Inhibition of nitric oxide synthase prevents magnesium-free-induced epileptiform activity in guinea-pig piriform cortex neurones in vitro

The effects of N-nitro-L-arginine methyl ester (L-NAME), a nitric oxide (NO) synthase inhibitor, were examined on Mg²⁺-free-induced epileptiform activity, in guinea-pig piriform cortex slices in vitro. L-NAME (0.1-1mM) had no effect on neuronal membrane properties or electrically-evoked postsynaptic potentials (PSPs). In contrast, during superfusion of the slices with Mg²⁺-free solution neurones exhibited spontaneous and stimulus-evoked epileptiform potentials that were suppressed in the presence of L-NAME (100 μΜ) or the selective NMDA receptor antagonist DL-APV (100 μΜ). The inhibitory effects induced by L-NAME were reversibly reduced by L-arginine (1mM), but not D-arginine (1mM), the latter drug not being a substrate for NO formation. It was concluded that L-NAME can suppress epileptiform activity induced by Mg2⁺-free exposure primarily through a decrease in presynaptic transmitter release, although additional actions on the NMDA-receptor complex were also considered. Received: 24 October 1996 / Accepted: 7 January 1997     

Interaction between P2Y and NMDA receptors in layer V pyramidal neurons of the rat prefrontal cortex

In the first part of this study, monosynaptic excitatory postsynaptic potentials (EPSPs) in layer V of the rat prefrontal cortex were evoked by electrical stimulation of layer I. Recordings by intracellular sharp microelectrodes showed that EPSPs were concentration-dependently facilitated by the P2 receptor antagonistic ATP analogue 2-methylthio ATP (2-MeSATP), while ATP itself depressed the synaptic potentials. The inhibitory effect of ATP turned into facilitation in the presence of the adenosine A(1) receptor antagonist DPCPX. The 2-MeSATP-induced potentiation of EPSP amplitudes were prevented by the P2 receptor antagonists PPADS and Suramin. The EPSP was almost abolished by coapplication of the NMDA receptor antagonist AP-5 and the AMPA/kainate receptor antagonist CNQX. After blockade of the NMDA receptor-mediated part of the EPSP by AP-5, the stimulatory effect of 2-MeSATP disappeared. When NMDA or AMPA were pressure-applied onto pyramidal cells, only the NMDA-induced depolarization was potentiated by 2-MeSATP. In the second part of the study, NMDA-induced currents were measured by whole-cell patch-clamp pipettes. ATP, 2-MeSATP, UDP and UTP potentiated the response to NMDA, while ADP-beta-S was inactive. PPADS antagonized the effect of ATP. Synaptic isolation of pyramidal neurons by a Ca(2+)-free medium or tetrodotoxin did not alter the effect of ATP which, however, was markedly depressed when GTP in the micropipette was replaced by GDP-beta-S. These observations suggest that in layer V pyramidal neurons of the prefrontal cortex postsynaptically localized P2Y receptors interact with NMDA receptor-channels.     

Mechanisms of bi-directional modulation of thalamocortical transmission in barrel cortex by presynaptic kainate receptors

Presynaptic kainate receptors play an important role in synaptic transmission and short-term plasticity to profoundly regulate network activity in many parts of the mammalian brain. In primary sensory neocortex, where short-term synaptic plasticity is important for receptive field structure and information processing, kainate receptors are highly expressed and regulate thalamocortical inputs, particularly during development. However, the mechanisms of the kainate receptor-dependent presynaptic regulation of thalamocortical transmission are unclear. We therefore investigated this issue using electrophysiology in neonatal thalamocortical slices of barrel cortex combined with pharmacology and biochemical analyses. We show that presynaptic kainate receptors can both facilitate or depress synaptic transmission depending on the extent of their activation. This bi-directional regulation is mediated in part by kainate receptors that directly influence thalamocortical axonal excitability, but also likely involves receptors acting at thalamocortical terminals to regulate transmitter release. The efficacy of kainate in regulating thalamocortical transmission is low compared to that reported for other inputs. Consistent with this low efficacy, our biochemical analyses indicate that the presynaptic kainate receptors regulating neonatal thalamocortical inputs likely lack the high kainate affinity GluK4 and 5 subunits. Thus thalamocortical transmission can be bi-directionally regulated by low affinity kainate receptors through two mechanisms. Such presynaptic regulation provides a potentially powerful mechanism to influence sensory processing during development of barrel cortex.     

In vivo methylmercury exposure induced long-lasting epileptiform activity in layer II/III neurons in cortical slices from the rat

Prenatal and postnatal methylmercury (MeHg) exposure has been shown to increase neuronal excitability and seizure susceptibility. To determine if early postnatal MeHg exposure causes a similar effect, we examined changes in field potentials in layer II/III neurons in cortical slices of rat following in vivo MeHg treatment. Rats received 0 (0.9% NaCl), 0.75 mg/kg/day or 1.5 mg/kg/day MeHg subcutaneously for 15 or 30 days beginning on postnatal day 5, after which cortical slices were prepared for field potential recordings. In slices from rats treated with vehicle, single pulse stimulation of layer IV of cortical slices induced a typical field excitatory postsynaptic potential (fEPSP) with a single spike. This type of fEPSPs was also seen in slices from rats with 15 day treatment with 0.75 mg/kg/day or 1.5 mg/kg/day MeHg. However, 30-day treatment with either MeHg dose resulted in fEPSPs with multiple spikes (epileptiform activity) in 40% of animals examined. This epileptiform activity remained observable in 50–60% animals in which MeHg exposure had been terminated for 30 days. However, slices from control animals still showed fEPSPs with single spike. Thus, these data suggest that postnatal MeHg exposure in vivo altered neuronal excitability and induced a long-lasting hyperexcitability in cortical neurons.     

Positive coupling of cholinergic muscarinic receptors to adenylate cyclase activity in membranes of rat olfactory bulb

Summary In membranes of rat olfactory bulb acetylcholine stimulated adenylate cyclase activity in a concentration-dependent manner. The maximal stimulation corresponded to 53% increase of basal enzyme activity and was obtained with 100 M acetylcholine. The concentration of the cholinergic agonist eliciting a half-maximal effect was 0.4 M. The stimulatory effect of acetylcholine was antagonized by 0.1 M atropine but not by 10 M (+)-tubocurarine. Moreover, the addition of micromolar concentrations of GTP was absolutely required for the enzyme stimulation by acetylcholine. The results demonstrate the presence in rat olfactory bulb of muscarinic receptors coupled to stimulation of adenylate cyclase probably via a GTP regulatory protein and provide evidence for a novel signal transduction mechanism of central muscarinic receptors. Send offprint requests to P. Onali at the above address     

Lesion of forebrain nuclei reveals possible presynaptic cholinergic muscarinic receptors in rat cerebral cortex

In rats, three days after unilateral lesion of magnocellular basal forebrain nuclei, binding of L-[3H]-quinuclidinyl benzilate, and acetylcholinesterase activity decreased significantly in the ipsilateral and, to a lesser extent, in the contralateral cerebral cortex. This result suggests the existence of presynaptic muscarinic receptors in the cortical projections of these nuclei. After 14 days, the binding increased on both sides while the level of acetylcholinesterase activity remained low. These findings suggest that deafferentation causes ultimately an increase in postsynaptic receptors.     

Potentiation of sympathetic neuromuscular transmission mediated by muscarinic receptors in guinea pig isolated vas deferens

In guinea-pig isolated vasa deferentia, purinergic neurogenic contractions and responses to applied adenosine 5-triphosphate (ATP) were potentiated by carbachol; responses to adrenergic transmission and applied noradrenaline were not. Following blockade of P2 receptors and -adrenoceptors, the residual neurogenic response was massively potentiated by carbachol, suggesting the presence of a non-purinergic, non-adrenergic component. In the presence of guanethidine, carbachol had no significant effect, indicating that sympathetic transmission was the only element involved. Use of oxotremorine and selective muscarinic receptor antagonists suggested that the potentiating effect of carbachol and oxotremorine was mediated via M3 muscarinic receptors without involvement of nicotinic receptors.     

Effects of estrogen on intracellular signaling pathways linked to activation of muscarinic acetylcholine receptors and on acetylcholinesterase activity in rat hippocampus.

The aim of the present study was to investigate the effects of estrogen lack and estrogen replacement on the production of total [3H]inositol phosphate ([3H]IP) induced by the activation of muscarinic acetylcholine receptors (mAChRs) and on the mechanisms for inactivation of acetylcholine. Hippocampi were obtained from rats in proestrus (PE), ovariectomized for 15 days (C15), ovariectomized for 15 days and then treated with 17beta-estradiol for 7 days (E7) and ovariectomized and immediately treated with 17beta-estradiol for 21 days (E21). Ovariectomy did not change the basal level of total [3H]IP in the hippocampus. 17beta-Estradiol replacement (E7 and E21) reduced the basal level of total [3H]IP. In all experimental groups, carbachol (CCh) caused a concentration-dependent rise in total [3H]IP. The maximum effect was reached with 10(-4) M CCh. The response to 10(-4) M CCh in the hippocampi from C15 and E7 rats was twofold higher than in hippocampi from PE and E21 animals and was blocked by pirenzepine, but not by methoctramine. Ovariectomy or 17beta-estradiol treatment for 7 days did not change neither the total acetylcholinesterase (AChE) activity nor the relative amount of mono- and dimeric G1/G2 and tetrameric G4 globular forms. Conversely, hormonal treatment for 21 days induced an increase in AChE activity of G1/G2 and G4 forms, indicating that 17beta-estradiol stimulates both synthesis and assembly of AChE molecular forms. The present results suggest that the duration and/or a critical period with regard to the initiation of estrogen therapy are important to regulate the function of mAChRs and AChE activity in female rat hippocampus.     

Effect of chronic ethanol treatment in vivo on excitability in mouse cortical neurones in vitro

1. The effects of cessation of chronic ethanol ingestion on seizure activity in vivo and on the characteristics of the evoked synaptic potentials in cortical neurones in vitro have been investigated in mice. Withdrawal from chronic ethanol treatment increased handling seizure ratings in mice between 4 and 16 h post-withdrawal. This ethanol-induced increase in seizure rating was unaffected by carbamazepine (30 mg kg⁻¹) but significantly reduced at a higher concentration (130 mg kg⁻¹).
2. Intracellular recordings were made from cortical layer II neurones in vitro from control mice and from mice following chronic ethanol ingestion. Evoked synaptic potentials were generated in these neurones through intralaminar stimulation.
3. Neurones from control mice displayed an evoked potential consisting of a fast excitatory postsynaptic potential (e.p.s.p.) mediated by AMPA-type glutamate receptors and an inhibitory postsynaptic potential (i.p.s.p.) mediated via GABA_A receptors. Application of pentylenetetrazole (PTZ) or bicuculline onto these neurones inhibited the i.p.s.p., caused a large increase in both the amplitude and duration of the e.p.s.p. and initiated spontaneous excitatory activity. The resulting large evoked e.p.s.p. was mediated via both NMDA- and AMPA-type glutamate receptors.
4. Most neurones (77%) from ethanol treated mice displayed an evoked potential which comprised a large e.p.s.p. and no i.p.s.p. The e.p.s.p. consisted of several distinct components and in addition these neurones displayed spontaneous paroxysmal depolarizing shifts. This multi-component e.p.s.p. was mediated through both NMDA- and AMPA-type glutamate receptors. A population (23%) of neurones from ethanol treated mice exhibited evoked potentials which possessed both inhibitory and excitatory components and these neurones were effectively identical to those obtained from control mice.
5. Carbamazepine reduced the duration of the e.p.s.p. in neurones from ethanol treated mice and in PTZ-treated control neurones.
6. Prolonged ethanol ingestion is known to create a neurochemical imbalance in cortical neurones resulting in abnormal neurotransmission. The present study highlights the functional consequences that arise as a result of these neurochemical changes leading to over-excitation of neurones and pronounced epileptiform activity.

     

Synergistic role of muscarinic acetylcholine and tachykinin NK-2 receptors in intestinal peristalsis

It is known that tachykinins (substance P, neurokinin A) participate in the excitatory neural pathways subserving peristaltic motor activity in the intestine. The aim of the present study was to elucidate the types of tachykinin receptor (NK-1 or NK-2) involved in peristalsis by the use of receptor subtype-selective antagonists. Peristaltic motility in isolated segments of the guinea-pig ileum was induced by pumping fluid into the oral end of the intestinal segment. By way of the intraluminal pressure the compliance of the intestinal wall during the preparatory phase and the pressure threshold to trigger the emptying phase of peristalsis were recorded. The tachykinin antagonists were used at concentrations that were at least 30 times in excess of the equilibrium dissociation constants which had previously been evaluated with receptor subtype-selective agonists on the guineapig ileum circular muscle. The NK-1 selective antagonist CP-96,345 (0.3 M) had a slight stimulant influence on peristalsis, whereas the NK-2 selective antagonists MEN-10,376 (10 M), GR-94,800 (0.3 M) and SR-48,968 (0.1 M) led to a small inhibition of motor activity. However, when given after exposure of the ileum to a threshold concentration of atropine (5–20 nM) causing little depression of peristalsis, the tachykinin NK-2 receptor antagonists invariably abolished peristalsis. This synergistic interaction was not seen when SR-48,968 was administered after the ileal segments had been exposed to concentrations of hexamethonium, isoproterenol or calcitonin gene-related peptide that by themselves caused a slight inhibition of peristalsis only. CP-96,345 was without effect on peristalsis when it was applied in the presence of a threshold concentration of atropine. These findings indicate that transmission via tachykinin NK-2, but not NK-1, receptors synergizes with cholinergic transmission via muscarinic receptors in the relay of excitatory enteric pathways subserving intestinal peristalsis. Correspondence to: P. Holzer at the above address     

Blockade of NMDA receptors in the prefrontal cortex increases dopamine and acetylcholine release in the nucleus accumbens and motor activity

Objectives  The present study investigates the effects of injections of a specific N-methyl-d-aspartic acid (NMDA) antagonist 3-[(R)-2-carboxypiperazin-4-yl]-propyl-1-phophonic acid (CPP) into the prefrontal cortex (PFC) on the extracellular concentrations of dopamine and acetylcholine in the nucleus accumbens (NAc) and on motor activity in the freely moving rat. Materials and methods  Sprague–Dawley male rats were implanted with guide cannulas into the medial PFC and NAc to perform bilateral microinjections and microdialysis experiments. Spontaneous motor activity was monitored in the open field. Results  Injections of CPP (1 μg/0.5 μL) into the PFC produced a significant increase of the baseline extracellular concentrations of dopamine (up to 130%), dihydroxyphenylacetic acid (DOPAC; up to 120%), homovanillic acid (HVA; up to 130%), and acetylcholine (up to 190%) in the NAc as well as motor hyperactivity. In the NAc, perfusion of the NMDA and alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) glutamate antagonists CPP (50 μM)+6,7-dinitroquinoxaline-2,3-dione (DNQX; 50 μM) through the microdialysis probe blocked acetylcholine release, but not DOPAC and HVA increases produced by CPP injections into the PFC. Also, increases in motor activity produced by prefrontal injections of CPP were significantly reduced by bilateral injections into the NAc of a mixed D1/D2 antagonist, flupenthixol (5 and 25 μg/0.5 μL). Injections into the NAc of the muscarinic antagonist scopolamine (1 and 10 μg/0.5 μL) further increased, and of the nicotinic antagonist mecamylamine (1 and 10 μg/0.5 μL) did not change, the increases in motor activity produced by prefrontal CPP injections. Conclusions  These results suggest that the dysfunction of NMDA receptors in the PFC could be a key factor in the neurochemical and motor effects associated with corticolimbic hyperactivity.     

Stereoselective effects of the novel anticonvulsant lacosamide against 4-AP induced epileptiform activity in rat visual cortex in vitro

We examined effects of the novel anticonvulsant lacosamide and its inactive isomer (SPM 6953) in an in vitro model of epileptiform activity. Focal field potential recordings (34+/-0.2 degrees C) were obtained from 17 to 22 day old rat brain slices. Physiological synaptic transmission (fEPSP amplitude and duration) in CA1 of rat hippocampus was not significantly altered (P > 0.05, n = 4) by lacosamide (1 microM-1 mM). Recording from visual cortex during application of 4-aminopyridine (4-AP; 100 microM) revealed both spontaneous and evoked 'ictal like' discharges. Spontaneous ictal like discharges in the visual cortex were blocked by 100 microM carbamazepine (CBZ), 100 microM pentobarbital and 200 microM phenobarbital (PHB) but were insensitive to the anti-absence drug ethosuximide (750 microM; n = 4, P > 0.05). Lacosamide reduced tonic duration and maximal firing frequency with EC(50)s of 41 and 71 microM, respectively. In contrast, the S stereoisomer (100-320 microM) produced no significant effect on spontaneous ictal activity (n = 3-4, P > 0.05). Seizures induced by high frequency (100 Hz, 1s) stimulation were selectively reduced in amplitude by PHB (200 microM) and frequency by CBZ (100 microM; n = 6) and lacosamide (100 microM; n = 4). GABAergic negative going potentials were attenuated by CBZ (irreversible with washing) and lacosamide (reversible) but not by PHB. We conclude that lacosamide blocks 4-AP induced epileptiform activity in the visual cortex. This novel anticonvulsant drug appears to inhibit epileptogenesis (seizure spread) by interacting with a stereoselective, but as yet unidentified, target site in rodent neocortex in the mid-micromolar range.     

Activation of midbrain presumed dopaminergic neurones by muscarinic cholinergic receptors: an in vivo electrophysiological study in the rat

1. Extracellular single-unit recording and iontophoresis were used to examine the effects of different cholinoceptor agonists and antagonists on the firing rate and firing pattern of A9 and A10 presumed dopaminergic neurones in the anaesthetized rat.
2. Administration of low currents (1–5 nA) of the selective muscarinic agonists oxotremorine M (Oxo M) and muscarine and of the non-selective muscarinic/nicotinic agonist carbamylcholine (CCh) produced a dose-dependent increase in firing rate in most of the A9 and A10 presumed dopaminergic neurones tested. Oxo M-induced activation could be completely blocked by iontophoretic application of the muscarinic antagonist butyl-scopolamine or systemic administration of the muscarinic antagonist scopolamine (300 μg kg⁻¹, i.v.).
3. Iontophoretic application of the selective nicotinic agonist methylcarbamylcholine (MCCh), but not nicotine, induced a consistent increase in firing rate. Surprisingly, the excitatory effect of MCCh was significantly reduced by the selective muscarinic antagonist scopolamine (300 μg kg⁻¹, i.v.), but not by the selective nicotinic antagonist mecamylamine (2.2 mg kg⁻¹, i.v.). Mecamylamine (3 mg kg⁻¹, i.v.) was also ineffective in reducing the CCh-induced activation of presumed dopamine neurones, suggesting that both CCh and MCCh increased the activity of dopamine neurones via an interaction with muscarinic receptors.
4. Iontophoretic application of the endogenous agonist acetylcholine (ACh) had no or little effect on the firing activity of A10 presumed dopaminergic neurones. However, concomitant application of neostigmine, a potent cholinesterase inhibitor, with acetylcholine induced a substantial activation of these neurones. This activation consisted of two components; one, which was prevalent, was scopolamine (300 μg kg⁻¹, i.v.)-sensitive, and the other was mecamylamine (2 mg kg⁻¹, i.v.)-sensitive.
5. In addition to their effect on firing activity, Oxo M, muscarine and concomitant neostigmine/ACh caused a significant increase in burst firing of A10 neurones, but not of A9 neurones.
6. These data suggest that dopamine cells, both in the A9 and A10 regions, possess functional muscarinic receptors, the activation of which can increase their firing rate and, for A10 neurones, their amount of burst activity. These cholinoceptors would be able to influence the activity of the midbrain dopamine system greatly and may play a role in, and/or be a therapeutic target for, brain disorders in which dopamine is involved (e.g., Parkinson''s disease, drug addiction and schizophrenia).

     

Presynaptic inhibition of synaptic transmission in the rat hippocampus by activation of muscarinic receptors: involvement of presynaptic calcium influx

1. Modulation of presynaptic voltage-dependent calcium channels (VDCCs) by muscarinic receptors at the CA3–CA1 synapse of rat hippocampal slices was investigated by using the calcium indicator fura-2. Stimulation-evoked presynaptic calcium transients ([Ca_pre]_t) and field excitatory postsynaptic potentials (fe.p.s.ps) were simultaneously recorded. The relationship between presynaptic calcium influx and synaptic transmission was studied.
2. Activation of muscarinic receptors inhibited [Ca_pre]_t, thereby reducing synaptic transmission. Carbachol (CCh, 10 μM) inhibited [Ca_pre]_t by 35% and reduced fe.p.s.p. by 85%. The inhibition was completely antagonized by 1 μM atropine. An approximate 4^th power relationship was found between presynaptic calcium influx and postsynaptic responses.
3. Application of the N-type VDCC-blocking peptide toxin ω-conotoxin GVIA (ω-CTx GVIA, 1 μM) inhibited [Ca_pre]_t and fe.p.s.ps by 21% and 49%, respectively, while the P/Q-type VDCC blocker ω-agatoxin IVA (ω-Aga IVA, 1 μM) reduced [Ca_pre]_t and fe.p.s.ps by 35% and 85%, respectively.
4. Muscarinic receptor activation differentially inhibited distinct presynaptic VDCCs. ω-CTx GVIA-sensitive calcium channels were inhibited by muscarinic receptors, while ω-Aga IVA-sensitive channels were not. The percentage inhibition of ω-CTx GVIA-sensitive [Ca_pre]_t was about 63%.
5. Muscarinic receptors inhibited presynaptic VDCCs in a way similar to adenosine (Ad) receptors. The percentage inhibition of ω-CTx GVIA-sensitive [Ca_pre]_t by Ad (100 μM) was about 59%. There was no significant inhibition of ω-Aga IVA-sensitive channels by Ad. The inhibitions of [Ca_pre]_t by CCh and Ad were mutually occlusive.
6. These results indicate that inhibition of synaptic transmission by muscarinic receptors is mainly the consequence of a reduction of the [Ca_pre]_t due to inhibition of presynaptic VDCCs.

     

Blockade of glutamate transporters leads to potentiation of NMDA receptor current in layer V pyramidal neurons of the rat prefrontal cortex via group II metabotropic glutamate receptor activation

Membrane currents of layer V pyramidal cells in slices of the rat prefrontal cortex (PFC) were recorded with the patch-clamp technique. In an Mg(2+)-free superfusion medium l-trans-pyrrolidine-2,4-dicarboxylic acid (PDC), a preferential blocker of astrocytic glutamate transporters, caused inward current due to the activation of NMDA receptors. The blockade of conducted action potentials by tetrodotoxin did not interfere with this effect. ATP was inactive when given alone and potentiated the NMDA-induced current in an Mg(2+)-containing but not Mg(2+)-free superfusion medium. Agonists of group I ((S)-3,5-dihydroxyphenylglycine; DHPG) and II ((1R,4R,5S,6R)-4-amino-2-oxabicyclo[3.1.0]hexane-4,6-dicarboxylic acid; LY 379268) metabotropic glutamate receptors (mGluRs) also potentiated responses to NMDA, whereas the group III mGluR agonist l-(+)-2-amino-4-phosphonobutyric acid (l-AP4) did not affect them. In contrast to ATP, PDC evoked inward current in the absence but not in the presence of external Mg(2+), when given alone, and facilitated the NMDA effect Mg(2+)-independently. The PDC-induced facilitation of NMDA responses was blocked by group II ((2S)-2-amino-2-[(1S,2S)-2-carboxycycloprop-1-yl]-3-(xanth-9-yl) propanoic acid; LY 341495), but not group I ((RS)-1-aminoindan-1,5-dicarboxylic acid; AIDA) or III (alpha-methyl-3-methyl-4-phosphonophenylglycine; UBP 1112) mGluR antagonists. In conclusion, the blockade of astrocytic glutamate uptake by PDC may lead to a stimulation of group II mGluRs, while the triggering of exocytotic glutamate release from astrocytes by ATP may cause activation of group I mGluRs, both situated postsynaptically at layer V PFC pyramidal cells. Either group of mGluRs may interact with NMDA receptors in a positive manner.     

Serotonergic modulation of neurotransmission in the rat subicular cortex in vitro: a role for 5-HT1B receptors

Summary We have studied the effect of serotonin on synaptic transmission in rat hippocampal subiculum slices. Electrical stimulation of the alveus induced a field potential in the subiculum. The non-NMDA glutamate receptor antagonist, NBQX (3 × 10^–6 mol/l) suppressed the response by 78%, indicating that the signal involves glutamatergic neurons. Application of serotonin suppressed (EC₅₀ = 3.6 × 10^–6 mol/l) the amplitude of he evoked potentials in a reversible, concentration-dependent manner. The responses to 5-HT were not altered after pretreatment with the 5-HT uptake blocker, fluvoxamine (10^–5 mol/l) or a combination of the MAO inhibitor pargyline (10^–5 mol/l) and ascorbic acid (10^–4 mol/l). The responses to 5-HT were also unaffected by pretreatment with the 5-HT_1A selective antagonist NAN-190 (10^–6 mol/l), the 5-HT_2A antagonist ketanserin (10^–6 mol/l) or the 5-HT₃/5-HT₄ antagonist ICS 205–930 (10^–6 mol/l).The 5-HT_1B selective agonist CP 93,129 mimicked the effects of serotonin, but was more potent (EC₅₀ 4.1 × 10^–7 mol/l). The 5-HT_1B receptor antagonist, (±)21-009 (3 × 10^–7 mol/l), antagonized the response to 5-HT and CP 93,129 with a pK_B value of 7.1 and 7.2, respectively. These results suggest that the effect of 5-HT in the rat subiculum is mediated by 5-HT_1B receptors.Correspondence to: H.W.G.M. Boddeke at the above address     

Both agonists and antagonists of the strychnine-insensitive glycine site of N-methyl-D-aspartate receptors modulate polysynaptic excitations in slices of mouse olfactory cortex

Summary In this study, it is reported that bath application of D-serine and, to a lesser extent glycine, potentiated polysynaptic but not monosynaptic excitations evoked in slices of mouse olfactory cortex perfused with solution containing Mg²⁺ (1 mmol/l), picrotoxin and strychnine (both 25 μmol/l). Effects were largely confined to the longer latency components of the field potentials and occurred at amino acid concentrations of between 0.01 and 1 mmol/l. The effects of D-serine and glycine were antagonized by 7-chlorokynurenate and indole-2-carboxylate, antagonists of the glycine regulatory site of the N-methyl-D-aspartate (NMDA) receptor complex. D-Serine (glycine not tested) also potentiated, and 7-chlorokynurenate partially inhibited the longer latency components of the polysynaptic field potentials evoked in slices perfused in the absence of picrotoxin and strychnine. However, neither D-serine nor glycine potentiated responses evoked by the bath application of NMDA. It is concluded that under the present experimental conditions, the glycine regulatory sites of those NMDA receptor involved in the mediation of polysynaptic excitations in the mouse olfactory cortex are not saturated with endogenous glycine.     

In vivo modulation of histamine release by autoreceptors and muscarinic acetylcholine receptors in the rat anterior hypothalamus

The modulation of histamine release by histamine and muscarinic acetylcholine receptors was investigated by using the push-pull technique. The anterior hypothalamic area of the conscious, freely moving rat was superfused through the push-pull cannula with CSF or with CSF containing drugs and the release of endogenous histamine was determined in the superfusate.Hypothalamic superfusion with tetrodotoxin (10 mol/1) led to a pronounced and sustained decrease in the histamine release rate. Superfusion with compound 48/80 (100 mg/1) was ineffective. Hypothalamic superfusion with the H₃ agonist (R)--methylhistamine inhibited, while superfusion with the H₃ antagonist thioperamide enhanced the release of histamine. The release of histamine was inhibited on hypothalamic superfusion with the muscarinic receptor agonists carbachol or oxotremorine. Histamine release was enhanced by atropine, and this release-enhancing effect was abolished by oxotremorine. The selective M₁ antagonist pirenzepine (100 mol/I) and 4-diphenylacetoxy-N-methylpiperidine (4-DAMP, 10 ol/1), which blocks M₁ and M₃ receptors, also enhanced the release rate of histamine. On the other hand, 50 and 100 moI/I methoctramine (M₂ receptor antagonist) 10 and 100 moI/l p-fluoro-hexahydro-siladifenidol (p-F-HHSiD, a M₃ receptor antagonist) were ineffective.It is concluded that histamine released in the hypothalamus originates predominantly from neurons. The release of histamine is modulated by H₃ autoreceptors. The histamine release is also modulated by cholinergic neurons which modify histamine release from histaminergic neurons by stimulating M₁ muscarinic acetylcholine heteroreceptors probably located on histaminergic neurons.Supported by the Fonds zur Förderung der wissenschaftlichen Forschung Correspondence to: H. Prast at the above address     

Effects of agonists and antagonists of D1 and D2 dopamine receptors on self-stimulation of the medial prefrontal cortex in the rat

The possible participation of D1 versus D2 dopamine receptors in mediating dopaminergic neurotransmission of self-stimulation (SS) in the medial prefrontal cortex (MPC) of the rat was studied neuropharmacologically. Intracerebral as well as intraperitoneal injections of agonists and antagonists of dopamine receptors were used in this study. In all experiments performed with systemic injections, spontaneous motor activity (SM) was measured parallel to self-stimulation behavior as control for non specific effects of the drugs. Intracranial injections were done unilaterally serving SS of the contralateral side (not injected or injected with 0.9% NaCl) as control in the same animals. Spiroperidol and pimozide were used as D1-D2 dopamine antagonists, while sulpiride was used as a specific D2 antagonist. Apomorphine was used as D1-D2 agonist, while bromocriptine and lergotrile were used at doses in which these ergot drugs are considered predominantly D2 agonists. Sulpiride, intraperitoneally or intracerebrally injected at the same locus at which the stimulating electrode was located produced no effect on SS. On the contrary, the D1-D2 antagonists, spiroperidol and pimozide intraperitoneally or intracerebrally injected produced a dose-dependent decrease on SS. On the basis of these data it is suggested, that the dopamine neurotransmission involved in SS of the MPC is mediated via D1 dopamine receptors. This suggestion is further emphasized by the results obtained with the agonists, apomorphine, bromocriptine and lergotrile. Apomorphine produced a dose-related decrease on SS and a decrease at lower doses and an increase at higher doses on SM. Bromocriptine and lergotrile had, on the contrary, no effect on SS and a dose-related decrease on SM.     

Blockade of the locomotor stimulant effects of amphetamine by group I,group II,and group III metabotropic glutamate receptor ligands in the rat nucleus accumbens: possible interactions with dopamine receptors

Previous investigations have shown that mGlu receptors would be involved in the amphetamine-induced motor response. However, data are somewhat controversial across studies where methodological protocols vary. The aim of the present study was to determine the involvement of mGlu receptors in the NAcc in the locomotor-activating properties of amphetamine in rats well habituated to their experimental environment, a condition known to modulate the motor response to amphetamine. Focal infusion of the group I mGlu receptor antagonist S-4-CPG, which has no effect on basal motor activity, virtually suppressed the locomotor response to amphetamine, while infusion of the group II mGlu receptor antagonist LY 341495 or the group III mGlu receptor agonist AP4, at the minimal dose that produces locomotor activation, reduced it by approximately a half. These effects were blocked by the group I mGlu receptor agonist DHPG, the group II mGlu receptor agonist APDC, and the group III mGlu receptor antagonist MPPG, respectively. These data confirm that mGlu receptors in the NAcc contribute to the psychostimulant motor effect of amphetamine. Results are discussed from the view of recent neuropharmacological studies that have defined the effects of these mGlu receptor ligands on basal motor activity and DA receptor agonists-induced locomotor responses in rats exposed to similar experimental procedures (Eur J Neuroscience 13 (2001) 2157; Neuropharmacology 41 (2001) 454; Eur J Neuroscience 13 (2001) 869). It is suggested that the contribution of mGlu receptors to the amphetamine-induced motor response may result mainly from their functional, either direct or indirect, interactions with D1-like receptors in the NAcc.