Interleukin-1 stimulates hypothalamic inhibitory amino acid neurotransmitter release

In order to get a better insight into the function of amino acid residues located in the second transmembrane domain of the cystic fibrosis transmembrane conductance regulator (CFTR) protein, all exon 18 mutations found in cystic fibrosis (CF) patients were characterized at the protein and at the electrophysiological level. Of the different mutations present in transmembrane helix 12 (M1137V, M1137R, I11139V and deltaM1140), and the intracytoplasmic loop connecting TM12 and NBD2 (D1152H and D1154G), only M1137R interfered with the proper maturation of the protein. Permeability studies performed after injection of the different wild-type and mutant cRNAs in Xenopus laevis oocytes indicated that the mutations did not alter the permeability sequence of the CFTR channels. The whole cell cAMP activated chloride currents, however, were significantly reduced for M1137V, I1139V, D1152H and D1154G and close to zero for deltaM1140, indicating that these mutations interfere with the proper gating of the chloride channels.     

ATP receptor-mediated enhancement of fast excitatory neurotransmitter release in the brain

In recent years considerable effort has been invested toward understanding the molecular mechanisms that regulate and restrict DNA replication to once per each cell cycle. An important contribution came from studying the phenomenon of endoreduplication-an endonuclear duplication of chromosomes which occurs in the absence of mitosis leading to the production of chromosomes with doubling series of chromatids. Because endoreduplicating nuclei retain the capability of replication without passing through mitosis, they provide a unique system for studying the molecular mechanisms that restrict DNA replication to once per cycle. Three types of endoreduplication can be identified: I, multiple initiations within a given S phase; II, reoccurring S phase; and III, repeated S and Gap phases. Each of these illuminates a different control level acting over the onset of S phase, which coordinately restrict DNA synthesis to once per each cell cycle.     

Ionic and haemodynamic changes influence the release of the excitatory amino acid glutamate in the posterior hypothalamus

Argyrophilic and tau-positive abnormal structures occurring in glial cells are called glial fibrillary tangles. In the astrocyte, a conspicuous tau-positive structure is known to appear in progressive supranuclear palsy (PSP). In this report, another type of argyrophilic and tau-positive astrocytes is reported. The morphology of this new type is quite different from that of the previously reported tau-positive astrocyte in PSP and they are designated here as thorn-shaped astrocytes (TSA). TSA have an apparently argyrophilic cytoplasm with a few short processes and often have a small eccentric nucleus, whose appearance resembles that of a reactive astrocyte. Immunohistochemically, TSA are positive to anti-tau antibodies but are negative for ubiquitin. Simultaneous immunostaining revealed the coexistence of tau and glial fibrillary acidic protein epitopes in the same cytoplasm. Electron microscopically, bundles of 15-nm straight tubules were included in the cytoplasm together with abundant glial filaments. In the vicinity of a cluster of TSA, related structures of perivascular or subpial tau-positive linings, which correspond to astrocytic end-feet, are sometimes observed. In almost all cases, a few TSA are generally located in a confined area of subpial and subependymal regions. Although TSA appear to be intimately associated with some diseases, they are also found in a wide range of cytoskeletal disorders including the aged brain with neurofibrillary tangles. TSA are presumed to be a secondary induced product in relation to astrocytic reaction.     

Prenatal ethanol exposure diminishes activity-dependent potentiation of amino acid neurotransmitter release in adult rat offspring

Prenatal ethanol exposure has been associated with long-lasting intellectual impairments in children. Previous studies suggest that these deficits are, in part, linked to neurochemical abnormalities that reduce the ability to sustain long-term potentiation (LTP) in hippocampal formation of adult offspring. One presynaptic component of LTP that manifests during the first half-hour after tetanic stimulation is an enhancement of amino acid neurotransmitter release. Given that the onset of enhanced neurotransmitter release correlates temporally with the decay of hippocampal LTP in prenatal ethanol-exposed offspring, we tested the hypothesis that prenatal ethanol exposure reduces tetanus stimulus-induced potentiation of electrically evoked amino acid release in hippocampal slices. Rat dams consumed 1 of 3 diets throughout gestation: (1) a BioServ liquid diet containing 5% (v/v) ethanol (26% ethanol-derived calories) that produces a maternal peak blood ethanol concentration of 83 mg/dl; (2) pair-fed an isocalorically equivalent amount of 0% ethanol liquid diet; or (3) Purina rat chow ad libitum. Hippocampal slices were prepared from adult offspring from each experimental diet group. Neither the amount of hippocampal slice tissue protein nor the incorporation of [3H]-D-aspartate (D-ASP) was affected by prenatal ethanol exposure. Furthermore, spontaneous efflux and electrically evoked D-ASP release were similar among the three diet groups. However, tetanus stimulus-induced potentiation of evoked D-ASP release in prenatal ethanol-exposed offspring was reduced to about one-third of the potentiation of D-ASP release observed in the control diet groups. These results suggest that prenatal ethanol exposure produces long-lasting deficits in the neurochemical mechanisms responsible for activity-dependent potentiation of amino acid transmitter release without affecting the synaptic machinery responsible for amino acid uptake, storage, and release.     

Role of excitatory amino acids in the regulation of dopamine synthesis and release in the neostriatum

We have explored the role of excitatory amino acids in the increased dopamine (DA) release that occurs in the neostriatum during stress-induced behavioral activation. Studies were performed in awake, freely moving rats, using in vivo microdialysis. Extracellular DA was used as a measure of DA release; extracellular 3,4-dihydroxyphenylalanine (DOPA) after inhibition of DOPA decarboxylase provided a measure of apparent DA synthesis. Mild stress increased the synthesis and release of DA in striatum. DA synthesis and release also were enhanced by the intra-striatal infusion of N-methyl-D-aspartate (NMDA), an agonist at NMDA receptors, and kainic acid, an agonist at the DL-alpha-amino-3-hydroxy-5-methyl-4-isoxazole-4-propionate (AMPA)/kainate site. Stress-induced increase in DA synthesis was attenuated by co-infusion of 2-amino-5-phosphonovalerate (APV) or 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX), antagonists of NMDA and AMPA/kainate receptors, respectively. In contrast, intrastriatal APV, CNQX, or kynurenic acid (a non-selective ionotropic glutamate receptor antagonist) did not block the stress-induced increase in DA release. Stress-induced increase in DA release was, however, blocked by administration of tetrodotoxin along the nigrostriatal DA projection. It also was attenuated when APV was infused into substantia nigra. Thus, glutamate may act via ionotropic receptors within striatum to regulate DA synthesis, whereas glutamate may influence DA release via an action on receptors in substantia nigra. However, our method for monitoring DA synthesis lowers extracellular DA and this may permit the appearance of an intra-striatal glutamatergic influence by reducing a local inhibitory influence of DA. If so, under conditions of low extracellular DA glutamate may influence DA release, as well as DA synthesis, by an intrastriatal action. Such conditions might occur during prolonged severe stress and/or DA neuron degeneration. These results may have implications for the impact of glutamate antagonists on the ability of patients with Parkinson's disease to tolerate stress.     

Direct and indirect presynaptic control of dopamine release by excitatory amino acids

Dopamine (DA) release from nerve terminals of the nigrostriatal DA neurons not only depends on the activity of nigral DA cells but also on presynaptic regulation. Glutamatergic neurons of cortical origin play a prominent role in these presynaptic regulations. The direct glutamatergic presynaptic control of DA release is mediated by N-methyl-D-aspartate (NMDA) and alpha-amino-3-hydroxy-5-methyl-4-isoxazole-4-propionate (AMPA) receptors, located on DA nerve terminals. In addition, by acting on striatal target cells, these glutamatergic neurons contribute also to indirect regulations of DA release involving several transmitters such as GABA, acetylcholine and neuropeptides. Diffusible messengers such as nitric oxide (NO) or arachidonic acid (AA) which are particularly formed under the stimulation of NMDA receptors may also participate to the regulation of DA release. In the present study, it will be shown that the co-application of NMDA and carbachol synergistically increases the release of [3H]-DA and that this effect is reduced by mepacrine or 4-bromophenacylbromide (10(-7) M), two inhibitors of PLA2. Therefore endogenously released AA induced by the co-stimulation of NMDA and cholinergic receptors seems to be involved, at least partly, in the release of DA.     

Serum factor in Miller-Fisher variant of Guillain-Barré syndrome and neurotransmitter release

Serum IgG autoantibodies to GQ1b ganglioside are associated with the acute phase of the Miller-Fisher syndrome (MFS). We investigated the effects of three anti-GQ1b-positive MFS sera in the mouse phrenic-nerve/diaphragm preparation. Miniature endplate potential frequencies increased eight-fold within 25 min, declined rapidly, and ceased altogether after 3 h, when nerve stimulation no longer evoked a response. One MFS convalescent serum (anti-GQ1b negative) and sera from healthy controls and from patients with other neurological diseases were without effect. Thus muscle weakness in MFS may be caused by a serum factor, likely to be GQ1b antibody, that leads to failure of acetylcholine release from motor nerve terminals.     

The effect of losigamone (AO-33) on electrical activity and excitatory amino acid release in mouse cortical slices

1. Losigamone is a novel anticonvulsant the mechanism of action of which is not known. This study investigated the effect of losigamone on spontaneous, NMDA- and AMPA-induced depolarizations in the cortical wedge preparation of the DBA/2 mouse (which are susceptible to sound-induced seizures) and on endogenous amino acid release from BALB/c mouse cortical slices. 2. Cortical wedges exhibit spontaneous depolarizations in magnesium-free medium and losigamone was effective in significantly reducing these spontaneous depolarizations at concentrations of 100 microM and above. 3. NMDA-induced depolarizations were significantly reduced by losigamone at concentrations of 25 microM and above. Losigamone had no effect on AMPA-induced depolarizations. 4. Veratridine (20 microM) and potassium (60 mM) were used to stimulate the release of amino acids from mouse cortex. Veratridine-stimulated release of glutamate was significantly reduced by losigamone at concentrations of 100 microM and above, while potassium-stimulated release was significantly reduced by losigamone at 200 microM. 5. NMDA antagonism and inhibition of excitatory amino acid release may contribute to the anticonvulsant effect of losigamone.     

Preoptic rather than mediobasal hypothalamic amino acid neurotransmitter release regulates GnRH secretion during the estrogen-induced LH surge in the ovariectomized rat

Inhibitory and excitatory amino acid neurotransmitters have been suggested to participate in the feedback actions of estradiol (E2) on LH secretion. In the rat estrogen-receptive neurons have been demonstrated in the preoptic/anterior hypothalamic area (POA) and mediobasal hypothalamus/median eminence (MBH) and many of these neurons utilize gamma-aminobutyric acid (GABA) as neurotransmitter. The actions of excitatory amino acids (EAA) differ in ovariectomized (ovx) and ovx E2-substituted rats indicating that EAAs also participate in the positive feedback action of E2 on LH release. However, little information is available as to whether in vivo these transmitters exert their effects in the POA, where most of the GnRH perikarya are located, or in the MBH, i.e. at the nerve terminals. Therefore we conducted push pull cannula perfusions to compare the release rates of GABA, aspartate (ASP) and glutamate (GLU) in the MBH and POA. A subcutaneous implant of a silastic tube containing E2 resulted in LH surges in the afternoon of all treated animals. Prior to and during this LH surge the MBH release rates of neither GABA nor ASP nor GLU were significantly altered. In contrast, a conspicuous drop in preoptic GABA release occurred prior to and during the time of estrogen-induced LH surges and this was accompanied by enhanced preoptic secretion of ASP and GLU. In conclusion, we present the first data about amino acid release in the MBH during the E2-induced LH surge. Since only in the POA the LH surge is associated with changes in amino acid release, it appears that both inhibitory and excitatory amino acids act at the level of the GnRH cell bodies and/or dendrites and not on GnRH nerve terminals to mediate the feedback mechanism of E2 on LH release.     

Recent basic findings in support of excitatory amino acid hypotheses of schizophrenia

1. Several clinical and post-mortem tissue findings have suggested a role for excitatory amino acid neuronal systems in the pathophysiology of schizophrenia. 2. These include the ability of NMDA antagonists, phencyclidine and ketamine, to cause both negative and positive symptoms in healthy subjects, and abnormalities in the densities of some types of excitatory amino acid receptors in the postmortem tissue of schizophrenic brains. 3. The present review describes recent basic findings that have examined the involvement of excitatory amino acids in the mechanism of action of antipsychotic drugs. These include studies on the functional links between glutamatergic and dopaminergic systems, effect of acute and chronic antipsychotic drug treatment on excitatory amino acid function, and stress-induced activation of excitatory amino acid release, in particular in the prefrontal cortex.     

Effects of nitric oxide synthase gene knockout on neurotransmitter release in vivo

Previous studies suggest that low bone mass is a complication of alcoholic liver disease. Nevertheless, little is known about bone mass and bone metabolism in viral cirrhosis. To evaluate the prevalence and magnitude of hepatic osteopenia in these patients, bone remodeling status, and its relationship with the severity of liver disease and serum levels of insulin-like growth factor I (IGF-I), we studied 32 consecutive patients with viral cirrhosis and no history of alcohol intake. Bone mineral density (BMD) was measured by dual x-ray absorptiometry in the lumbar spine (LS) and femoral neck (FN), and the values were expressed as the z score. Bone metabolism markers and hormone profiles were measured. Patients with viral cirrhosis showed reduced BMD in all sites (LS: -1.27 +/- 1.06, P < .001; FN: -0.48 +/- 0.96; P < .01). Of the 32 patients, 53% met the diagnostic criteria for osteoporosis. In patients, urine deoxypyridinoline (D-Pyr) as a marker of bone resorption and serum bone alkaline phosphatase (b-AP) as a marker of bone formation were significantly higher than in control subjects (P < .001 and P < .01, respectively). Serum IGF-I was lower than in control subjects (P < .001), and significant differences were also found between patients with and without osteoporosis (P < .05). BMD in LS correlated with severity of the disease, with serum levels of IGF-I, and with urine D-Pyr. Our findings show that viral cirrhosis is a major cause of osteoporosis in men, and that low serum IGF-I levels seem to play a role in the bone mass loss in these patients. The biochemical markers of bone remodeling suggest high-turnover osteoporosis in patients with viral cirrhosis.     

Enhancement of neurotransmitter release induced by brain-derived neurotrophic factor in cultured hippocampal neurons

Brain-derived neurotrophic factor (BDNF), like other neurotrophins, has long-term effects on neuronal survival and differentiation; furthermore, recent work has shown that BDNF also can induce rapid changes in synaptic efficacy. We have investigated the mechanism(s) of these synaptic effects on cultured embryonic hippocampal neurons. In the presence of the GABAA receptor antagonist, picrotoxin, the application of BDNF (100 ng/ml) for 1-5 min increased the amplitude of evoked synaptic currents by 48 +/- 9% in 10 of 15 pairs of neurons and increased the frequency of EPSC bursts to 205 +/- 20% of the control levels. There was no detectable effect of BDNF on various measures of electrical excitability, including the resting membrane potential, input resistance, action potential threshold, and action potential amplitude. In addition, BDNF did not change the postsynaptic currents induced by the exogenous application of glutamate. BDNF did increase the frequency of miniature EPSCs (mEPSCs) (268.0 +/- 46.8% of control frequency), however, without affecting the mEPSC amplitude. The effect of BDNF on mEPSC frequency was blocked by the tyrosine kinase inhibitor K252a and also by the removal of extracellular calcium ([Ca2+]o). Fura-2 recordings showed that BDNF elicited an increase in intracellular calcium concentration ([Ca2+]c). This effect was dependent on [Ca2+]o; it was blocked by K252a and by thapsigargin, but not by caffeine. The results demonstrate that BDNF enhances glutamatergic synaptic transmission at a presynaptic locus and that this effect is accompanied by a rise in [Ca2+]c that requires the release of Ca2+ from IP3-gated stores.