Formation of extracellular glutamate from glutamine: exclusion of pyroglutamate as an intermediate

A 4.6-fold increase in interstitial glutamate was observed following the reverse microdialysis of 5 mM glutamine into the rat hippocampus. Two possible mechanisms of glutamine hydrolysis were examined: (a) an enzymatic glutaminase activity and (b) a non-enzymatic mechanism. Injection of 14C-glutamine at the site of microdialysis followed by microdialysis with artificial cerebral spinal fluid allowed isolation of 14C-glutamine (63%), 14C-glutamate (14%), and a compound tentatively identified as pyroglutamate (22%). In this study, we determined if non-enzymatic pyroglutamate formation from glutamine contributed to the synthesis of glutamate. Pyroglutamate is in chemical equilibrium with glutamate, although under physiological conditions, the chemical equilibrium is strongly in the direction of pyroglutamate. In vitro stability studies indicated that 14C-glutamine and 14C-pyroglutamate are not subject to significant non-enzymatic breakdown at pH 6.5-7.5 at 37 degrees C for up to 8 h. Reverse microdialysis with 1 mM pyroglutamate did not increase interstitial glutamate levels. Following injection of 14C-pyroglutamate and microdialysis, radioactivity was recovered in 14C-pyroglutamate (88%) and 14C-glutamine (11%). Less than 1% of the radioactivity was recovered as glutamate. Our data do not support a role of pyroglutamate as an intermediate in the formation of extracellular glutamate following the infusion of glutamine. However, it confirms that pyroglutamate, a known constituent in brain, is actively metabolized in brain cells and contributes to glutamine in the interstitial space.     

Neonatal exposure to monosodium L-glutamate induces loss of neurons and cytoarchitectural alterations in hippocampal CA1 pyramidal neurons of adult rats

Glutamatergic post-synaptic receptors are closely related to the known excitotoxic effects of high doses of L-glutamate. Several behavioral abnormalities, glial reaction, and an increase of expression of the NMDA receptor sub-units have been observed in the rat hippocampus after early monosodium glutamate exposure. Thus, a quantitative morphological study was carried out to determine the effects of early exposure to monosodium glutamate on post-synaptic structures that mediate glutamate excitatory neurotransmission in the hippocampal CA1 field. Four milligrams per gram body weight of monosodium glutamate was subcutaneously injected into neonatal Wistar rats, at 1, 3, 5, and 7 days. Cell loss and several cytoarchitectonic parameters were evaluated in pyramidal cells from the hippocampal CA1 field in the treated rats at 60 days of age. An untreated group of rats were used as controls. Cell number in the hippocampus of experimental rats was 11.5% less than that in control animals. In addition, both dendritic arborization and dendritic spine density were adversely affected, and thin and mushroom-shaped spines became proportionally more numerous, while the opposite occurred to stubby spines. These results strongly suggest the occurrence of cell death and also show some cytoarchitectural modifications in the surviving neurons. These could lead to functional alterations in the hippocampal integrative activity, due to an early cytoexcitotoxic effect of monosodium glutamate.     

N1-dansyl-spermine: a potent polyamine antagonist

The potential polyamine antagonist action of N1-dansyl-spermine (a potent NMDA antagonist) was assessed in two in vivo mouse models of polyamine action. Co-administration of N1-dansyl-spermine (2-10 microg, i.c.v.) with spermine (100 microg, i.c.v.) resulted in a dose-dependent antagonism of the spermine-induced CNS excitation (body tremor and fatal tonic convulsions). In addition, the same dose of N1-dansyl-spermine antagonised spermine's enhancement of NMDA-induced convulsions. These results suggest that N1-dansyl-spermine is in vivo a potent antagonist of the CNS effects of spermine and of its action at the positive polyamine modulatory site on the NMDA receptor.     

Contribution of mitogen-activated protein kinases to NMDA-induced neurotoxicity in the rat retina

We examined the contributions of the mitogen-activated protein kinases (MAPKs) family [extracellular signal-regulated kinase (ERK), p38 kinase (p38), and c-Jun N-terminal kinase (JNK)] to N-methyl-D-aspartate (NMDA)-induced neurotoxicity in the rat retina. Detection of apoptotic cell death in the retinal ganglion cell layer (RGCL) and the inner nuclear layer (INL) by terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick-end labeling (TUNEL) staining began 6 h after intravitreal NMDA (100 nmol) injection and continued to increase thereafter. Western blot analysis showed that phosphorylated MAPKs (p-MAPKs) were expressed in the retina following a temporal manner: maximal expression of phosphorylated ERK (p-ERK) at 1 h, maximal expression of phosphorylated p38 (p-p38) at 6 h, and beginning of phosphorylated JNK (p-JNK) significant increase at 6 h after injection. An immunohistochemical/TUNEL co-localization study showed that p-JNK- and p-p38-positive cells in the RGCL were frequently TUNEL-positive, whereas few p-ERK-positive cells were TUNEL-positive. Moreover, co-injection of inhibitors for JNK (0.2 nmol SP600125) and/or p38 (2.0 nmol SB203580) with NMDA was effective in ameliorating NMDA-induced apoptotic cell loss in the RGCL 12 h after injection, as shown by TUNEL-positive cell counts. These inhibitors also protected the inner retina as shown by morphometric studies such as cell counts in the RGCL and measurement of the IPL thickness 7 days after injection. On the other hand, an ERK inhibitor (2.0 nmol U0126) did not suppress NMDA-induced cell death in the RGCL nor thinning of the IPL. These findings suggest that JNK and p38 are proapoptotic in NMDA-induced cell death in the RGCL, but not ERK.     

Protective effects of betaxolol in eyes with kainic acid-induced neuronal death

In the present study, we investigated whether betaxolol, a selective beta1-adrenoceptor antagonist, has neuroprotective effect on kainic acid (KA)-induced retinal damage. Neurotoxicities were induced in adult male rats by intravitreal injection of KA (total amount, 6 nmol). To examine the neuroprotective effects of betaxolol, rats were pretreated with betaxolol topically 60 min before KA injection to the rat eyes and twice daily for 1, 3, and 7 days after KA injection. The neuroprotective effects of betaxolol were estimated by measuring the thickness of the various retinal layers, and by counting the number of choline acetyltransferase (ChAT)- and tyrosine hydroxylase (TH)-positive cells in each retinal layer. The retina is highly vulnerable to KA-induced neuronal damage. Morphometric analysis of retinal damage in KA injected eyes, the thickness of the retinal layers decreased markedly after KA injection period of both 3 and 7 days. Furthermore, the numbers of ChAT- and TH-positive cells were significantly reduced by intravitreal injection of KA. However, when two drops of betaxolol, once before KA injection and twice daily for 7 days after KA injection, were continuously administered, the reductions in the retinal thickness and the retinal ChAT- and TH-positive cells were significantly attenuated. The present study suggests that topically applied betaxolol has neuroprotective effect on the retinal cell damage due to KA-induced neurotoxicity.     

Spontaneous synchronized calcium oscillations in neocortical neurons in the presence of physiological [Mg(2+)]: involvement of AMPA/kainate and metabotropic glutamate receptors

Primary cultures of neocortical neurons exhibit spontaneous Ca(2+) oscillations under zero or low extracellular [Mg(2+)] conditions. We find that mature murine neocortical neurons cultured for 9 days also produce spontaneous Ca(2+) oscillations in the presence of physiological [Mg(2+)]. These Ca(2+) oscillations were action potential mediated inasmuch as tetrodotoxin eliminated their occurrence. AMPA receptors were found to regulate the frequency of Ca(2+) oscillations. In contrast, Ca(2+) oscillations were independent of activation of L-type Ca(2+) channels, and NMDA receptors provided only a minor contribution. Release of intracellular Ca(2+) stores was involved in the oscillatory activity since thapsigargin reduced the amplitude and frequency of the oscillations. S-4-carboxyphenylglycine (S)-4CPG), an antagonist of group I metabotropic glutamate receptor (mGluR), also reduced the amplitude of oscillations. In addition, 1-aminocyclopentane-trans-1,3-dicarboxylic acid (trans-ACPD), a group I mGluR agonist, increased the oscillation frequency, suggesting a critical role for mGluR in the generation of Ca(2+) oscillations. The mGluR-mediated release of intracellular Ca(2+) stores appeared to be mediated by phospholipase C (PLC) since the PLC inhibitor U73122 eliminated the Ca(2+) oscillations. These results indicate that Ca(2+) oscillations in neocortical cultures in the presence of physiologic [Mg(2+)] are primarily initiated by excitatory input from AMPA receptors and involve mobilization of intracellular Ca(2+) stores following activation of mGluR.     

AMPA receptor stimulation mediates the antidepressant-like effect of a group II metabotropic glutamate receptor antagonist

(1R,2R,3R,5R,6R)-2-amino-3-(3,4-dichlorobenzyloxy)-6-fluorobicyclo[3.1.0]hexane-2,6-dicarboxylic acid (MGS0039), a selective group II metabotropic glutamate receptor (mGluR) antagonist, exhibits antidepressant-like activities in rodent models. In the present studies, to clarify the involvement of alpha-amino-3-hydroxy-5-methylisoxazole-4-propionate (AMPA) receptor activation in exhibition of the antidepressant-like properties of MGS0039, we examined the effect of an AMPA receptor antagonist, 2,3-dihydroxy-6-nitro-7-sulfamoylbenzo(f)quinoxaline (NBQX), on the antidepressant-like effect of MGS0039 in the mouse tail suspension test. We also examined the effects of NBQX on increased serotonin release after treatment with MGS0039 in the rat medial prefrontal cortex (mPFC) using in vivo microdialysis evaluation. In the tail suspension test, MGS0039 (0.3-3 mg/kg, i.p.) treatment dose-dependently and significantly reduced immobility time. Pretreatment with NBQX (10 mg/kg, s.c.) significantly prevented the antidepressant-like effect of MGS0039 in the tail suspension test, while NBQX itself had no effect on immobility time. In the microdialysis evaluation, administration of MGS0039 (10 mg/kg, i.p.) significantly increased serotonin levels in mPFC in freely moving rats, while NBQX (1 mg/kg, i.p.) itself had no effect on serotonin release in this region. Pretreatment with NBQX significantly attenuated the increase in serotonin release by MGS0039. These findings suggest that stimulation of postsynaptic AMPA receptors plays a role in mediating the pharmacological effects of MGS0039.     

Thyrotropin-releasing hormone (protirelin) inhibits potassium-stimulated glutamate and aspartate release from hippocampal slices in vitro

Excess excitatory amino acid release is involved in pathways associated with seizures and neurodegeneration. Thyrotropin-releasing hormone (TRH; protirelin), a brain-derived tripeptide, has shown efficacy in the treatment of such disorders, yet its mechanism of neuroprotection is poorly understood. Using superfused hippocampal slices, we tested the hypothesis that TRH could inhibit evoked glutamate/aspartate release in vitro. Rat hippocampal slices were first equilibrated in oxygenated Krebs buffer (KRB) (120 min) then superfused for 10 min with KRB (control), or KRB containing 0.1, 1, or 10 microM TRH respectively, prior to and during 5 min depolarization with high potassium KRB (50 mM [K(+)] +/- TRH). Fractions (1 min) were collected during the 5 min stimulation and for an additional 10 min thereafter and analyzed for glutamate and aspartate by HPLC. TRH had no effect on baseline glutamate/aspartate release, while all three TRH doses significantly (P < 0.05) inhibited peak 50 mM [K(+)]-stimulated glutamate/aspartate release, and glutamate remained below control (P < 0.05) at 15 min post stimulation. A 5 min pulse of TRH (10 microM) had no affect on basal glutamate/aspartate release, whereas the TRH pre-pulsed slices failed to release glutamate/aspartate by [K(+)]-stimulation given 15 min later. These results are the first to show a potent and prolonged inhibitory effect of TRH on evoked glutamate/aspartate release in vitro. These initial studies suggest that exogenous and/or endogenous TRH may function, in part, to modulate excess glutamate release in specific CNS loci. Additional studies are in progress to fully understand the mechanism of this potent effect of TRH and its implication in various CNS disorders.     

Lack of the alanine-serine-cysteine transporter 1 causes tremors, seizures, and early postnatal death in mice

The Na(+)-independent alanine-serine-cysteine transporter 1 (Asc-1) is exclusively expressed in neuronal structures throughout the central nervous system (CNS). Asc-1 transports small neutral amino acids with high affinity especially for D-serine and glycine (K(i): 8-12 microM), two endogenous glutamate co-agonists that activate N-methyl-D-aspartate (NMDA) receptors through interacting with the strychnine-insensitive glycine binding-site. By regulating D-serine (and possibly glycine) levels in the synaptic cleft, Asc-1 may play an important role in controlling neuronal excitability. We generated asc-1 gene knockout (asc-1(-/-)) mice to test this hypothesis. Behavioral phenotyping combined with electroencephalogram (EEG) recordings revealed that asc-1(-/-) mice developed tremors, ataxia, and seizures that resulted in early postnatal death. Both tremors and seizures were reduced by the NMDA receptor antagonist MK-801. Extracellular recordings from asc-1(-/-) brain slices indicated that the spontaneous seizure activity did not originate in the hippocampus, although, in this region, a relative increase in evoked synaptic responses was observed under nominal Mg(2+)-free conditions. Taken together with the known neurochemistry and neuronal distribution of the Asc-1 transporter, these results indicate that the mechanism underlying the behavioral hyperexcitability in mutant mice is likely due to overactivation of NMDA receptors, presumably resulting from elevated extracellular D-serine. Our study provides the first evidence to support the notion that Asc-1 transporter plays a critical role in regulating neuronal excitability, and indicate that the transporter is vital for normal CNS function and essential to postnatal survival of mice.     

Effect of two noncompetitive AMPA receptor antagonists GYKI 52466 and GYKI 53405 on vigilance, behavior and spike-wave discharges in a genetic rat model of absence epilepsy

The present study was conducted to investigate the effects of two noncompetitive alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor antagonists, GYKI 52466 and GYKI 53405 (the racemate of talampanel) on the generation of spike-wave discharges (SWD) parallel with the vigilance and behavioral changes in the genetic absence epilepsy model of WAG/Rij rats. Intraperitoneal (i.p.) administration of GYKI 52466 (1-[4-aminophenyl]-4-methyl-7,8-methylenedioxy-5H-2,3-benzodiazepine; 3, 10 and 30 mg/kg, i.p.), the prototypic compound of the 2,3-benzodiazepine family, caused a fast dose-dependent increase in the number and cumulative duration of SWD. These changes were accompanied by dose-dependent increase in duration of light slow wave sleep (SWS1) and passive awake, vigilance states associated with the presence of SWD. In addition a short, transient behavioral activation occurred that was followed by strong ataxia and immobility, decrease of active wakefulness and increase in deep slow wave sleep. GYKI 53405 (7-acetyl-5-(4-aminophenyl)-8-methyl-8,9-dihydro-7H-1,3-dioxolo[4,5-b][2,3]benzodiazepine, the racemate of talampanel, 16 mg/kg, i.p.) failed to affect any measure of SWD and vigilance. When used as a pretreatment, GYKI 52466 (10 mg/kg) slightly attenuated SWD-promoting effects of the 5-HT1A receptor agonist 8-OH-DPAT, it decreased cumulative duration and average time of paroxysms. In conclusion, AMPA receptors play moderate role in regulation of epileptic activity, and some of these effects are connected to their effects on vigilance in this model.     

Effect of delta-aminolevulinic acid treatment on N-methyl-D-aspartate receptor at different ages in the rat brain

We report here the effects of the chronic treatment with the oxidant agent delta-aminolevulinic acid (ALA) on the N-methyl-D-aspartate (NMDA) receptors in 4-, 12- and 24-month-old male Wistar rats. ALA was administered daily for 15 days (40 mg/kg i.p). The study was performed by membrane homogenate binding and autoradiography, using tritiated 5-methyl-10, 11-dihydro-5H-dibenzo(a,d)cycloheptan-5,10-imine maleate ([3H]MK-801). [3H]MK-801 binding was significantly decreased in most areas studied (cortex and hippocampus) at all ages in treated rats with respect to their controls. Furthermore, Western blot assays were performed using antibodies against the NMDA receptor NR2A subunit, which is widely distributed in the brain, mainly in cortex and hippocampus. In cortex but not in hippocampus, the ALA treatment induced significant decreases in the amounts of NR2A subunit in 12- and 24-month-old animals. We conclude that chronic treatment with ALA is able to induce NMDA receptor decreases in an age-independent way and that NR2A subunit seems to be involved in these decreases in cerebral cortex, but not in the other structures studied.     

Nefopam blocks voltage-sensitive sodium channels and modulates glutamatergic transmission in rodents

In order to specify the nature of interactions between the analgesic compound nefopam and the glutamatergic system, we examined the effects of nefopam on binding of specific ligands on the three main subtypes ionotropic glutamate receptors: N-methyl-D-aspartate (NMDA), alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA), or quisqualic acid (QA) and kainic acid (KA) in rat brain membrane preparations. Functionally, we investigated the effects of nefopam against the seizures induced by agonists of these excitatory glutamate receptors in mice. Since the synaptic release of glutamate mainly depends upon the activation of membrane voltage-sensitive sodium channels (VSSCs), the nature of interactions between nefopam and these ionic channels was studied by evaluating the effects of nefopam on binding of 3H-batrachotoxinin, a specific ligand of the VSSCs in rat brain membrane preparations. The functional counterpart of the binding of nefopam on VSSCs was evaluated by its effects on the 22Na uptake-stimulated by veratridine on human neuroblastoma cells and in the maximal electroshock test in mice. Nefopam showed no affinity for the subtypes of ionotropic glutamate receptors up to 100 microM. On the other hand, nefopam was effective against NMDA, QA and KA induced clonic seizures in mice. Nefopam displaced 3H-batrachotoxinin and inhibited the uptake of 22Na in the micromolar range and it protected mice against electroshock induced seizures. Nefopam may block the VSSCs activity: consequently, at the presynaptic level, this effect led to a reduction of glutamate release and at the postsynaptic level, it led to a decrease of the neuronal excitability following activation of the glutamate receptors.     

Tyrosine administration does not affect desipramine-induced dopamine levels as measured in vivo in prefrontal cortex

Using in vivo microdialysis, we examined whether tyrosine administration would potentiate the desipramine (DMI)-induced elevation of medial prefrontal cortex (MPFC) dopamine (DA) levels. DMI (10 or 20 mg/kg IP) increased MPFC DA levels but not DOPA accumulation. Tyrosine (12.5-100 mug/ml) administered by reverse microdialysis did not affect DMI-induced MPFC DA levels. The data support our hypothesis that DA synthesis must be significantly increased in order for administered tyrosine to increase extracellular DA levels.     

VIP enhances synaptic transmission to hippocampal CA1 pyramidal cells through activation of both VPAC1 and VPAC2 receptors

We previously described that vasoactive intestinal peptide (VIP) increases synaptic transmission to hippocampal CA1 pyramidal cells at concentrations known to activate VIP-selective receptors (VPAC1 and VPAC2) but not the PACAP-selective PAC1 receptor. We now investigated the involvement of VPAC1 and VPAC2 receptors in the effects elicited by VIP as well as the transduction pathways activated by VIP to cause enhancement of synaptic transmission. Blockade of either VPAC1 or VPAC2 receptors with PG 97-269 (100 nM) or PG 99-465 (100 nM) inhibited VIP-induced enhancement of synaptic transmission. Selective activation of VPAC1 receptors with [K15, R16, L27] VIP(1-7)/GRF(8-27) (10 nM) or of VPAC2 receptors with RO 25-1553 (10 nM) increased synaptic transmission to CA1 pyramidal cells, and this increase was larger when both agonists were applied together. Inhibition of either PKA with H-89 (1 microM) or PKC with GF109203X (1 microM) attenuated the effect of VIP (1 nM). GF109203X (1 microM) abolished the effect of the VPAC1 agonist [K15, R16, L27] VIP(1-7)/GRF(8-27) (10 nM) on hippocampal synaptic transmission but that effect was not changed by H-89 (1 microM). The effect of RO 25-1553 (100 nM) obtained in the presence of both the PAC1 and VPAC1 antagonists, M65 (30 nM) and PG 97-269 (100 nM), was strongly inhibited by H-89 (1 microM) but not GF109203X (1 microM). It is concluded that VIP enhances synaptic transmission to CA1 pyramidal cell dendrites through VPAC1 and VPAC2 receptor activation. VPAC1-mediated actions are dependent on PKC activity, and VPAC2-mediated actions are responsible for the PKA-dependent actions of VIP on CA1 hippocampal transmission.     

Theta bursts set up glutamatergic as well as GABA-ergic plasticity in neonatal rat hippocampal CA1 neurons

gamma-Aminobutyric acid (GABA) is inhibitory in adult, but excitatory in neonatal, neurons. The switch from excitatory to inhibitory action is due to a negative shift in the equilibrium potential for the GABA(A) receptor-mediated postsynaptic current (E(GABA-PSC)). Here, we report that, in neonatal rat hippocampal CA1 neurons, presynaptic theta-burst activation induces not only a shift in E(GABA-PSC) towards that in adult neurons, but also a recruitment of alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA)-receptor-mediated postsynaptic currents.     

Mice lacking D-amino acid oxidase activity display marked attenuation of stereotypy and ataxia induced by MK-801

The behavioral effects produced by MK-801 (0.4 mg/kg) were compared in mutant DAO-/- mice lacking D-amino acid oxidase activity and normal DAO+/+ mice. Mutant mice display marked diminution of stereotypy and ataxia induced by MK-801 compared to normal mice. Because the D-serine level in the brain of mutant mice is significantly higher than that of normal mice, the elevated D-serine in the brain of mutant mice could antagonize MK-801-induced stereotypy and ataxia.     

Oxytocin increases the density of high affinity alpha(2)-adrenoceptors within the hypothalamus, the amygdala and the nucleus of the solitary tract in ovariectomized rats

Oxytocin induces long-term changes in, for example, blood pressure, spontaneous motor activity and corticosterone levels in rats. Previous studies in male rats have suggested a role for alpha(2)-adrenoceptors within the central nervous system in these effects. The aim of the present study was to investigate if oxytocin treatment in female rats would influence alpha(2)-adrenoceptors within the hypothalamus, the amygdala and the nucleus of the solitary tract (NTS). For this purpose, female ovariectomized (OVX) rats were treated with oxytocin (1 mg/kg s.c.) or saline once a day for 10 days. Rats were decapitated 5 days after the last injection, and brains and plasma were collected. Quantitative receptor autoradiography for characterization of high affinity alpha(2)-adrenoceptor agonist binding and radioimmunoassay for corticosterone were performed. Oxytocin increased the B(max) values of the alpha(2)-adrenoceptor agonist [3H]UK14.304 binding sites significantly in all the analyzed areas (P<0.05). K(d) values were unchanged. Plasma levels of corticosterone were significantly decreased in the oxytocin-treated rats (P<0.05). These findings are in further support of an interaction between oxytocin receptors and alpha(2)-adrenoceptors and show that oxytocin treatment may increase alpha(2)-adrenoceptor recognition probably leading to an increase in alpha(2)-adrenoceptor signaling in several parts of the brain.     

Role of NMDA receptor subtypes in the induction of catalepsy and increase in Fos protein expression after administration of haloperidol

The increase of Fos expression in the striatum induced by haloperidol, an antagonist of the dopamine D2 receptor, might be related to the activation of glutamatergic neurotransmission, especially that of N-methyl-D-aspartate (NMDA) receptors. In this study, using behavioral and immunohistochemical techniques, we examined the effects of a noncompetitive NMDA antagonist, (+)-MK-801, and an NMDA receptor NR2B subunit antagonist, ifenprodil, on catalepsy, an extrapyramidal symptom; in this context, we also considered the expression of Fos protein in the forebrain after the administration of haloperidol. Catalepsy in mice, induced by the administration of haloperidol (1 mg/kg), was inhibited by pretreatment with (+)-MK-801 (0.2 mg/kg) or ifenprodil (10 mg/kg). Furthermore, pretreatment with (+)-MK-801 (0.2 mg/kg) significantly attenuated the induction of Fos-immunoreactive (IR) cells in the dorsomedial, dorsolateral, and ventrolateral striatum, but not in the shell region of the nucleus accumbens after the administration of haloperidol, whereas pretreatment with ifenprodil (10 mg/kg) significantly attenuated the induction of Fos-IR cells in all of these areas. It is known that ifenprodil binds sigma receptors and alpha-1 adrenergic receptors with high affinity. Pretreatment with the sigma receptor antagonist BD-1407 (3 mg/kg) or the alpha-1 adrenergic receptor antagonist prazosin (3 mg/kg) affected neither catalepsy nor the expression of Fos-IR cells after the administration of haloperidol. However, pretreatment with CP-101,606 (1 mg/kg), a selective antagonist for the NR2B subunit of the NMDA receptor, significantly attenuated catalepsy and the expression of Fos-IR cells in the forebrain after the administration of haloperidol. These results suggest that the NMDA receptor antagonists attenuated the induction of catalepsy and Fos-IR cells in forebrain after the administration of haloperidol. It was also suggested that haloperidol-induced expression of Fos-IR cells in the shell region of the nucleus accumbens might be differentially regulated by NMDA receptor subunits. Therefore, it appears that selective antagonists for the NR2B subunit of the NMDA receptor (e.g., CP-101,606) might be useful drugs for the treatment of extrapyramidal side effects (EPS) associated with the chronic use of typical antipsychotics such as haloperidol.     

Nicotinic-receptor potentiator drugs, huprine X and galantamine, increase ACh release by blocking AChE activity but not acting on nicotinic receptors

The main goal of the present study was to analyse the effects of (+/-)-huprine X ((+/-)-HX) and galantamine (GAL), with potentiating action on nicotinic receptors, and huperzine A (HPA), devoid of nicotinic activity, on [3H]-acetylcholine ([3H]-ACh) release in striatal slices of rat brain. All compounds are non-covalent and reversible inhibitors of AChE. Addition of (+/-)-HX (0.01 microM), GAL (10 microM) and HPA (0.1 microM) to the superfusion medium decreased the release of the ACh neurotransmitter to a similar extent: 36%, 30% and 34%, respectively (P<0.01). This effect was reverted in the presence of atropine (ATR; 0.1 microM), which blocks the pre-synaptic muscarinic M2 receptor. After that, a wide range of concentrations of drugs, concomitantly with ATR (0.1 microM), was studied in the presence of haloperidol (HAL; 0.01 microM), a dopamine D2 antagonist. In these conditions, a dose-dependent increase of [3H]-ACh release was observed in the presence of (+/-)-HX, GAL and HPA. To test the role of nicotinic receptors in the drugs' effects on [3H]-ACh release, mecamylamine (MEC) 100 microM was used to block such receptors. MEC alone significantly decreased neurotransmitter release by 18% (P<0.05), but no change was obtained in the presence of both ATR and MEC. Under these conditions, (+/-)-HX, GAL and HPA increased the release of [3H]-ACh by 37%, 25% and 38%, respectively (P<0.01). Taking into account all of these data, the present results suggest that the effects induced by (+/-)-HX and GAL nicotinic-receptor potentiators seem to be mainly due to their ability in inhibiting acetylcholinesterase activity, but not by interaction on the nicotinic receptors.     

Low intracerebroventricular doses of cholinotoxin AF64A do not affect the morphology of gonadotropin hormone-releasing hormone (GnRH)-immunoreactive fibers in the rat septum

Ethylcholine aziridinium (AF64A) induces cholinergic lesion in animal models of AD. Although higher concentrations of AF64A are known to induce nonspecific, cholinergic, and non-cholinergic lesions, low concentrations are believed to be selectively cholinotoxic. However, morphological evidence of this phenomenon has not been demonstrated yet. The present study demonstrates that while AF64A damaged septal cholinergic fibers, periventricular GnRH-immunoreactive fibers remained intact, confirming the highly selective cholinotoxicity of AF64A at appropriate concentrations.