Increase In Extracellular Concentration Research Articles

GLT-1-Dependent Disruption of CNS Glutamate Homeostasis and Neuronal Function by the Protozoan Parasite Toxoplasma gondii.

The immune privileged nature of the CNS can make it vulnerable to chronic and latent infections. Little is known about the effects of lifelong brain infections, and thus inflammation, on the neurological health of the host. Toxoplasma gondii is a parasite that can infect any mammalian nucleated cell with average worldwide seroprevalence rates of 30%. Infection by Toxoplasma is characterized by the lifelong presence of parasitic cysts within neurons in the brain, requiring a competent immune system to prevent parasite reactivation and encephalitis. In the immunocompetent individual, Toxoplasma infection is largely asymptomatic, however many recent studies suggest a strong correlation with certain neurodegenerative and psychiatric disorders. Here, we demonstrate a significant reduction in the primary astrocytic glutamate transporter, GLT-1, following infection with Toxoplasma. Using microdialysis of the murine frontal cortex over the course of infection, a significant increase in extracellular concentrations of glutamate is observed. Consistent with glutamate dysregulation, analysis of neurons reveal changes in morphology including a reduction in dendritic spines, VGlut1 and NeuN immunoreactivity. Furthermore, behavioral testing and EEG recordings point to significant changes in neuronal output. Finally, these changes in neuronal connectivity are dependent on infection-induced downregulation of GLT-1 as treatment with the ß-lactam antibiotic ceftriaxone, rescues extracellular glutamate concentrations, neuronal pathology and function. Altogether, these data demonstrate that following an infection with T. gondii, the delicate regulation of glutamate by astrocytes is disrupted and accounts for a range of deficits observed in chronic infection.

Propriétés pharmacologiques de la vortioxetine et leurs conséquences pré-cliniques

Selective Serotonin Reuptake Inhibitors (SSRIs) are extensively used for the treatment of major depressive disorder (MDD). SSRIs are defined as indirect receptor agonists since the activation of postsynaptic receptors is a consequence of an increase in extracellular concentrations of serotonin (5-HT) mediated by the blockade of serotonin transporter. The activation of some serotoninergic receptors (5-HT1A, post-synaptic, 5-HT1B post-synaptic, 5-HT2B, and 5-HT4), but not all (5-HT1A, pre-synaptic, 5-HT1B pre-synaptic, 5-HT2A, 5-HT2C, 5-HT3, and probably 5-HT6), induces anxiolytic/antidepressive - like effects. Targetting specifically some of them could potentially improve the onset of action and/or efficacy and/or prevent MD relapse. Vortioxetine (Brintellix, 1- [2-(2,4-dimethylphenyl-sulfanyl)-phenyl]-piperazine) is a novel multi-target antidepressant drug approved by the Food and Drug Administration (FDA) and by European Medicines Agency. Its properties are markedly different from the extensively prescribed SSRIs. Compared to the SSRIs, vortioxetine is defined as a multimodal antidepressant drug since it is not only a serotonin reuptake inhibitor, but also a 5-HT1D, 5-HT3, 5-HT7 receptor antagonist, 5-HT1B receptor partial agonist and 5-HT1A receptor agonist. This specific pharmacological profile enables vortioxetine to affect not only the serotoninergic and noradrenergic systems, but also the histaminergic, cholinergic, gamma-butyric acid (GABA) ergic and glutamatergic ones. Thus, vortioxetine not only induces antidepressant-like or anxiolytic-like activity but also improves cognitive parameters in several animal models. Indeed, vortioxetine was shown to improve working memory, episodic memory, cognitive flexibility and spatial memory in young adult rodents and also in old animal models. These specific effects of the vortioxetine are of interest considering that cognitive dysfunction is a common comorbidity to MDD. Altogether, even though this molecule still needs to be investigated further, especially in the insufficient-response to antidepressant drugs, vortioxetine is already an innovative therapeutic option for the treatment of major depression.

Staphylococcal α-hemolysin is neurotoxic and causes lysis of brain cells in vivo and in vitro

Formation of a bacterial brain abscess entails loss of brain cells and formation of pus. The mechanisms behind the cell loss are not fully understood. Staphylococcus aureus, a common cause of brain abscesses, produces various exotoxins, including α-hemolysin, which is an important factor in brain abscess formation. α-Hemolysin may cause cytolysis by forming pores in the plasma membrane of various eukaryotic cells. However, whether α-hemolysin causes lysis of brain cells is not known. Nor is it known whether α-hemolysin in the brain causes cell death through pore formation or by acting as a chemoattractant, recruiting leukocytes and causing inflammation. Here we show that α-hemolysin injected into rat brain causes cell damage and edema formation within 30min. Cell damage was accompanied by an increase in extracellular concentrations of zinc, GABA, glutamate, and other amino acids, indicating plasma membrane damage, but leukocytic infiltration was not seen 0.5–12h after α-hemolysin injection. This was in contrast to injection of S. aureus, which triggered extensive infiltration with neutrophils within 8h. In vitro, α-hemolysin caused concentration-dependent lysis of isolated nerve endings and cultured astrocytes. We conclude that α-hemolysin contributes to the cell death inherent in staphylococcal brain abscess formation as a pore-forming neurotoxin.

Characterization of saxitoxin production and release and phylogeny of sxt genes in paralytic shellfish poisoning toxin-producing Aphanizomenon gracile

The freshwater cyanobacterium Aphanizomenon gracile is one of the most widely distributed producers of the potent neurotoxins saxitoxin (STX) and its derivatives (paralytic shellfish poisoning toxins, PSP toxins). However, the phylogeny of STX biosynthesis genes and the regulation of STX production and release remain poorly studied in the genus Aphanizomenon. In this study, two A. gracile strains from Spanish freshwaters were grown in semi-continuous cultures under three temperatures (15, 20 and 28°C) and their STX production and release were determined by Enzyme-Linked ImmunoSorbent Assay (ELISA). STX production was stable along the temperature range, with 1.4–2.3-fold shifts in biomass-standardized STX contents, and maxima of 0.22μg equivalent STXmg−1 dry weight 15.3fg equiv STX cell−1 and 15.1μg equiv STXmg−1 Chl a. The extracellular fraction was remarkably high (13.6–35.3%), not clearly affected by temperature but with nitrate-depleted medium (BG110) inducing a 2-fold increase in extracellular content. STX production and release were not directly related to growth rates. The 16S rRNA phylogenetic analyses in sixteen A. gracile strains from Spanish and German freshwaters showed that PSP-producing A. gracile grouped within a monospecific and highly supported cluster, together with PSP-producing Aphanizomenon sp. NH-5 and clearly separated from a monospecific Aphanizomenon flos-aquae cluster. The sixteen A. gracile strains formed also monospecific and highly supported clusters for PSP-biosynthesis genes (sxtG, sxtI, sxtH and sxtX) together with Aphanizomenon sp. NH-5. This study evidences an elevated extracellular proportion of STX in A. gracile with importance for risk assessment, and supports the identification of Aphanizomenon sp. NH-5 as A. gracile.

Overdose of Drugs for Attention-Deficit Hyperactivity Disorder: Clinical Presentation, Mechanisms of Toxicity, and Management

The prevalence of attention-deficit hyperactivity disorder (ADHD) in the USA is estimated at approximately 4-9% in children and 4% in adults. It is estimated that prescriptions for ADHD medications are written for more than 2.7 million children per year. In 2010, US poison centers reported 17,000 human exposures to ADHD medications, with 80% occurring in children <19 years old and 20% in adults. The drugs used for the treatment of ADHD are diverse but can be roughly separated into two groups: the stimulants such as amphetamine, methylphenidate, and modafinil; and the non-stimulants such as atomoxetine, guanfacine, and clonidine. This review focuses on mechanisms of toxicity after overdose with ADHD medications, clinical effects from overdose, and management. Amphetamine, dextroamphetamine, and methylphenidate act as substrates for the cellular monoamine transporter, especially the dopamine transporter (DAT) and less so the norepinephrine (NET) and serotonin transporter. The mechanism of toxicity is primarily related to excessive extracellular dopamine, norepinephrine, and serotonin. The primary clinical syndrome involves prominent neurological and cardiovascular effects, but secondary complications can involve renal, muscle, pulmonary, and gastrointestinal (GI) effects. In overdose, the patient may present with mydriasis, tremor, agitation, hyperreflexia, combative behavior, confusion, hallucinations, delirium, anxiety, paranoia, movement disorders, and seizures. The management of amphetamine, dextroamphetamine, and methylphenidate overdose is largely supportive, with a focus on interruption of the sympathomimetic syndrome with judicious use of benzodiazepines. In cases where agitation, delirium, and movement disorders are unresponsive to benzodiazepines, second-line therapies include antipsychotics such as ziprasidone or haloperidol, central alpha-adrenoreceptor agonists such as dexmedetomidine, or propofol. Modafinil is not US FDA approved for treatment of ADHD; however, it has been shown to improve ADHD signs and symptoms and has been used as an off-label pharmaceutical for this diagnosis in both adults and children. The mechanism of action of modafinil is complex and not fully understood. It is known to cause an increase in extracellular concentrations of dopamine, norepinephrine, and serotonin in the neocortex. Overdose with modafinil is generally of moderate severity, with reported ingestions of doses up to 8 g. The most common neurological effects include increased anxiety, agitation, headache, dizziness, insomnia, tremors, and dystonia. The management of modafinil overdose is largely supportive, with a focus on sedation, and control of dyskinesias and blood pressure. Atomoxetine is a selective presynaptic norepinephrine transporter inhibitor. The clinical presentation after overdose with atomoxetine has generally been mild. The primary effects have been drowsiness, agitation, hyperactivity, GI upset, tremor, hyperreflexia, tachycardia hypertension, and seizure. The management of atomoxetine overdose is largely supportive, with a focus on sedation, and control of dyskinesias and seizures. Clonidine is a synthetic imidazole derivative with both central and peripheral alpha-adrenergic agonist actions. The primary clinical syndrome involves prominent neurological and cardiovascular effects, with the most commonly reported features of depressed sensorium, bradycardia, and hypotension. While clonidine is an anti-hypertensive medication, a paradoxical hypertension may occur early with overdose. The clinical syndrome after overdose of guanfacine may be mixed depending on central or peripheral alpha-adrenoreceptor effects. Initial clinical effects may be drowsiness, lethargy, dry mouth, and diaphoresis. Cardiovascular effects may depend on time post-ingestion and may present as hypotension or hypertension. The management of guanfacine overdose is largely supportive, with a focus on support of blood pressure. Overdose with ADHD medications can produce major morbidity, with many cases requiring intensive care medicine and prolonged hospital stays. However, fatalities are rare with appropriate care.

Caffeine and Substance Use Disorders

Caffeine and Substance Use Disorders

Effect of paroxetine and bupropion on human resting brain perfusion: An arterial spin labeling study

Due to the physiological coupling with metabolism, brain perfusion is a potential aid in understanding the effects of medication on neural activity in vivo. In this study, arterial spin labeling (ASL) was used to quantify the effect of seven days of administration of paroxetine (20 mg), bupropion (150 mg) on cerebral blood flow levels at rest in a double-blind, placebo-controlled crossover design in N=21 healthy participants. Paroxetine administration was associated with diffuse cortical perfusion decrements, more marked in the prefrontal region, and with decrements in the striatum, caudate and the basal forebrain. The effect of bupropion on the cortex was similar but less conspicuous. In the brainstem, there was suggestive evidence of perfusion decrements in the main serotonergic and noradrenergic nuclei. These results demonstrate the existence of effects on rest perfusion of antidepressant medication of potential use in monitoring its action. The observed perfusion reductions may be related to compensatory decrements of monoaminergic neural activity in the initial phases of treatment that are observed under both drugs following the increase in extracellular concentration of transmitters.

Effects of a novel mGlu2/3 receptor agonist prodrug, LY2140023 monohydrate, on central monoamine turnover as determined in human and rat cerebrospinal fluid

Accumulating evidence suggests that the primary symptoms of schizophrenia may be associated with altered central glutamate transmission. LY2140023 monohydrate is the methionine prodrug of the selective mGlu(2/3) receptor agonist LY404039 and is currently being assessed for the treatment of schizophrenia. The objective of this study was to evaluate the central pharmacological activity of LY2140023 monohydrate in preclinical and clinical studies. Effects on neurotransmitter/metabolite concentrations were assessed in male rats following single oral doses of LY2140023 monohydrate (microdiasylates from the prefrontal cortex), single intraperitoneal injection of LY404039 [cerebrospinal fluid (CSF)], or LY2140023 monohydrate dosed once daily for 7 days (CSF). A clinical study in 16 healthy subjects assessed the effects of LY2140023 monohydrate 40 mg orally twice daily for 14 days in lumbar CSF. Rat studies: Acute dosing with LY2140023 monohydrate resulted in significant dose-dependent increases in extracellular concentrations of dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA), but not 5-hydroxyindoleacetic acid (5-HIAA), in the prefrontal cortex. LY2140023 monohydrate dosing for 7 days elevated the concentrations of HVA in CSF, while acutely dosed LY404039 increased the concentrations of DOPAC, HVA, and methoxy-hydroxyphenylglycol (MHPG), but not 5-HIAA. Clinical study: Significant increases were seen for DOPAC, HVA, 5-HIAA, and MHPG in the CSF of subjects receiving LY2140023 monohydrate, but not placebo. LY2140023 monohydrate and/or LY404039 dosing potently affected dopamine turnover and also significantly affected serotonin turnover in the human and rat central nervous systems. The measurement of biogenic amine metabolites such as DOPAC and HVA may serve as useful biomarkers of LY2140023 monohydrate and/or LY404039 central pharmacodynamic activity.

Perception of the Arabidopsis Danger Signal Peptide 1 Involves the Pattern Recognition Receptor AtPEPR1 and Its Close Homologue AtPEPR2

Plasma membrane-borne pattern recognition receptors, which recognize microbe-associated molecular patterns and endogenous damage-associated molecular patterns, provide the first line of defense in innate immunity. In plants, leucine-rich repeat receptor kinases fulfill this role, as exemplified by FLS2 and EFR, the receptors for the microbe-associated molecular patterns flagellin and elongation factor Tu. Here we examined the perception of the damage-associated molecular pattern peptide 1 (AtPep1), an endogenous peptide of Arabidopsis identified earlier and shown to be perceived by the leucine-rich repeat protein kinase PEPR1. Using seedling growth inhibition, elicitation of an oxidative burst and induction of ethylene biosynthesis, we show that wild type plants and the pepr1 and pepr2 mutants, affected in PEPR1 and in its homologue PEPR2, are sensitive to AtPep1, but that the double mutant pepr1/pepr2 is completely insensitive. As a central body of our study, we provide electrophysiological evidence that at the level of the plasma membrane, AtPep1 triggers a receptor-dependent transient depolarization through activation of plasma membrane anion channels, and that this effect is absent in the double mutant pepr1/pepr2. The double mutant also fails to respond to AtPep2 and AtPep3, two distant homologues of AtPep1 on the basis of homology screening, implying that the PEPR1 and PEPR2 are responsible for their perception too. Our findings provide a basic framework to study the biological role of AtPep1-related danger signals and their cognate receptors.

Gliopathy ensures persistent inflammation and chronic pain after spinal cord injury

Research focused on improving recovery of function, including the reduction of central neuropathic pain (CNP) after spinal cord injury (SCI) is essential. After SCI, regional neuropathic pain syndromes above, at and below the level or spinal injury develop and are thought to have different mechanisms, but may share common dysfunctional glial mechanisms. Detloff et al., [Detloff, M.R., Fisher, L.C., McGaughy, V., Longbrake, E.E., Popovich, P.G., Basso, D.M., Remote activation of microglia and pro-inflammatory cytokines predict the onset and severity of below-level neuropathic pain after spinal cord injury in rats. Exp. Neurol. (2008), doi: 10.1016/j.expneurol.2008.04.009.] describe events in the lumbar region of the spinal cord after a midthoracic SCI injury, the so called “below-level” pain and compares the findings to peripheral nerve lesion findings. This commentary briefly reviews glial contributions and intracellular signaling mechanisms, both neuronal and glial, that provide the substrate for CNP after SCI, including the persistent glial production of factors that can maintain sensitization of dorsal horn neurons in segments remote from the spinal injury; ie. dorsal horn hyperexcitability to formerly non noxious stimuli that become noxious after SCI resulting in allodynia. The term “gliopathy” is proposed to describe the dysfunctional and maladaptive response of glial cells, specifically astrocytes and microglia, to neural injury that is initiated by the sudden injury induced increase in extracellular concentrations of glutamate and concomitant production of several proinflammatory molecules. It is important to understand the roles that different glia play in “gliopathy”, a condition that appears to persist after SCI. Furthermore, targeted treatment of gliopathy will attenuate mechanical allodynia in both central and peripheral neuropathic pain syndromes.

The selective delta opioid agonist SNC80 enhances amphetamine-mediated efflux of dopamine from rat striatum

The highly selective delta opioid agonist, SNC80, elicits dopamine-related behaviors including locomotor stimulation and conditioned place-preference. In contrast, it has been reported that SNC80 fails to promote dopamine efflux from the striatum of freely moving rats. However, SNC80 does enhance behavioral responses to the stimulants, amphetamine and cocaine, suggesting an interaction between delta opioids and psychostimulants. Since the increase in locomotor activity elicited by amphetamine and related stimulants acting at the dopamine transporter is associated with increases in extracellular concentrations of dopamine within the striatum, we hypothesized that SNC80 enhances this activity by potentiating the overflow of dopamine through the transporter. To test this hypothesis, striatal preparations from Sprague Dawley rats were assayed for dopamine efflux in response to amphetamine challenge. SNC80 was given either in vivo or in vitro directly to rat striatal tissue, prior to in vitro amphetamine challenge. Both in vivo and in vitro administration of SNC80 enhanced amphetamine-mediated dopamine efflux in a concentration- and time-dependent manner. However, SNC80 in either treatment paradigm produced no stimulation of dopamine efflux in the absence of amphetamine. The effect of SNC80 on amphetamine-mediated dopamine overflow, but not the effect of amphetamine alone, was blocked by the delta selective antagonist, naltrindole and was also observed with other delta agonists. The results of this study demonstrate that even though SNC80 does not stimulate dopamine efflux alone, it is able to augment amphetamine-mediated dopamine efflux through a delta opioid receptor mediated action locally in the striatum.

Suppressive effect of Yokukansan on excessive release of glutamate and aspartate in the hippocampus of zinc-deficient rats

Yokukansan (TJ-54), a herbal medicine, has been used as a cure for insomnia and irritability in children. Yokukansan also improves behavioral and psychological symptoms such as agitation, aggression and irritability in patients with dementia including Alzheimer's disease, in which the glutamatergic neurotransmitter system is perturbed. However, the action of Yokukansan in synaptic neurotransmission is unknown. In the present study, the action of Yokukansan in the glutamatergic neurotransmitter system was examined in zinc-deficient rats, a neurological disease model, in which the glutamatergic neurotransmitter system is perturbed. Administration of Yokukansan significantly suppressed the increase in extracellular concentrations of glutamate and aspartate in the hippocampus after stimulation with 100 mM KCl, but not the increase in extracellular concentrations of glycine and taurine, suggesting that Yokukansan is involved in modulation of excitatory neurotransmitter systems. The present study demonstrates that Yokukansan is a possible medicine for prevention or cure of neurological diseases associated with excitotoxicity.

The neurobiology of cocaine-induced reinforcement.

Cocaine has potent pharmacological actions on a number of monoaminergic systems in the brain, including those that use noradrenaline, dopamine and serotonin as neurotransmitters. There is growing evidence that cocaine's effects on dopaminergic neurons, particularly those that make up the mesolimbic system, are closely associated with its rewarding properties. For example, low doses of dopamine receptor antagonists reliably influence cocaine self-administration, whereas noradrenaline and serotonin receptor antagonists are without consistent effects. Similarly, selective lesions of dopaminergic terminals in the nucleus accumbens, a major target of the mesolimbic dopamine projection, disrupt cocaine self-administration in a manner that is consistent with loss of cocaine-induced reward. The introduction of in vivo brain microdialysis as a tool with which to investigate the neurochemical correlates of motivated behaviour has provided new opportunities for investigating the role of dopamine in the nucleus accumbens in the acquisition and maintenance of cocaine self-administration. Although the body of literature that has been generated by this approach appears to contain some important inconsistencies, these probably reflect the use of inappropriate microdialysis conditions by some investigators. A critical review of the literature suggests that microdialysis results are generally consistent with a role for mesolimbic dopamine in cocaine-induced reward, although it does not seem to be the case that animals will work to maintain consistent increases in extracellular concentrations of dopamine in the nucleus accumbens in all experimental conditions. Elucidation of the complete neural circuitry of cocaine-induced reward remains an important priority for future research.

6585 POSTER Accelerated radical radiotherapy for non-small cell lung cancer (NSCLC) using two common regimens: a single centre audit of outcome

An experimental model for the study of conduction disturbances in the proximal His-Purkinje system after acute myocardial ischemia was produced in 24 dogs by ligation of the anterior septal artery. Eighteen of the 19 dogs that survived the initial vulnerable arrhythmic period showed different degrees of conduction disturbance in the bundle of His, right and left bundles or any combination thereof usually within 1 to 2 12 hours after ligation. In 14 experiments normal conduction was restored within 8 hours. In all cases with bundle branch block, the block was initially demonstrated on increasing or slowing the heart rate, with a range of intermediate rates at which conduction was normal. Within 45 minutes the block became constant at all heart rates, but within 2 to 6 hours was again replaced by normal conduction at Intermediate rates and, finally, consistent normal conduction. In 12 experiments showing intra-His bundle block (always demonstrated by splitting of the His potential into two deflections, Hb1 and Hb2) increasing the rate resulted in second or third degree block with the blocked impulse localized between the two deflections. In 5 of 12 experiments, slowing the rate gave rise to first degree intra-His bundle block (lengthening of the Hb1-Hb2 interval).Our in vitro observations suggested that the initial phase of tachycardia- and bradycardia-dependent block in the proximal His-Purkinje system can be explained by characteristic changes in the electrophysiologic properties of two groups of Purkinje fibers which may be due to differential localization of metabolites released in the ischemic zone. Tachycardia-dependent block was based on a time-dependent response of Purkinje fibers partially depolarized by an increase in extracellular concentration of potassium ion. Bradycardia-dependent block was explained by a slight to moderate decrease of resting potential, a normal slope of phase-4 depolarization and reduced membrane responsiveness associated with shift of the threshold potential toward zero. Contrary to previous data, our findings indicate no relation between enhanced automatlcity of pacemaker cells and bradycardia-dependent conduction disturbances.

Selective increase in the extracellular d-serine contents by d-cycloserine in the rat medial frontal cortex

A partial agonist of the N-methyl- d-aspartate (NMDA) receptor, d-cycloserine, acting at its glycine modulatory site, ameliorates the neuropsychiatric symptoms that are mimicked by NMDA antagonists and include cognitive disturbances, antipsychotic-resistant schizophrenic symptoms and cerebellar ataxia. To obtain a further insight into the mechanisms of the therapeutic efficacies of d-cycloserine, we investigated the effects of the systemic administration of d-cycloserine on the extracellular contents of an endogenous NMDA co-agonist, d-serine, in the medial frontal cortex of the rat using an in vivo dialysis technique. An acute intraperitoneal injection of d-cycloserine (50 and 100 mg/kg) caused an increase in extracellular concentrations of d-serine without significant effects on those of l-serine, glycine, l-glutamate, l-aspartate, l-glutamine, l-asparagine, l-alanine, l-threonin and taurine in the medial frontal cortex. The selective increase in the extracellular d-serine contents may, at least partially, be associated with the facilitating effects of d-cycloserine on the NMDA receptor functions in addition to its direct stimulation of the NMDA receptor glycine site.

Preemptive analgesia with lidocaine prevents Failed Back Surgery Syndrome

Failed Back Surgery Syndrome (FBSS) is commonly encountered in pain-treatment settings in the United States. We tested whether potential key factors in this syndrome, such as extracellular concentrations of excitatory amino acids (EAAs), are increased in the dorsal horn by synaptic release due to unintentional stretch and/or deformation/compression/transection of dorsal spinal structures during surgery. We hypothesized that pharmacological nerve block as a form of preemptive analgesia prior to any insult to dorsal root neurons will prevent an abnormally high increase in extracellular concentrations of EAAs in the dorsal horn and ultimately the establishment of central sensitization during back surgery. The L4 and L5 dorsal roots were cut bilaterally near the spinal cord to provide an adequate model to test for preemptive analgesia. Amino acid concentrations were measured by dorsal horn microdialysis sampling; EAAs aspartate and glutamate were significantly increased by 80% and 65% respectively, as were other amino acids compared to sham control values. Topical application of 1% Lidocaine ®, a voltage-gated Na + channel blocker, for 10 min prior to L4 and L5 bilateral dorsal rhizotomy (BDR) significantly attenuated the increase in EAA concentrations such that their values were not different from sham controls. Behavioral tests demonstrated significant hindlimb mechanical allodynia after BDRs that was significantly attenuated by Lidocaine® pretreatment. Thus, Lidocaine® pretreatment could offer a safe measure for prevention of chronic pain for back surgical procedures if given by intramuscular injection, topical administration onto spinal nerves and/or the dorsal spinal surface during surgical procedures that include nerve entrapment release, intervertebral disc modification and laminectomies.

Systemic administration of a dopamine, but not a serotonin or norepinephrine, transporter inhibitor reinstates cocaine seeking in the rat

Reinstatement of cocaine-seeking behavior can be elicited by a systemic priming injection of cocaine or a non-selective biogenic amine transporter inhibitor. In order to determine which biogenic amine is responsible for this effect, selective dopamine (GBR 12909), serotonin (fluoxetine) or norepinephrine (nisoxetine) transporter inhibitors were systemically administered in order to assess their ability to induce cocaine seeking in rats. Administration of GBR 12909, but not nisoxetine or fluoxetine, dose-dependently reinstated cocaine seeking in rats. Furthermore, administration of the non-selective dopamine receptor antagonist flupenthixol into the nucleus accumbens shell attenuated GBR 12909-induced reinstatement of cocaine seeking. These results suggest that increases in extracellular concentrations of dopamine, specifically in the nucleus accumbens shell, are primarily responsible for promoting cocaine priming-induced reinstatement of drug seeking in rats.

Dual effect of initial [K] on vascular tone in rat mesenteric arteries

A slight increase in extracellular concentration of potassium ([K(+)](o)) can act as a vasodilator in rat mesenteric vascular bed. However, in recent years, several groups have failed to consistently observe relaxation of rat mesenteric arteries in these conditions. The aim of the present study was to provide a mechanistic understanding of this discrepancy. In rat small mesenteric arteries, 37 of 40 arteries mounted for measurement of isometric force and pre-contracted with phenylephrine (PE) did not relax when ([K(+)](o) was raised from 5.9 mM (control ([K(+)](o) to 11.2 or 21.2 mM. However, when ([K(+)](o) was briefly lowered to 1.2 mM, increasing ([K(+)](o) to between 5.9 and 41.2 mM evoked relaxation. This relaxation was not reduced by barium or by removal of the endothelium, but was abolished by 0.1 mM ouabain. Raising ([K(+)](o) from concentrations between 0 and 5.9 mM to 13.8 mM elicited a relaxation of PE-induced tone that was inversely proportional to initial ([K(+)](o). Relaxation was associated with a ouabain-sensitive hyperpolarization of smooth muscle cells. In arteries exposed to dihydroouabain (DHO), raising ([K(+)](o) from 5.9 to 13.8 mM and simultaneously washing out DHO resulted in relaxation of PE-induced force. These results suggest that only when the initial ([K(+)](o) is less than approximately 5 mM do small elevations in ([K(+)](o) evoke smooth muscle hyperpolarization and relaxation via activation of Na,K-ATPase, and not inwardly rectifying K(+) channels. Therefore, small differences in the initial ([K(+)](o) (4.6 vs 5.9 mM) can strongly influence the variations of vascular tone to increases in ([K(+)](o).

Selective dopamine D 3 receptor antagonists enhance cortical acetylcholine levels measured with high-performance liquid chromatography/tandem mass spectrometry without anti-cholinesterases

The present study compared the effects of two selective dopamine (DA) D 3 receptor antagonists, SB-277011A (3, 10 and 30 mg/kg i.p.) and SB-414796A (3, 10 and 30 mg/kg i.p.) on extracellular levels of acetylcholine (ACh) in the rat medial prefrontal cortex (mPFC) by using a LC/MS–MS analytical method that permitted the detection of ACh without the necessity of adding acetylcholinesterase inhibitors to the perfusate. Furthermore, the present LC/MS–MS method permitted the simultaneous measurement of the respective concentrations of SB-277011A and SB-414796A in the same extracellular samples from the mPFC. The systemic administration of both selective DA D 3 receptor antagonists produced a significant increase in extracellular levels of Ach compared to vehicle-treated animals, which was associated with increases in extracellular concentrations of SB-277011A and SB-414796. Overall, the present findings further strengthen the likelihood of a modulation of cortical cholinergic function through a DA D 3-mediated mechanism and suggest that selective DA D 3 receptor antagonism may be beneficial in the treatment of psychiatric diseases, such as schizophrenia, which are characterized by cognitive dysfunction.

Identifying the Components in Eggshell Membrane Responsible for Reducing the Heat Resistance of Bacterial Pathogens

The biological activity (D-value determination) of eggshell membrane (ESM) was examined to determine the membrane components and mechanisms responsible for antibacterial activity. Biological and enzymatic activities (i.e., β-N-acetylglucosaminidase [β-NAGase], lysozyme, and ovotransferrin) of ESM denatured with trypsin, lipases, or heat were compared with those of untreated ESM. Trypsin-treated ESM lost all biological activity (D-values at 54°C were 5.12 and 5.38 min for immobilized and solubilized trypsin, respectively) but showed no significant loss of enzymatic activities. Treatments with porcine lipase and a lipase cocktail did not impact biological or enzymatic activities. Heat denaturation of ESM (at 80 and 100°C for 15 min) resulted in significant decreases in biological activity (D-values of 3.99 and 4.43 min, respectively) and loss of β-NAGase activity. Lysozyme and ovotransferrin activities remained but were significantly reduced. Purified ESM and hen egg white components (i.e., β-NAGase, lysozyme, and ovotransferrin) were added to Salmonella Typhimurium suspensions (in 0.1% peptone water) at varying concentrations to evaluate their biological activity. D-values at 54°C were 4.50 and 3.68 min for treatment with lysozyme or β-NAGase alone, respectively, and 2.44 min for ovotransferrin but 1.47 min for a combination of all three components (similar to values for ESM). Exposure of Salmonella Typhimurium cells to a mixture of ovotransferrin, lysozyme, and β-NAGase or ESM resulted in significant increases in extracellular concentrations of Ca2+, Mg2+, and K+. Transmission electron microscopic examination of Salmonella Typhimurium cells treated with a combination of ovotransferrin, lysozyme, and β-NAGase revealed membrane disruption and cell lysis. The findings of this study demonstrate that ovotransferrin, lysozyme, and β-NAGase are the primary components responsible for ESM antibacterial activity. The combination of these proteins and perhaps other ESM components interferes with interactions between bacterial lipopolysaccharides, sensitizing the outer bacterial membrane to the lethal affects of heat and possibly pressure and osmotic stressors.