaptic storage contributes to short duration of action by dopamine. Vesicular monoamine transporter-2 (VMAT-2) controls intraneuronal dopamine storage by packaging dopamine into synaptic vesicles, thereby allowing exocytotic release of dopamine. Using primary fibroblast doubly transduced with VMAT-2 and aromatic l-amino acid decarboxylase (AADC) genes, we previously demonstrated the beneficial effects of such double gene transduction in the production, storage, and gradual release of dopamine in vitro and in vivo. In this study, we further evaluate the effect of achieving sustained level of dopamine within the striata by VMAT-2 gene on behavioral response of parkinsonian rats after chronic intermittent l-DOPA administration. Primary fibroblast (PF) cells were genetically modified with AADC and VMAT-2 genes. We grafted primary fibroblast cells, PF with AADC (PFAADC), or doubly transduced PF with AADC and VMAT-2 (PFVMAA) ( n = 6 for each group) into parkinsonian rat striata and administered l-DOPA (25 mg/kg/day) intermittently for 4 weeks. For behavioral study, we employed a model of akinesia using forepaw adjusting steps (FAS) that have been well characterized to reflect the effect of the lesion and the antiparkinsonian effect of dopaminergic drugs and transplants. The duration of FAS response to l-DOPA was sustained for a longer duration in rats grafted with PFVMAA cells than in those grafted with either control cells or cells with AADC alone. In PFVMAA-grafted animals, prolonged duration of FAS responses to l-DOPA was sustained even 6 weeks after discontinuation of 4-week intermittent l-DOPA treatment. These findings suggest that the restoration of dopamine storage capacity could enhance the efficacy of l-DOPA therapy and attenuate the motor fluctuations that result from chronic intermittent l-DOPA administration. The gene therapy expressing AADC and VMAT-2 along with systemic l-DOPA therapy could provide a novel treatment strategy to prevent motor fluctuations.