Differential effects of arginine, glutamate and phosphoarginine on Ca2+-activation properties of muscle fibres from crayfish and rat

The effects of two amino acids, arginine which has a positively charged side-chain and glutamate which has a negatively charged side-chain on the Ca²⁺-activation properties of the contractile apparatus were examined in four structurally and functionally different types of skeletal muscle; long- and short-sarcomere fibres from the claw muscle of the yabby (a freshwater decapod crustacean), and fast- and slow-twitch fibres from limb muscles of the rat. Single skinned fibres were activated in carefully balanced solutions of different pCa (-log₁₀[Ca²⁺]) that either contained the test solute (“test”) or not (“control”). The effect of phosphoarginine, a phosphagen that bears a nett negative charge, was also compared to the effects of arginine. Results show that (i) arginine (33-36 mmol l^-1) significantly shifted the force–pCa curve by 0.08–0.13 pCa units in the direction of increased sensitivity to Ca²⁺-activated contraction in all fibre types; (ii) phosphoarginine (9–10 mmol l^-1) induced a significant shift of the force–pCa curve by 0.18–0.24 pCa units in the direction of increased sensitivity to Ca²⁺ in mammalian fast- and slow-twitch fibres, but had no significant effects on the force–pCa relation in either long- or short-sarcomere crustacean fibres; (iii) glutamate (36–40 mmol l^-1), like arginine affected the force–pCa relation of all fibre types investigated, but in the opposite direction, causing a significant decrease in the sensitivity to Ca²⁺-activated contraction by 0.08–0.19 pCa units; (iv) arginine, phosphoarginine and glutamate had little or no effect on the maximum Ca²⁺-activated force of crustacean and mammalian fibres. The results suggest that the opposing effects of glutamate and arginine are not related to simply their charge structure, but must involve complex interactions between these molecules, Ca²⁺ and the regulatory and other myofibrillar proteins.

Wireless instantaneous neurotransmitter concentration system-based amperometric detection of dopamine, adenosine, and glutamate for intraoperative neurochemical monitoring - laboratory investigation

Object  In a companion study, the authors describe the development of a new instrument named the Wireless Instantaneous Neurotransmitter Concentration System (WINCS), which couples digital telemetry with fast-scan cyclic voltammetry (FSCV) to measure extracellular concentrations of dopamine. In the present study, the authors describe the extended capability of the WINCS to use fixed potential amperometry (FPA) to measure extracellular concentrations of dopamine, as well as glutamate and adenosine. Compared with other electrochemical techniques such as FSCV or high-speed chronoamperometry, FPA offers superior temporal resolution and, in combination with enzyme-linked biosensors, the potential to monitor nonelectroactive analytes in real time.

Methods  The WINCS design incorporated a transimpedance amplifier with associated analog circuitry for FPA; a microprocessor; a Bluetooth transceiver; and a single, battery-powered, multilayer, printed circuit board. The WINCS was tested with 3 distinct recording electrodes: 1) a carbon-fiber microelectrode (CFM) to measure dopamine; 2) a glutamate oxidase enzyme–linked electrode to measure glutamate; and 3) a multiple enzyme–linked electrode (adenosine deaminase, nucleoside phosphorylase, and xanthine oxidase) to measure adenosine. Proof-of-principle analyses included noise assessments and in vitro and in vivo measurements that were compared with similar analyses by using a commercial hardwired electrochemical system (EA161 Picostat, eDAQ; Pty Ltd). In urethane-anesthetized rats, dopamine release was monitored in the striatum following deep brain stimulation (DBS) of ascending dopaminergic fibers in the medial forebrain bundle (MFB). In separate rat experiments, DBS-evoked adenosine release was monitored in the ventrolateral thalamus. To test the WINCS in an operating room setting resembling human neurosurgery, cortical glutamate release in response to motor cortex stimulation (MCS) was monitored using a large-mammal animal model, the pig.

Results   The WINCS, which is designed in compliance with FDA-recognized consensus standards for medical electrical device safety, successfully measured dopamine, glutamate, and adenosine, both in vitro and in vivo. The WINCS detected striatal dopamine release at the implanted CFM during DBS of the MFB. The DBS-evoked adenosine release in the rat thalamus and MCS-evoked glutamate release in the pig cortex were also successfully measured. Overall, in vitro and in vivo testing demonstrated signals comparable to a commercial hardwired electrochemical system for FPA.

Conclusions  By incorporating FPA, the chemical repertoire of WINCS-measurable neurotransmitters is expanded to include glutamate and other nonelectroactive species for which the evolving field of enzyme-linked biosensors exists. Because many neurotransmitters are not electrochemically active, FPA in combination with enzyme-linked microelectrodes represents a powerful intraoperative tool for rapid and selective neurochemical sampling in important anatomical targets during functional neurosurgery.

The specificity of platelet glutamate receptor supersensitivity in psychotic disorders.

Hypoglutamatergic function is implicated in the pathogenesis of schizophrenia, and supersensitivity of platelet NMDA receptors has been reported in schizophrenia. The aim of this study was to examine the platelet glutamate receptor sensitivity in patients with schizophrenia (n=12), mania with psychotic features (n=10) and depression with psychotic features (n=10) and matched controls (n=12) in order to assess if this is a marker of schizophrenia or occurs in other psychotic conditions. Glutamate receptor sensitivity was assessed using the intracellular calcium response to glutamate measured with spectrofluorometry. The percentage response of the schizophrenic and depressed psychotic subjects to glutamate stimulation was significantly greater than control subjects (p<0.005). The mania with psychotic features group was not significantly different to controls. This data suggests that platelet glutamate receptors may be supersensitive in schizophrenia and depression with psychotic features. Furthermore, the platelet may be a possible peripheral marker of glutamate function in schizophrenia and depression with psychotic features.

Platelet glutamate receptor supersensitivity in major depressive disorder.

Dysregulation of glutamate has been described in depression, and supersensitivity of platelet glutamate receptors has been found in both psychotic major depression and schizophrenia. The aim of this study was to examine the platelet glutamate receptor sensitivity in patients with nonpsychotic, unipolar major depression to assess whether this is a marker of depression or of psychosis. Glutamate receptor sensitivity was assessed using the platelet intracellular calcium response to glutamate (0-100 micromol) measured by spectrofluorometry. The depression group showed a significantly greater platelet intracellular calcium response to glutamate stimulation than the control group, both in terms of absolute values (p = 0.007) and percentage of response from baseline (p = 0.030). These data suggest that platelet glutamate receptors may be supersensitive in depression and that the platelet may be a possible peripheral marker of glutamate function in depression.

Supersensitive platelet glutamate receptors as a possible peripheral marker in schizophrenia.

Hypoglutamatergic function is implicated in the pathogenesis of schizophrenia. The aim of this study was to examine the platelet intracellular calcium response to glutamate using spectroflourometry in 15 schizophrenic patients and 15 matched control individuals as an index of platelet glutamate receptor sensitivity. Patients with schizophrenia had significantly lower baseline intracellular calcium levels than matched control individuals (P = 0.03). The percentage response of the schizophrenic individuals to glutamate stimulation was significantly greater than control individuals (P < 0.001). These data suggest that platelet glutamate receptors may be supersensitive in schizophrenia. Furthermore, the platelet may be a possible peripheral marker of glutamate function in schizophrenia.

Metabotropic glutamate receptors as targets for novel antipsychotic treatments

In recent years metabotropic glutamate receptors have emerged as key targets for the design of new antipsychotic medications for schizophrenia, in particular mGluR5 and mGluR2/3. These receptors exhibit diverse interactions with other neurotransmitter receptors and critical elements of intracellular signalling cascades known to be important to the pharmacotherapy of schizophrenia. In addition, mGluR5 and mGluR2/3 are intimately involved in behavioural domains related to the symptoms of this disorder. Both animal and clinical studies using novel drugs targeting these receptors have provided encouraging results. The number of patents registered for drugs targeting metabotropic glutamate receptors has grown dramatically, and positive allosteric modulators for both receptors show particular promise.

Clozapine reverses schizophrenia-related behaviours in the metabotropic glutamate receptor 5 knockout mouse: association with N-methyl-d-aspartic acid receptor up-regulation

Abnormalities in glutamatergic signalling are proposed in schizophrenia in light of the schizophreniform psychosis elicited by NMDA antagonists. The metabotropic glutamate receptor 5 (mGluR5) interacts closely with the NMDA receptor and is implicated in several behavioural endophenotypes of schizophrenia. We have demonstrated that mice lacking mGluR5 have increased sensitivity to the hyperlocomotive effects of the NMDA antagonist MK-801. Mice lacking mGluR5 also show abnormal locomotor patterns, reduced prepulse inhibition (PPI), and deficits on performance of a short-term spatial memory task on the Y-maze. Chronic administration of the antipsychotic drug clozapine ameliorated the locomotor disruption and reversed the PPI deficit, but did not improve Y-maze performance. Chronic clozapine increased NMDA receptor binding ([3H]MK-801) but did not alter dopamine D2 ([3H]YM-09151), 5-HT2A ([3H]ketanserin), or muscarinic M1/M4 receptor ([3H]pirenzepine), binding in these mice. These results demonstrate behavioural abnormalities that are relevant to schizophrenia in the mGluR5 knockout mouse and a reversal of behaviours with clozapine treatment. These results highlight both the interactions between mGluR5 and NMDA receptors in the determination of schizophreniform behaviours and the potential for the effects of clozapine to be mediated by NMDA receptor regulation.
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Cross-fostering reduces obesity induced by early exposure to monosodium glutamate in male rats

Maternal obesity programmes a range of metabolic disturbances for the offspring later in life. Moreover, environmental changes during the suckling period can influence offspring development. Because both periods significantly affect long-term metabolism, we aimed to study whether cross-fostering during the lactation period was sufficient to rescue a programmed obese phenotype in offspring induced by maternal obesity following monosodium L-glutamate (MSG) treatment. Obesity was induced in female Wistar rats by administering subcutaneous MSG (4 mg/g body weight) for the first 5 days of postnatal life. Control and obese female rats were mated in adulthood. The resultant pups were divided into control second generation (F2) (CTLF2), MSG-treated second generation (F2) (MSGF2), which suckled from their CTL and MSG biological dams, respectively, or CTLF2-CR, control offspring suckled by MSG dams and MSGF2-CR, MSG offspring suckled by CTL dams. At 120 days of age, fat tissue accumulation, lipid profile, hypothalamic leptin signalling, glucose tolerance, glucose-induced, and adrenergic inhibition of insulin secretion in isolated pancreatic islets were analysed. Maternal MSG-induced obesity led to an obese phenotype in male offspring, characterized by hyperinsulinaemia, hyperglycaemia, hyperleptinaemia, dyslipidaemia, and impaired leptin signalling, suggesting central leptin resistance, glucose intolerance, impaired glucose-stimulated, and adrenergic inhibition of insulin secretion. Cross-fostering normalized body weight, food intake, leptin signalling, lipid profiles, and insulinaemia, but not glucose homeostasis or insulin secretion from isolated pancreatic islets. Our findings suggest that alterations during the lactation period can mitigate the development of obesity and prevent the programming of adult diseases.

Modifying the bitterness of selected oral pharmaceuticals with cation and anion series of salts

Purpose. NaCl has proven to be an effective bitterness inhibitor, but the reason remains unclear. The purpose of this study was to examine the influence of a variety of cations and anions on the bitterness of selected oral pharmaceuticals and bitter taste stimuli: pseudoephedrine, ranitidine, acetaminophen, quinine, and urea.
Method. Human psychophysical taste evaluation using a whole mouth exposure procedure was used.
Results. The cations (all associated with the acetate anion) inhibited bitterness when mixed with pharmaceutical solutions to varying degrees. The sodium cation significantly (P < 0.003) inhibited bitterness of the pharmaceuticals more than the other cations. The anions (all associated with the sodium cation) also inhibited bitterness to varying degrees. With the exception of salicylate, the glutamate and adenosine monophosphate anions significantly (P < 0.001) inhibited bitterness of the pharmaceuticals more than the other anions. Also, there were several specific inhibitory interactions between ammonium, sodium and salicylate and certain pharmaceuticals.
Conclusions. We conclude that sodium was the most successful cation and glutamate and AMP were the most successful anions at inhibiting bitterness. Structure forming and breaking properties of ions, as predicted by the Hofmeister series, and other physical-chemical ion properties failed to significantly predict bitterness inhibition.

The influence of sodium salts on binary mixtures of bitter-tasting compounds

In order to study potential mixture interactions among bitter compounds, selected sodium salts were added to five compounds presented either alone or as binary bitter- ompound mixtures. Each compound was tested at a concentration that elicited ‘weak’ perceived bitterness. The bitter compounds were mixed at these concentrations to form a subset of possible binary mixtures. For comparison, the concentration of each solitary compound was doubled to measure bitterness inhibition at the higher intensity level elicited by the mixtures. The following sodium salts were tested for bitterness inhibition: 100 mM sodium chloride (salty), 100 mM sodium gluconate (salty), 100 and 20 mM monosodium glutamate (umami), and 50 mM adenosine monophosphate disodium salt (umami). Sucrose (sweet) was also employed as a bitterness suppressor. The sodium salts differentially suppressed the bitterness of compounds and their binary combinations. Although most bitter compounds were suppressed, the bitterness of tetralone was not suppressed, nor was the bitterness of the binary mixtures that contained it. In general, the percent suppression of binary mixtures of compounds was predicted by the average percent suppression of its two components. Within the constraints of the present study, the bitterness of mixtures was suppressed by sodium salts and sucrose independently, with few bitter interactions. This is consistent with observations that the bitter taste system integrates the bitterness of multi-compound solutions linearly.

Effects of dietary omega-3 polyunsaturated fatty acids on brain gene expression

Polyunsaturated fatty acids (PUFA) are essential structural components of the central nervous system. Their role in controlling learning and memory has been well documented. A nutrigenomic approach with high-density microarrays was used to reveal brain gene-expression changes in response to different PUFA-enriched diets in rats. In aged rats fed throughout life with PUFA-enriched diets, genes with altered expressions included transthyretin, α-synuclein, and calmodulins, which play important roles in synaptic  plasticity and learning. The effect of perinatal omega-3 PUFA supply on gene expression later in life also was studied. Several genes showed similar changes in expression in rats fed omega-3-deficient diets in the perinatal period, regardless of whether they or their mothers were fed omega-3 PUFA-sufficient diets after giving birth. In this experiment, among the down-regulated genes were a kainate glutamate receptor and a DEAD-box polypeptide. Among the up-regulated genes were a chemokine-like factor, a tumor necrosis factor receptor, and cytochrome c. The possible involvement of the genes with altered expression attributable to different diets in different brain regions in young and aged rats and the possible mode of regulatory action of PUFA also are discussed. We conclude that PUFA-enriched diets lead to significant changes in expression of several genes in the central nervous tissue, and these effects appear to be mainly independent of their effects on membrane composition. The direct effects of PUFA on transcriptional modulators, the downstream developmentally and tissue-specifically activated elements might be one of the clues to understanding the beneficial effects of the omega-3 PUFA on the nervous system.

Larval settlement of the tropical abalone, Haliotis asinina Linnaeus, using natural and artificial chemical inducers

Many kinds of chemical and biological materials have been used as inducers of settlement of abalone larvae, as well as other species of marine gastropods, with responses being highly variable, even to the same chemical cue. The present study tested chemical inducers, γ-aminobutyric acid (GABA), δ-aminovaleric acid (5-AVA) and l-glutamic acid (GA) and the effects they have on larval settlement of Haliotis asinina. Additionally, a relatively inexpensive commercial substance, monosodium glutamate (MSG), was trialed. The datum provided shows all chemicals to be active inducers of settlement in this study, in order of effectiveness of 5-AVA, GABA, MSG to GA. Induction as adjudged from larval numbers settled was best at 6 h 62%, with 10⁻¹ mM 5-AVA. At 24 h, induction was the highest at 78% when exposed to 10⁻² mM 5-AVA. Larvae that were allowed to settle up to 72 h showed the highest numbers of settled larvae, and declined back to 60% when exposed to 10⁻² 5-AVA and 10⁻¹ mM GABA respectively. Monosodium glutamate, although third in settlement standings would bypass the other chemicals, with regard to cost versus yield. The assessment of settlement surface, rough or smooth proved to be irrelevant, which had no significant impact on larval settlement.